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Jenkinson JW. Experimental Embryology. (1909) Claredon Press, Oxford.

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Experimental Embryology

Experimental Embryoijogy

By

J. W. Jenkinson. M.A.. D.Sc.

Lecturer in Embryology in the University of Oxford

(1909)

Preface

For the biologist there are, I conceive, in the main two prohlems. One is to give an account of those activities or functions by means of which an organism maintains its specific form in an environment. The other is to ﬁnd the causes which determine the production of that form, whether in the race or in the individual. The solution ol":tlie first of these problems is the business of physiology, in the usual sense of the term. The second falls to morphology.

It is with the origin of form that we are here concerned, and in particular with its origin in the individual. The endeavour to discover by experiment the causes of this process——as distinct from the mere description of the process its(-lf—is a comparatively new branch of biological science, for Experimental Embryology, or, as some prefer to call it, the Mechanics of Development (E'n(w.'i¢'/clungsiner/zcmik), or the Physiology of Dexelopment, really dates from Roux's production of a half-embryo from a. half-blaatomere, and the consequent formulation of the ‘ Mosaik-Theorie’ of self(llffel'0ni«ltttl0l1. That hypothesis has been the focus of much fruitful criticism and controversy, the experiment has been followed by many others of the same kind, and the present volume is an attempt to sketch the progress of these researches and speculations on the nature and essence of differentiation, as well as of those which deal with growth, cell-division, and the external conditions of development.

In writing this review I have had the very great advantage of an excellent model in the textbook of Korsehelt and Heider (Lehrbuch cler fucrgleichemleat Entwio/cluugsgeschiclzte (Zer 1ve'rbelZo.~e'n T/u'c7'e, Allgemeiner Theil, Jena, 1902). I have iv PREFACE

indeed followed the general arrangement adopted by these authors fairly closely except in one respect. I believe so strongly that the processes of growth and cell-division, though they always (in the Metazoa) accompany, are yet distinct from, differentiation, that I have felt justiﬁed in treating them in a chapter apart from the other internal factors of development. The external factors—whether of growth, celhdivision, or differentiation-—-are discussed in Chapter III, and the ground is thus cleared for a consideration of the real problem ——the differentiation of speciﬁc form.

The last chapter is devoted to the theories, scientific and philosophical, of Hans Driesch. I sincerely hope that Herr Driesch will allow my great admiration for the former to atone in some measure for my inability to accept the tenets of nee-vitalism.

It is a. very great pleasure to 1110 tolacknowledge my indebtedness to the Delegates and Secretaries of the Clarendon Press, and in particular to Professor Osler, for undertaking the publication of this book, as well as for the pains which have been expended in its preparation. Dr. Osler also took the trouble to read through the whole of the manuscript, and Mr. G. ‘V. Smith and Dr. Haldane have been kind enough to look through certain chapters.

To Dr. ltamsden I am under great obligations for his assistance in that part of Chapter 1], Section 1, in which surface-tensions are discussed; to Dr. Vernon for calling my attention to Roberts’s work on Anthropometry, and to Mr. Grosvenor for the information embodied in the foot-note on p. 89. Mr. A. D. Lindsay has given me invaluable assistance in those sections of Chapter V which deal with the philosophy of Kant, while, for Aristotle, I was fortunately able to attend Professor Bywatei-’s lectures on the De Am'ma.

I can hardly express the debt I owe to Mr. J. A. Smith for

much friendly counsel and criticism, although he is, of course, 1’REI<‘A(,‘E v

in no way responsible for the philosophical speculations in which I have ventured to indulge.

The illustrations are largely borrowed from Korschelt and Heider’s work, and I must thank Herr Gustav Fischer, of Jena, for his readiness in supplying the blocks. Others are from the original publications‘, and I am obliged to the proprietors for permission to make use of them. A few are my own.

In the appendices will he found an account of some recent work on the relation between the symmetry of the egg and that of the embryo in the Frog, and on the part played by the nucleus in ditt'c1-entiation.

’ Pru('e('zIin_q.s- of the Bo.»-ton S'0c1'eI_:/ of Natm'ul 1‘II'.\'l0)'_I/, the Journal of E.rpm-[mental Zoology (VVillia1ns 8; Wilkins, Baltimore), the Anm'ir(rn Journal of I‘hysz'ulo_'/_I/ (Ginn & C0., Boston), ZeIIrn~Sfu(Iim (Fischer, Jena), l’erhamIlmI_r/en 410;" A/mlumis-1-hm G(‘.s'¢'”N(‘7I((fl (Fischer, Jena), Er;/cbnisse fiber din Ii'on.m'tzm'ou dcr cIu'onmta'scIzm Kernsubslmz: (Fischer, Jena), .[r¢-kin fiir mik)'osk0])i.s¢*7¢1: .»lm¢tomi(' (Cohen, Bonn), Archizv ff/"r Entwiclcluuysnwvlzanik (Engelinunn, Leipzig), and the Popular Science .llontM3/ (Appleton & L‘o., New York).

1. The modern form of the prefurmationist doctrine 2. Amphibia 3. Pisces 4. Amphioxus 5. Coe-lenterata 6. Ecliinodcrmata 7. Nemertinen . . . . . . . . 204 8. (.‘tenopho1':i . . . . . . . . 208 9. Chaetopoda and Mollusca . . . . . . 213 10. Ascidia . . . . . . . . . 229 11. General consiileratious and conclusions . . . 240 12. The part. played by the spernmtozoon in the determination of egg-.<ztructm'e . . . . . 247 13. The part played by the nucleus in iliﬂ'e1'enti;iti0n . . 251 (2) The actions of the parts of the developing oiganism on one another 271 Cll.\l"l‘El{ V Dmi~:scu‘s '1‘1n:on11a.x' or l)l'2\.'l-2l.()l‘M.l-INT. (}J«:.\'r;RAI. ll:-:i«‘m:c~ noxs AND (,.‘0Nc1.1.'s1o.\'s . . . . . . 27!)

APPENDIX A On the .’~y)1)l)lL'l2l'y of the egg, the symmetly of scglnentation, and the symmetry of the embryo in the Frog

APPENDIX B

On the part played by the nucleus in (lifferenti:L’tion

I.\'m<;x or AUTl{0I{

Ixmzx or SUBJPJCTS

ADDENDA

Chapter I

INTRODUCTORY

THAT living creatures reproduce their kind is a fact which is familiar to us all, but it is the peculiar privilege and province of the embryologist to observe and to reflect upon that marvellous series of changes whereby, out of a germ which is comparatively structureless and unformed, a new organism is developed which is, within the limits of variation, like the parents that gave it birth.

Development is the production of speciﬁc form. 1<‘rom a particular kind of germ only a particular kind of individual will normally arise, though unusual conditions may lead to the formation of an abnormality or monstrosity. Thus, while the germ is the material basis, development is the mechanism of inheritance. The student of heredity seeks to express in terms which shall be as exact as possible, ultimately mathematically exact, the degree of similarity between the offspring on the one hand, and parents and more remote ancestors on the other. The embryologist has under his very eyes the process by which that similarity is brought about, and even when the resemblance shall have been stated with all possible precision, it will still remain for him to give an explanation of those changes whereby the inheritable peculiarities of the species are handed on from one generation to the next.

Used in the widest sense of the word, development includes not merely the formation of a new individual from a single cell, whether fertilized or not, but also the phenomena of budding and regeneration. In a narrower sense, however, the term is restricted to the first of these processes, and a corresponding distinction is made, however artiﬁcially, between Experimental Embryology and Experimental Morphology, when the subject is treated from a physiological point of view.

uzxxmsox 13 2 INTRODUCTORY I

In development two factors are obviously involved. One is growth, or increase of volume, more correctly increase of mass; the other is differentiation, or increase of structure; and, in multicellular organisms, both these factors are accompanied by division of the nucleus and the cell.

Segmentation is the first sign, or almost the first sign, the developing ovum gives of its activity; and this cutting up of the egg-cell into parts, which marks the beginning, is also continued during the later stages of ontogeny, and goes on as long as the life of the organism endures.

Growth is especially characteristic of the embryonic and of the adolescent organism. It occurs at different rates in the different cells, and indeed the growth of a group of cells is in itself often an act of differentiation. Growth may depend upon the absorption of water or the assimilation of other substances, and this may lead simply to an increase in the size of internal cavities, as in the blastula of lrlc-hinoderms or the Mammalian blastocyst; to an increase in the volume of the living protoplasm ; or to the secretion of intracellular or intercellular substances, either organic (for example, the notochordal vacuoles, the matrix of cartilage and bone) or inorganic (the skeletal spicules of Eehinoderm larvae, Sponges, and Coelenterata). This increase of mass is not only conditioned by the presence of food in the form of substances found in the environment, but depends on such external circumstances as temperature, atmospheric and osmotic pressure, and so forth.

But while the embryo is dividing up its material—a material which is already to a certain extent heterogeneous, composed, for example, of protoplasm and deutoplasm or yolk——while it is increasing its mass, it is also undergoing a process of differentia tion; and, as even a superficial acquaintance with embryology.

will inform us, one of the most characteristic features of differentiation is that it occurs in a series of stages which follow upon one another in regular order and with increasing complexity. When segmentation has been accomplished—sometimes, indeed, during segmentation——certain sets of cells, the germ-layers, become separated from one another. Each germlayer contains the material for the formation of a deﬁnite set of I INTRODUCTORY 3

organs——the endoderm of a Vertebrate, for instance, contains the material for the alimentary tract and its derivatives-—gill-slits, lungs, liver, bladder, and the like ; the germ-layers are therefore not ultimate but elementary organs, and elementary organs of the first order. In the next stage these primary organs become subdivided into secondary organs——as the arehenteron of an Eehinoderm becomes portioned into gut and eoelom-sac, or the ectoderm of an Earthworm into epidermis, nervous system, and nephridia——and in subsequent stages these again become successively broken up into organs of the third and fourth orders and so on, until ﬁnally the ultimate organs or tissues are formed, each with special histological characters of its own. This end is, however, not necessarily reached by all the tissues at the same time. Indeed, it is no uncommon thing for certain of them to attain their ﬁnal structure while the others are yet in a rudimentary condition; thus, in some Sponges the scleroblasts begin to secrete spicules in the larval period, nematocysts may be formed in the Planula of the Coelenterates, notochordal tissue is differentiated in the newly hatched tadpole of the Frog; and, speaking generally, larval characters are developed at a very early stage.

To this regular sequence of ontogenetie events Driesch has applied the term ‘rhythm ’, the rhythm of development. The organs of the body are, however, by no means all formed of single tissues~—-bone, epithelium, blood, and the rest——but are compounded, frequently of very many tissues, and this ‘ composition ’, to quote a term of Driesch’s again, is another of the obvious features of organogeny.

VVhile, therefore, in the last resort all diiterentiation is histological, that ﬁnal result, the assumption by the cells of their definitive form, is only achieved after many changes have taken place in the position of the parts relatively to one another while the organs are being compounded, and so its speciﬁc shape conferred upon the whole body.

It is possible to find a few general expressions for the manifold changes that take place in the relative positions of the parts. Several years ago, in 1874, His compared the various layers of the chick embryo to elastic plates and tubes; out of these he

13 2 4 INTRODUCTORY I

suggested that some of the principal organs might be moulded by mere local inequalities of growth——the ventricles of the brain, for instance, the alimentary canal, the heart—-and he further succeeded in imitating the formation of these organs by folding, pinching, and cutting india-rubber tubes and plates in various ways. This analysis, however, deals only with the foldings of ﬂat layers, and must be supplemented by a more exhaustive catalogue of the processes concerned in ontogeny, such as that more recently suggested by Davenport. Davenport resolves the changes in question into the movements of cells or cell aggregates, the latter being linear, superﬁcial, or massive, and within the limits of these categories the phenomena are susceptible of further classiﬁcation. The catalogue proceeds as follows :——

I. THE Movmtaxrs or SINGLE Cams.

1. Migration of nodal thickenings in a network of protoplasm: e. g. the migration of the ‘ cells ’ to the surface of the Arthropod ovum to form a blastoderm, the movements of vitellophags, and yolk-nuclei (Fig. 1).

FIG. 1.—Sections of the egg of Geophilus ferrrlyiuezls showing two stages in the formation of the blastoderiu: bl, blastoderm; dp, yolk pyramids; gr, groups of blastoderni cells on what will be the dorsal side ; L-, nuclei surrounded by masses of protoplasm. (After Sograff, from

Korsehelt and Heider.) I INTRODUCTO RY 5

2. Migration of free amoeboid bodies: e. g. the mesenchyme cells in the Eehinoderm gastrula, the lower layer cells of Elasmobranchs, the blastomeres amongst the yolk-cells in Triclads and Salps.

3. Aggregation of isolated cells.

a. Linear aggregates: e. g. the kidney of Lamellibranchs, the yolk-gland of Turbellaria, capillary blood-vessels.

6. Superﬁcial aggregates: e. g. the blastoderm of Arthropods, the formation of the imaginal gut-epithelium in some Insects.

0. Massive aggregates : e. g. the gemmule of Sponges, the spleen of Vertebrates.

4. Attachment of isolated cells to another body: e. g. the union of muscles to the shell in Mollusca and Arthropoda, of tendon to bone in Vertebrates, the application of skeletal cells to the notochord.

5. Investment and penetration by isolated cells: e. g. the follicle cells between the blastomeres in Tnnicata, the muscles of the gut in va.rious animals, the septa of the corpus luteum, the formative cells of the vitreous body of the Vertebrate eye, the immigration of the nephric cells in the Earthworm.

6. '1‘ransportation of bodies by wandering cells: e. g. of the buds in Doliolidae.

7. Absorption by wandering cells: e. g. phagocytosis in Insect pupae a11d in the ta(lpole’s tail.

8. We may place here. the frequent alterations in the shapes of cells, which do not apparently involve growth: e. g. when {lat cells become columnar.

II. Tm»; Movi~:.\mNTs or (J1~:r.r. A(:eu1«;(:A'rEs. A. Linear Aggregates.

1. Growth in length: e. g. the growth of the roots and stems of plants, of the stolons and hydranths of Hydroids, the outgrowth of nerves, of the necks of unicellular glands, the growth of the blood-vessels from the area vaseulosa into the body of the Chick embryo, of blood-vessels towards a. parasite, the growth of mesoblastie and other germ-bands in Annelids, the back-growth of the Vertebrate segmental duct, and the like. 6 INTRODUCTORY I

2. Splitting.

a. At the end, that is, branching: e.g'. of nerves, bloodvessels, kidney tubules, glands, tentacles.

3. Throughout the length: e.g. the segmental duct of Elasmobranehs, the truncus arteriosus of Mammalia.

3. Anastomoses: e. g. of the dorsal and ventral roots of the spinal nerves, of nerve plexuses, of capillaries, of bile capillaries, of the excretory tubules of Platyhelmia.

4«. Fusion with other org'a11s: e. g-. of a nerve with its endorgan, of the vasa eiferenti-.1. with inesonephrie tubules, of nephriclia with the coelom in Annelida.

B. Superﬁcial aggregates. i. Increase of area. (1.. Growth of a sphere. 1. Equal in all dire(-tions: e.g. the blastula of ]*l(-hinoderms. 2. Unequal.

oz. Unequal in dilferent axes: e.g-. the conversion of a spherical blastula into an ellipsoid Planula in Coelenterata, or into an ellipsoid Sponge larva, or of the spherical into the ellipsoid blastocyst in l\Ia1mnalia.

,8. Unequal at dilferent poles: e. g. the formation of ovoid forms, such as 1’lannlae, the club-shaped glaild of .122/19/ziamzs, the auditory vesicle of Vertebrata.

6. Growth of a plane surface. 1. Equal in all directions: e.;,;'. the growtli of the blastodcrm over the yolk in Sauropsida, or Cephalopoda.

2. Unequal. a. When parts lying in one plane move out of that

plane: c. g. inva,t,;‘i11ations and evaginations of all descriptions.

(Fig. 2).

B. When parts—e. a row of eells——lying in one plane are moved in that plane: e. g. the germ-bands of C/cpsinc, by the growth of the epiblast (Fig. 3).

ii. Alterations of thickness. 1/. Increase: thickenings: e. g. the formation of the central nervous system in Teleostei, the formation of gonads from the I INTRODUCTORY 7

coelomic epithelium, the development of hair follicles, the trophoblast in the Mammalian placenta (Fig. 4).

/I noaaoanaaoonnananaaaa

Fm. 2.—-Three stages of an invagination or evagination. (After Korschelt and Ileider.)

A 0 0 DDDDDDIJDDDDDC 0

1?

7 ‘IR

l§a;: .».i?

I .1

anaﬂlﬁadﬁk

WWI

ﬂ DU UUUUUUUOUUDUQDQU ‘ ' FIG. 3.——Displacement of a row of cells in an epithelium. (After Kor- qQp§§ag!d9“’gQ‘éDO

schelt and Heider.) PI 4 F t _ tlef t, < .——— ours ages 1n 1 orma 1on

uf an epithelial thickening of many layers. (After Korsehelt and Heicler.)

6. Decrease: thinnings: e. g. in the roof of the thalamencephalon a11d medulla, the outer layer of the lens, the trophoblast of the Mammalian blastocyst. 8 INTRODUCTORY I

iii. Interruptions of continuity. a. By the atrophy of part of a, layer: e. g. when the ﬂoor of the archenteron tog-ether with the underlying paraderm dis appears in Amniota. (Fig. 5). X . ° UODCIDDUUDDDUDDU °

3 « <2 vacuum

5 canon» «aaaaaan

Fm. 5.——Three stages in the development of am interruption of von~ tinnity pcrpemliculzmr to the surfztce of am epithelium. (l'erfomtion.) (After Korschelt and Heidcr.)

/1. By the detachment of a part: c. g. of the medullary plate from the eetoderm in Amp//ioxus (Fig-. 6), of the notochord from the roof of the archenteron in Urodela. and I’e/r0myzo7z.

/[ ~ - - iﬂlllllll - WMDDO

FIG 6. «Scheme of the formation of the medullary canal in Amphi0.rIts. (After Korschclt and Heidcr.) I INTRODUCTORY 9

iv. Conerescence of layers. a. By their margins: e. g. the edges of the eetoderm over the medullary plate, the edges of the embryonic eetoderm inside

fl -‘ DOD!» 0000050

/2’

Eﬂﬂﬂﬂboaoﬂﬂﬂﬂﬂﬂ

Fm. 7.--—1<‘usion of two cell plates by their nmrgins. (After Korschclt and Heider.)

the serosa of Si/nmczzlzts, the embryonic plate with the trophohlast in some Mammals (Figs. 7, 9).

(2. By their surfa.c-es (Figs. 8, 9, 10) : e. g. when the stomodaeum or proctodaenm open into the gut, when the mcdullary

ﬂ

FIG. 8.— Fusion of two cell plates by their surfaces. (Aft-er Korsehelt and Heider.) folds meet, when the edges of the peritoneal groove close to form the canal of the oviduct in Amphihia and Amniota.

This concrescence is commonly followed by a communication of the cavities on opposite sides of the adherent layers, as when the stomodaeum opens into the gut, or the amnion-folds unite ; but not necessarily, as when the somatopleure fuses with the trophoblast, or the allantois with the somatopleure in Mammalia.

v. Splitting of a. layer into two: e. g. the inner wall of the pineal vesicle in Lacertilia (Fig. 11). 10 INTRODUCTORY I

\'v'-‘cacao ct -L

1.‘1(,<,, s)__.])iug1-;un to i11u_._~tm,tC Flu. 10 —- Scheme of the f0r11mtion of thefbrmationotthefore-gut(stom. the mcdullm-y mum] in .11, Vertebmtc. odueu1n)andit.s' opening into the (.-U101‘ Korschelt and Holder.) mid-gut: er‘, ectoderm ; nu], midgut; wl,stomodaeum. (After Korschclt and Heicler.)

FIG 11.—Th1-ee stages in the development of an interruption of continuity parallel to the surface of an epithelium. (Delanlination) (After K01-schclt and Heider.)

oouoouuoooou I INTRODUCTORY 11

0. Massive aggregates.

i. Changes in volume.

a. Unequal in different axes : c. g. when the spherical larva becomes cylindrical in Dicyemidae.

3. Unequal at diﬂerent points : e. g. the outgrowth of limbbuds of Vertebrates and other forms, of the buds of plants.

ii. Rearrangement of material.

a. Simple rear1‘angement of cells: e.g. in the formation of the concentric corpuscles of the thymus, in the development of kidney tubules in the metanephric. blastema of Amniotn, in the grouping of the cells to form ectoderm, gut and atrium in the Salps.

/1. Development of an internal cavity: e.g. segmentation cavities, lumina of ducts and blood-vessels, of the coelom and many generative organs.

0. Dispersion ol’ the elements of an aggregate: e. g. in gcmmule formation in certain Sponges, in unipolar immigration in some Sponges and some (‘oelcntcrates, in the liberation of the germ—celis.

iii. Division of masses.

(1. By constriction : c. g. the segmentation of the mesodcrm and neural crest.

/1. By splitting: e. the nervous system from the ectodcrm in Teleostei and many Invertebrates, the notochord from the roof of the {l.1'('l1(3lltCl'011.

iv. Fusion of masses : e. g. of originally separate nerve ganglia (Vertebrates, Arthropods, Annelids), of myotomes, of somites in Artln'opods.

v. Attachment of one mass to another: e. g. of selerotome to notoehord.

It will be seen that this ;'¢7.s-/rzzr/‘"3 17 1C principal kinds of movement executed by the dev 11111:) ‘arts extends Ilis’s principle of the local inequality of growth from ﬂat layers to linear and massive aggregates and at the same time includes the movements of isolated cells. Davenport, however, is not content merely to give a simple classiﬁcation ol the phenomena; he goes further, and endeavours to express them in terms of responses to stimuli, an idea due in the first instance to Ilerbst. 12 INTRODUCTORY I

Thus he suggests that the migrations of vitellophags and mesenchyme cells, the thickenings, thinnings, and perforations of ﬂat layers, the rearrangements of cells in a massive aggregate, their dispersion, the constriction, and splitting and fusion may be regarded as tactic responses, the growth in various ways of linear aggregates, the eoncrescence of layers and masses as so many tropic responses to stimuli which may be positive or negative and exerted by other organs or by agents in the world outside.

Now it is clear that the analyses both of His and Davenport aim at something more than a mere description of ontogenctic events, for a serious attempt is here made to give a causal, if you will a mechanical, explanation of those events, and the subject thereby raised at once from the level of mere morphology or morphography to a loftier, aetiological point of view.

There are, indeed, two methods by which embryology, like any other branch of zoology, may be investigated. One is purely descriptive, anatomical, morphological. By this method, truly, great results have been achieved. The life—l1i_stories of members of all the most important groups of the animal kingdom have been worked out, and the science of Comparative Embryology has been built up. Nor has an explanation of the process been lacking. For ontogeny is, the fundamental Biogenetic Law assures us, a recapitulation of and therefore explicable in terms of phylogeny; and since on this principle the individual repeats in its development the ancestry of its race, embryology affords a means of tracing out the relationships of the organism and establishing the homologies of its parts.

Unfortunately a more intimate acquaintance with the facts has made it abundantly clear that development is no mere repetition of the aneert ‘é~\(_‘ "e1,tliat the organism has manifold ways of attaining its sin k i" ‘.',(\at those resemblances in early stages which were held to .,.:sm. bthe most triumphant vindication of the Biogenetic Law bear no constant relation to the similarities of adult organization, that the attempt to ﬁnd in development an absolute criterion of homology is vain.

The facts thus remain unexplained, as in truth it was only to be supposed that they would. A method, however comparative, which relies on mere observation, and is content to wait for 1 INTRODUCTORY 13

Nature's own experiments, cannot hope to arrive at sound inductions, or to establish general laws of causation.

There is, however, another \vay. Development, the production of form, may be regarded as one of the activities, one of the functions of the organism, to be investigated. like any other function by the ordinary physiological method of experiment; and the ideal of the experimental or physiological embryologist is to give a complete causal account, whether the causes are external or internal, of each stage, and so of the whole series of ontogenctie changes, his weapon, to borrow Roux’s splendid phrase, ‘die Geistesanatomie, das analytische eausale Denken.’ 1

This effort is, of course, no modern one. Speculation into the nature and essence of development begins, indeed, with the Greeks, and theories of fertilization and development are to be found in the writings of Aristotle? In fertilization the male element, which, according to Aristotle, provides the formal and efficient causes in providing the necessary perceptive soul, acts upon the mere matter, endowed only with a nutritive soul, which is given by the female, in the same sort of way, to use his own illustration, as rennet coagulates milk. In the germ thus formed the parts of the embryo, which can only be said to pre-exist potentially, arise not simultaneously but in gradual succession, first the heart, then the blood, the veins from the heart and the various organs about the veins by a process of condensation and coagulation, the anterior parts of the body being built up first.

This Aristotelian doctrine appears to have persisted through the Middle Ages ; it reappears in the seventeenth century in the pages of Hieronymus Fabricius ab Aquapendentc and his pupil VVilliam Harvey in essentially the same form, although both authors diifer from Aristotle in certain matters of observational detail. Thus l*‘abricius3 states that ‘ ope generatricis facultatis pulli partes, quae prius non erant, produci atque ita ovum in pulli corpus migrate’, while Harvey“ gives to development as thus conceived of the name of ‘Epigenesin sivc partium superadditionem’, though he believes that in some cases (Insects) the

1 Roux, 1885. __ '2 Aristotle, De Gem, i. 20. 18; ii. 4. 43; 5. 2, 3, 10; 6. DeAM., 11.4. 2, 15 ; 5; 6. 3 Fabricius, 1. c., p. 22. ‘ Harvey, l. c., Ex. 44. 14 INTRODUCTORY I

process is one of ‘metamorphosis’ or the simultaneous origin of all parts. In generation properly so called, however——in the development of the Chick, for exa1nple——tl1e process ‘a parte aliqua, tanquam ab origine, incipit; eiusque ope reliqua membra adseiscuntur: atque haec per epigenesin ﬁeri dieimus: sensim nempe, partem post partem’, and this ‘pars prima genitalis’ Harvey held, in opposition to Aristotle, to be the blood.‘

But in spite of the exact observation and brilliant exposition of his followers, the teaching of Aristotle was destined to be overshadowed and eclipsed, tempoiarily at least, by a new l1ypo— thesis which, appearing ﬁrst towards the end of the seventeenth century, swept the schools and universities, and dominated biolog-ieal speculation for a hundred years.

This was the theory of Evolution or Preformation. According to it the future animal or plant is already present in miniature in the. germ with all its parts complete, invisible or hardly visible it may be, but still there, and not merely ‘ potenti-ft’; and in development there is no such thing as ‘generation’, but only growth, whereby that which was before impalpable and invisible becomes tangible and manifest to our eyes. A further and logical outcome of the hypothesis was the doctrine of ‘emboitemcnt’, enthusiastically described by Bonnet as ‘une (les plus belles vietoires que l’entendement pur ait remporté sur les sens ’.3 The organism present already in the germ, with all its parts complete, possesses of necessity the germs of the next generation, and so on in indefinite though not in inﬁnite regress, for as Bonnet is careful to tell us, ‘ Il ne faut pas supposer un emboitcment in l’inﬁni, ce qui scroit absurde. La divisibilité de la matiére 51 l’inﬁni par laquelle on prétendroit soutenir eet emboitement est une vérité géométrique et une erreur physiqne.’3 Swammerdam solved the diﬂiculty in another way. All the’ germs of the human race must have been present. in the bodies of our ﬁrst parents, and ‘exhaustis his ovis humani generis ﬁnem adesse ’.“

The theory became widely held. First put forward by Marcello

‘ Harvey, l. e., EX. 50.

2 Bonnet, Cont. de la nm‘., 71"“ partie, c. ix ; (1a'uLv'cs, vol. iv, p. 270. 3 Bonnet, Consid. sur les rorps 0131., c. viii; (lﬁuvres, vol. iii, 1). 74.

‘ Swammerdam, 1679, pp. 21, 22. I INTRODUCTORY 15

Malpighi in the memoir, ‘ De formatione pulli in ovo,’ which he presented to the Royal Society of London in 1673, it was not only adopted by biologists of prestige, by Swammerdam, Haller, who in his early days had been an advocate of Epigenesis, dc Buffon, and Bonnet, but secured the adherence of philosophers of such eminence as Malebranche and Leibniz.

In some cases it was accepted as a result of observation. Thus Malpighi,‘ in the treatise referred to, asserted that he had himself observed the chick in the unineubated egg, ‘ inelusum foetum animadvertebam, cuius caput cum appensae carinae staminibns patenter emergebat,’ while de Buifonz expresses himself even more categorically. ‘ J ’ai ouvert,’ he says, ‘ une grande quantité d’eeufs a diﬁérens temps, avant et apres Pineubation, et je me suis eonvaincu par mes yeux que le poulet existe en entier dans le milieu de la eicatricule a11 moment qu’il sorte (lu corps de la poule.’ To others, however, it was rather a matter of theoretical necessity. Ilaller explains his conversion from the contrary opinion by asking‘ the very pertinent question, ‘(‘ur vis ea essentialis quae sit unica tam divorsas in animali partes semper eodem loco, semper ad eun(lem arehetypum struit ; si materies inorganica mutabilis et ad omnem ﬁguram rccipiendam apta est ? Cur absque ullo errore ex gallinac mista materie ea vis semper pullum, ex pavo11e pavonem fabricatur? ’

‘Nil nisi vis dilatans et progrediens reeipitur. Ab ea nihil sperarem nisi vasorum rete tamdiu continue amplius futurum quamdiu vis expandens resistentiae superandae par est. Cur loeo eius retis cor, eaput, eerebrum, ren struuntur? Cur in singulo animali suus ordo partium? Ad eas quaestiones nulla. datur responsio,’ a charge which is, of course, perfectly just.“

Bonnet’s argument is different. The heart of the chick, he points out, is already present in the egg‘; and since anatomy teaches that all the parts of an animal ‘doivent avoir toujours coexisté ensemble’, preformation follows as a matter of course.“

The belief in preformation continued paramount till towards the end of the eighteenth ceutur , nor was it till the publication

‘ Malpighi, 1. e., p. 4. 2 do Buffon, 1. c., p. 351. 3 Haller, 1778, VIII. i. 29, p. 121. _ ‘ Bonnet, Cont. de la nat., 7""? partie, c. ix; (lc'm‘)‘ﬂs, vol. 1v, 1). 261. 16 INTRODUCTORY I

in 1774 of Caspar Friedrich Wolff's T/Ieoria Gczze/'atz'om'.9 that the evolutionists were aroused from their dogmatic slumbers. Putting speculation on one side, Wolff returned to the method of Harvey, Fabricius, and may we not say also of Aristotle, the method of exact observation. He demdnstrated the presence in the unincubated egg not of a complete organism, but of ‘globules’; ‘ partes enim eonstitutivae, ex quibus omnes corporis animalis partes in primis initiis componuntur, sunt globuli," and described the epigenetic formation of the heart and blood-vessels, the central nervous system, the limbs and the ‘ VVolffian’ bodies from these primary elements.

Development thus consists of the gradual production and organization of parts; ‘embryonis partes sensim produci, mea observata suadent,’ 2 and again, ‘ suppeditari prius partem, deinde cam organisari iutelligitur ’.3

The ground was thus taken from beneath the feet of the preformationists, and Epigenesis restored to its former place of honour as the fundamental expression of developmental fact.

Tacitly accepted by all the great embryologists of the nineteenth century—~Pander, von Baer, Reichert, Bischoif, Remak, Kolliker, Kowalewsky, IIaeckel——the epigenetie idea continued to control the progress of research. These were men who set themselves to describe the sequence of changes that the embryo passes through with all possible accuracy, and over as wide a range as might be of animal form. 'l‘hey made Comparative Embryology. On the facts that they discovered new light was shed by the doctrine of descent with m0(lilicati0n, or evolution in the wider sense of the word. Von Baer had pointed out that in any group of animals the embryos were more like one another than were the adult organisms, and this now became easily translated by Haeekel into the idea that the form which is in every g-roup——ultimately in all groups—thc common startingpoint of individual development is representative of the common ancestor of the race. Ontogeny was thus not merely expressed but explained in phylogenetic terms.

Now that, as we have already seen, the proposed explanation

‘ Wolf, 1. c., Praemonenda, lxviii. 2 Id. ib. lxxiii. 3 Id. ib., De Gen. An., § 240. I INTRODUCTORY 1 7

has very‘ largely broken down, Epigenesis, taken by itself, remains, not a theory in terms of cause and effect, but a mere description of what occurs, and it is the crying need for such a theory that has given birth to modern experimental embryology.

The new era opens with the publication by Wilhelm His in 1874, just a hundred years after the appearance of the '1’/eeoria G'e2wratiom's, of a remarkable series of essays entitled Uusere K5f])87f07'7)l marl Jae 1;/1_'/siologisc/10 Problem 2'/zrer Eutste/nm_q. In these essays His, who was already in revolt against the ‘ Biogenctie Law’, not only sought to give a mechanical explanation of differentiation, but also laid down his famous ‘Prinzip der organbildenden Keimbezirke.’ According to this principle of ‘germinal localization’, every spot in the blastoderm corresponds to some future organ : ‘ (las Material zur Anlage ist schon in der ebencn Keimseheibe vorhanden, abcr morphologisch nieht abgcglie(lert, und somit als solchesnicht ohne Weiteres erkennbar.’ 1 Conversely, every organ is represented by some region in the blastoderm, and ‘ wenn wir consequent sein wollen’ in the fertilized, or even unfertilized, egg. In other words, although the parts of the embryo cannot be said to be preformed in the germ, the materials for those parts are already there, prelocalized, arranged roughly, at least, as the parts themselves will be later on. In this material unequal growth produces the form of the parts, and so of the whole body. Whether there is a strict causal connexion between each material rudiment and the organ which arises from it, whet-her these rudiments could be interchanged without prejudice to the normality of subsequent development, is a question which is not touched upon by His. It was reserved for another anatomist, Wilhelm Roux, to raise what in IIis’s hands had been merely a principle to the rank of a theory, the ‘ Mosaiktheoric ’, or theory of self-diiferentiation.

For Roux, no doubt, the ‘Mosaik-theoric’ was in part the outcome of the theoretical necessity of explaining the speciﬁc nature of development ; but it rests also upon a basis of observation and experiment. The coincidence in a majority of Frogs’ eggs of the first furrow with the sagittal plane, the production of local defects in the embryo by local injuries to the egg, the

1 His, 1. c., §ii.

Jicnxmson C 18 INTRODUCTORY I

occurrence of certain natural monsters (Hemit/leria anteriora, for example) in which one half of the body is normally developed, the other entirely suppressed, and the experimental demonstration of the formation of a half-embryo from one of the first two blastomercs of the Frog’ s egg when its fellow had been killed, all led Roux to regard the development of the whole and of each part as essentially a process of self-differentiation, a process, that is to say, of which the causes reside wholly within the fertilized ovum and within each part as it is formed, though allowance was made for the possible formative inﬂuence of the parts on one another in later stages. External conditions, though they may be necessary in the same sense as they are generally necessary to the maintena.nce of life, are yet of no importance for diiferentiation regarded as a speciﬁc activity of the organism.

In the meantime, an experiment of Pﬂiiger’s had apparently shown that, however obviously each part of the egg-cell might be related to the production of a particular organ, the relation was no necessary one, but that, on the contrary, the parts were all equivalent and the ovum ‘isotropic’. Pfliigcr demonstrated that in a Frog-’s egg which had been prevented from assuming its normal position with the axis vertical, the planes of the segmentation furrows bore no constant relation to the original egg-axis, that is to say to the structure of the egg, though they exhibited the same relation to the vertical as when de— veloping in the normal position. Further, in such forcibly upturned eggs the plane which included the original egg-axis and the present vertical axis became the median plane of the embryo, whose axes were disposed with regard to the vertical as in normal cases. Any part of the egg, therefore, might give rise to any part of the embryo, according to the extent to which and the direction in which the egg-axis had been diverted out of its original vertical position, and hence the egg-substance was ‘isotropic’ ; the planes of segmentation and the embryonic axes being determined by gravity. In fact, Pﬂiiger went so far as to say that an egg only becomes what it does become because it is always placed under the same external conditions.

Nor was this conception of the isotropy of the ovum invalidated by Born’s proof that in these eggs there is a redistribution I INTRODUCTORY 19

of yolk and protoplasm owing to the sinking of the former to the lower, the rising of the latter to the upper side of the egg. For though the egg thus comes to acquire a secondary structure about an axis which is vertical, still the arrangement of the parts of the supposed rudiments of the organs must have been disturbed. Yet from such ova normal tadpoles are developed.

It became necessary, therefore, to locate the self-diﬁerentiating substance, the ‘idioplasma’, mainly, at any rate, in the nucleus; and this idioplasma was imagined as composed of dissimilar determinant units, each representative of some part or character of the organism and arranged according to a plan or architecture which corresponds in some way with the architecture in the embryo of the organs represented. All that is necessary, then, and all that happens, at least in the early stages of development, is the gradual sundering of these units from one another by successive qualitative divisions of the nucleus and their distribution to the cytoplasm, where each determines the assumption by the cell to which it is allocated of that character which it represents.

Roux’s ‘Mosaik—tl1eoric’ and VVeismann’s very similar but more elaborate hypothesis of the constitution and behaviour of the germ-plasm both frankly involve the belief that every separately inheritable quality of the body has its own representative in the germ, with the difference, however, that this preformation, extended by Weismann to the adult characters, is limited by Roux to those of the embryo. The renewed inquiry into the nature and essence of development has thus simply resulted in the resuscitation of the eighteenth-century doctrine of evolution, though in a far more subtle form. Once again we ﬁnd ourselves face to face with the old alternative, Preformation or Epigenesis ,- and it is to the desire of solving this problem that a very considerable proportion of modern experimental research is attributable. Though much of this has been directed against and been destructive of the ‘ Mosaik-theorie’, which as far as the nucleus is concerned has now been abandoned by Roux himself} renewed investigation has proved the existence in many cases of definite and necessary organ-forming substances in the cytoplasm, while the necessity for ﬁnding a causal explanation

’ Roux, 1903. 20 INTRODUCTORY I

of what is obviously in some sense a predetermined process, without presupposing the preformation in the germ of morphological units representing every possible iuheritable character, has issued in IIerbst’s and Driesclfs conception of the events of ontogeny as so many responses to stimuli exerted by the developing parts on one another. At the same time the need for inquiry into the external conditions and the part they may play in growth and differentiation has not been forgotten.

Thus though it would be vain to pretend that the ideal of a complete causal explanation has yet been realized, still some material has been gathered for an answer to each of the two main questions: what are the internal, what the external conditions that determine the course of development? These questions we shall discuss in the following pages. It will then only remain to inquire whether a causal explanation—-in the accepted sense of the phrase—a mechanics of ontogeny which resolves the single occurrences ﬁrst into general physiological laws, and these in the last resort into the generalizations of physics and chemistry, can ever afford a theory which may be said to be complete either from a scientific or from a philosophical point of view. Should the mechanical explanation prove to be scientiﬁcally insufficient, it may be necessary, with Driesch and the neo-vitalists, to invoke a consciousness of the end to be realized to guide and govern the merely material elements; but even were this not so it would still be incumbent upon us to consider whether the end itselt'—not the consciousness of it—is not the final, and yet none the less the principal determining cause of the whole process.

LITERATURE

ARISTOTLE. Dc gcneratione animalium, ed. Bekkcr, Oxford, 1837. De partibus animaliuin, ed. Bekker, Oxford, 1837. De anima, ed. Trendelenburg, Berlin, 1877.

C. BONNET. (Euvres, Neuchatel, 1781. Contemplation do In. nature, vii“‘° partie; Considerations sur les corps organises; Mémoire sur les gerlnes; Palingénésie philosophique.

G. BORN. Ueber den Einﬂuss der Schwere ant‘ das Froschei, Arch. mi/rr. Anaf. xxiv, 1885. I INTRODUCTORY 2 1

L. DE I1UFI~‘()N. Histoirc nuturelle, génc’-mle et. pzl1'ti('111ii:1‘e, vol. ii, 1’zu'is, 1749.

C. B. DAV!-:N1*oR'1‘. Studies‘ in morphogenesis: iv. A prelimimtry catalogue of the processes concerned in ontogeny, Bull. 1Iarmr«l Mus. xxvii, 1896.

H. DRIESOII. Anzrlytische Theorie der orgzmischen Entwicklung, Leipzig, 1894.

IL FABRICIUS AB AQUAPENDENTE. De formatione ovi et pulli. Opera. omnia, Lipsiae, 1687.

A. HALLER. Hernmnni Boerhanve pmelectiones ncndemicae: edidit at notas addidit Albertus Haller, v, Gottingne, 1744. Prinme lineae physiologiae, Gottingae, 1747. Elementa. physiologiae corporis humani, Lausannae, 1778.

W. HARVEY. Exercitationes dc genemtione :mima.1ium, London, 1651.

C. Ilmmsr. Ueber die Bedeutung dcr Reizphysiologie fiir die emisale A11f'f'zLss11ng' V011 Vorga"mgeu in der thierisclien Ontogenese, Biol. (‘mil-albl. xiv, xv, 1894, 1895.

\V. HIS. Unsere Korperforin nnd das physiologisclle Problem ihrer Entstehung, Leipzig, 1874.

G. W. LEIBNIZ. (Euvres philosophiques, ed. by 1’. Janet, Paris, 1900. Systeine nouvenu de 1:1. natlire, 1695. Principes de ]:1. nature at de 1:1. gr-free, 1714. Essnis do Théodicée. Préfmze, 1710.

N. MALEBRANCHE. Recherche dc la. vérité, Si1non’s edition, Paris, 1846.

M. MALPIGIII. De formatione pulli in ovo, London, 1673.

E. PFLUGER. Ueber den Einﬂuss der Schwerkmft uuf die Theilung der Zellen und auf die Entwieklung dos Embryo, Pﬂc7,«/m".s- An-h. xxxi, xxxii, 1883.

W. ROUX. Ein1eit.11n;;' zu den ‘ Bcitrfigen zur Entwicklungsmechunik des Embryo ’, Zeitsc-hr. Biol. xxi, 1885 ; also Ge.»-. Abh. 13, Leipzig, 1895.

W. Roux. Ueber die kiinstliehe Hervorbringung ‘ha.1ber’ Embryonen, V4'rrIum‘s Arch., 1888 ; also Gas-. Abh. 22.

W. Roux. Ueber Mosuikarbeit mid neuere Entwieklungslnypothesen, Anat. Ilrjffe, 1893 ; also G'(‘.s'. Abh. 27.

VV. Roux. Einleitung, Am-I1. Eut. Jllwclz. i, 1895.

VV. ROUX. Fiir unser 1’rogramn1 und seine Verwirkliehung, Arrlz. Ent. Mech. v, 1897.

VV. RUUX. Ueber die Ursuchen der Bestimmun,g' der H:lll])1.l'iC]|tnngen des Elnbryo im Frosclici, Anat. Anz. xxiii, 1903.

\V. ROUX. Vortri'Lgc und Aufsdtzc ﬁber Entwvicklungsliieelmnik (ler O1'ga.nismen, Leipzig, 1905.

J. SWAMMERDAM. Mimculum na.tura.c sive uteri muliebris fabrica, Lugdunulu Buta.vorun1, 1679. Histoire génémle des Insectes, Utrecht, 1682.

C. F. WOLFF. 'I‘heo1-in generationis, Hnlle a. d. Szmle, 1774. CHAPTER II

CELL-DIVISION AND G ROWTII

1. CELL-DIVISION

IN a future chapter we shall see that there is no necessary connexion between segmentation and differentiation. Nevertheless, since ccl1~division is the first sign, or almost the first sign, that a developing organism gives of its activity; since, moreover, cell-division accompanies the later processes of growth and differentiation, we may briefly discuss what is known of those factors which determine the direction of division in general, and in particular the pattern of segmentation.

We shall first presume that segmenting ova may be grouped under several distinct types, as follows :—

1. Tim radial I_1//2e. Here the first division is meridional, the second meridional and at right angles to the first, the third equatorial—or more often latitudinal—-and at right angles to both the preceding, the fourth meridional and at forty-ﬁve degrees to the first two, the fifth latitudinal. What is characteristic above all of this type is, first, that four surfaces of contact between cells meet in one line; for example, the four surfaees between the first four blastomeres meet in the egg-axis, while each pair of animal cells lies exactly over each pair of vegetative cells after the third division; and secondly, the blastomeres are radially arranged about the axis. This type has been observed in Sponges (Syconl (Sehulze)), in Coelenterates, in Crinoids, Holothurians, and Echinoids (Fig. 12) amongst the Echinoderms, in Eetoproctous Polyzoa, in A111];/ziowz/.e and the

Vertebrates, and in some C1'11stacea—Cc/or/zilzts ((:‘rrobben),’

Luc-g'f'er (Brooks), Cyclops (Hiickcr), L’/‘awe/u'p11.v (Braucr) and some Cirrhipedes. Certain of these cases present special

peculiarities. In Echinoids micromeres are formed at the vegetative pole

by the division of the fourth phase (Fig. 12,

' In 831001: the third cleavage is meridional, the fourth latitudinal. FIG. 12.-Normal development of the sea-urcliin SI;'ong3/Ionenlrolus ”l’i4Il(.\‘. (After Boveri, 1901.)

The animal pole is uppermost in all cases, and in the first two figures the jelly with the canal (micropyle) is shown.

a, primary oocytc, the pigment is uniformly peripheral.

b, ovum after extrusion of polar bodies. The pigment now forms a subequatorial band. The nucleus is ex-axial.

c, d, ﬁrst division (meridional).

e, 8 cells, the pigment almost wholly in the vegetative blastomeres.

f, formation of mesomeres (animal cells) by meridional division: the vegetative cells have divided into macromeres and micromeres.

_r/, blastula. h, mesenehyme hlastula.

1', j, k, imagination of the pigmented cells to form the archenteron of the gastrula. In j the primary mesenchyme is separated into two groups, in each of which, in Ir, a spicule has been secreted. In is the secondary, pigmented mesenchyme is being budded off from the inner end of the archenteron. 24 CELL-DIVISION AND GROVVTH II. I

In Asteroids and Ophiuroids the division is at first tetrahedral, and to be classed, therefore, with those of the following type; after the second furrow, however, the blastomeres are rearranged, and division theneeforward is radial.

In Vertebiata segmentation is altered in megaleeithal eggs by the amount of yolk present. It becomes meroblastic; still the radial type is preserved, though the sequence of the furrows is often altered, the third, for instance, being frequently meridional, and the fourth latitudinal. Amongst the Ascidians Pg/rosoma has a large-yolked, telolecithal, and radially segmenting egg. In the Placental Mammals the first two (livisions may conform to this type; but segmentation soon becomes irregular. The accumulation of yolk in the Arthropod egg has resulted in a totally different type of meroblastic segmentation. The yolk is here uniformly distributed about the central protoplasm. In the latter is placed the segmentation nucleus, and this central mass divides into a number of cells, which subsequently migrate to the surface and form a blastoderm ; the egg is then centrolecithal (Fig. 1). The stages of the development of this modiﬁcation may be seen in the Crustaeea. In certain forms-——thosc alluded to above (with the exception of the (‘irrhipedes)—— division is holoblastic and radial. In (fam/mru.\', Branc/zipux (Brauer), Pellogastcr (Smith) segmentation is at_first total, but the inner yolk-containing ends of the cells subsequently fuse. In C’1'a7z.r/‘on, ]l[0i/ca, I)aj)/mella, ])a[//erzia, Ore/wstia segmentation is superﬁcial. In Isopods and in Deeapods segmentation is internal. In all cases the result in the end is the same, a peripheral blastoderm, a central yolk. But the blastoderm is not always, though it is often, formed simultaneously over the whole surface. There are cases in which it appears first on the ventral side, and by what may be described as a still more precocious formation of the blastoderm, segmentation may begin at this, the future ventral, point, as in Jlfysis and 0m'scu.9. In these cases the egg is teloleeithal.

In the Insects, Arachnids, Myriapods, and Perz';)atus nomezealandiae, the segmentation is meroblastic and the egg comes to be centrolceithal. In Peripatzw capevms and in some other species it would appear that the yolk has been secondarily lost. II. I CELL-DIVISION 25

2. The‘ second type is the so-called spiral form of cleavage (Fig. 13). This is especially characteristic of the eggs of Polyclads, Nemert-ines, Molluscs (except Cephalopods), Annelids, and Sipunculoids (P//((800/(I-_\'0I1l(I). The peculiarity of this mode of division is that after the ’r'our-celled stage the blastomeres usually known as the 1nacromeres—give elf ‘quartettes’ of micromeres towards the animal pole, the first quartette being given off’ dexiotropically (except in cases oi.’ reversed cleavage),

c B

ZD

FIG. 13.— Diagram of a ‘ spi1'a‘-Jy ’ segmenting egg in the 16-cell stage. 2 A-2 D macromeres; 2 «:2 d nueromeres of second quartette; ] n 1, 1 n LL Id 1, 1 cl 2 nncronieres of first quartettc.

the second laeotropically, and so on in regular alternation, until four quartcttes have been produced. The cells of each quartette divide meanwhile in conformity with the same law of alternation of direction of cleavage. The direction of division is thus always oblique to the egg-axis, and this ()l’)ll(lﬂll]_Y can be observed in the division of the first two blastomcres, the result of which is that of the two sister cells A and B A is nearer to the animal pole than B, while in the other pair C is nearer that pole than D; A being to the left of B and C to the left of I) (to an observer standing in the axis with his head to the animal pole), the division is laeotropie. The arrangement of cells approaches the tetrahedral, especially when, as occurs very frequently, A and C are united by a cross, or polar, furrow above, B and D by a polar furrow at the vegetative pole, as in JVc=rci.v, Icﬁnoc/zitou, Limaw, Plano)‘/153, Lejii/Ionolus, Dixcocelis, and others. In Univ, however, it is B and D that are in contact at the animal, A and C at the vegetative, pole. In other cases 26 CELL-DIVISION AND GROWTH II. I

(I/yauassu, Capi/cl/a, Umbra!/a, Clmyiirlzz/a, Amp/u'6m'te, A/‘euicolu, for instance) the furrows are parallel, the same two blastomercs, B and D, being in contact at both poles. In Troc/ms both the ‘parallel furrows’ and the ‘ crossed furrows’ conditions are found. A similar disposition is to be observed amongst the mieromeres of the ﬁrst quartette. These mieromeres, also, alternate with the macromeres. Not more than three contact surfaces between blastomeres, therefore, meet in one line.

The eggs of certain Lamellibranchs—-Ykrerio, Cg/clas—i11 which the ‘spiral’ arrangement is obscured by the large size of the D maeromere, and possibly the ova of the Rotifera, are to be referred to this type.

The tetrahedral arrangement of the first four cells is conspicuous in Asteroids and Ophiuroids, where the planes of division of the first two cells are at right angles to one another. Before the next division, however, the cells shift their positions and come to lie in one plane, in which, however, the sister cells are not adjacent, but opposite, to one another.

The eggs of Amp/u'0.mzs sometimes segment spirally (Wilson).

After the completion of the spiral period of division, segmentation beeomes radial and then bilateral.

3. The third type of cleavage is the bilateral. The first two divisions intersect in the axis; the next. may he equatorial, as in Aseidi-ans. In this case the hilaterality becomes evident in the succeeding phase, in which the divisions in two adjacent cells of the animal hemisphere meet the first furrow, while in the other two they meet the second. The bilaterality is marked in the reverse way in the vegetative half of the egg. The egg is thus divided into what will be anterior and posterior, dorsal and ventral, and right and left halves. In future divisions the bilateral symmetry is retained.

The egg of ./Imp/ez'o.avzz.s' may divide in this fashion (Wilson), and this is the normal method, according to Roux, in Ram: esculwz/a.

In the Teleostei and some Ganoids (Lepz'do.w‘ws) the bilaterality becomes evident in the third division, which is parallel to the first, the fourth being parallel to the second division. The egg is in fact iso-bilateral.

The Ctenophorc egg also possesses two planes of symmetry, II. 1 CELL-DIVISION 27

for the third division is meridional and unequal in such a manner that the next stag'e~—eig11t cells—is composed of two opposite pairs of small and two opposite pairs of large cells.

The mesomeres in the sixteen—celled stage of Ecliinoids are bilaterally arranged.

In the Cephalopoda the egg is large-yolked, and segmentation consequently meroblastic. After the ﬁrsttwo meridional (livisions

V‘ I1.’ I 01' 1’ Flo. 14.—-Tln-ee segmentation stages in the blastoderm of S'I—'pi(I o_)_7i 4-inalis; the segmentation is of the bilateral type. I, left; 2-, right; I— V,

first to ﬁfth cleavages. The top sides of the ﬁg111‘es are anterior. (After Vialleton, from Korschelt and Heider.)

the bilateral disposition sets in, for the furrows of the third phase are unequally inclined to the first furrow in two halves—— the future anterior and posterior halves—oE the egg (Fig. 14). The egg of A.s'aa7'2's megalocep/m/a also exhibits a bilateral cleavage, but not on the plan just described. The iirst division is equatorial. Then the animal cell divides meridionally, and, as it will prove, transversely, the vegetative cell latitudinally. 28 CELL-DIVISION AND GROVVTII II. I

Betore the next division the most vegetative cell (P._,) slips round to what will be the posterior side. All four cells are bilaterally arranged about the plane in which they all lie, and this will become the sagittal plane of the embryo. The anterior and posterior ends, and therewith the right and left sides, are likewise now determined. The bilateral symmetry is preserved in future divisions, at least in the vegetative hemisphere ; in the animal part of the egg the blastomeres of the left side become t.ilted forwards, those of the right side backwards (Fig. 155, p. 255).

4. In the Triclad Turbellarians, in Trematoda and Cestoda, segmentation is irregular, the blastomeres separate from one another and lie amongst the yolk-cells. The same phenomenon may be witnessed in the Salps, and the separation and subsequent reunion of the blastomeres has also been described in Coelenterates and in Asteroids.

Although these types of segmentation are distinct enough from one another, intermediate conditions are readily found. The radial easily passes into the spiral type for example, for in many eggs of the former kind the ‘cross furrows ’ have been observed at either one or both poles, while the animal blastomercs may be rotated slightly on the vegetative, and so lie not over, but in between, them. The radial symmetry again may become bilateral, as when the meridional furrows of the fourth phase, instead of passing through the animal pole, meet the first or second furrow, symmetrically on either side of one of these divisions; this occurs as a variation in I?ana_/'/mew and (normally (Roux)) in Ifamz excz/Zr’):/rt.

In ()phiuroids and Asteroids the tetrahedral arrangement is lost, and the egg segments radially. In Amp/riuamr: all three types occur.

All three forms may therefore have been derived from one, though what that one was we do not know. In any case, however, one feature is common to them all; in all cases successive divisions are at right angles to one another. This is the law formulated by Sachs long since for the divisions of the cells of plants. It holds good for the segmenting animal ovum, though exceptions may, of course, be found. The alternation of dexiotropic and laeotropic divisions, for instance, in spirally segmenting ova continues for a long period with striking regularity, II. I CELL-DIVISION 29

and it is comparatively rare for a cell to disobey the rule. The rule is, however, no universal law of cell—division. Every embryologist will recollect the continued division of a teloblast in the same direction to form a germ-band, which is such a coilspicuous fact in the development of Molluscs, Annelids, and Arthropods. The four polar nuclei of Insect eggs, lying in one straight line, may also be cited.

The direction of division and the size of the blasfomeres are not, however, the only factors which determine the actual pattern of segmentation. The cells can, and do, shift their positions on one another. This is of common occurrence, and a few examples will suﬂice. The rearrangement of the tetrahedrally disposed cells in Asteroids and Ophiuroids has been ‘noticed already. In many ‘spiral’ ova the micromeres have been observed to rotate on the macromeres, or one quartette to be pushed out of position by the cells of another. In z1.vca;'is the cell P._, slips to one side. 14‘urther, cells change their shape.

Two factors are therefore involved in the production of the pattern of cleavage, the direction of division, and the movements of the cells, and these factors in their turn demand explanation. To these must be added the shape, the size, and the rate of division of the cells.

The two latter depend very largely upon the amount of yolk present in the egg ; yolk-cells are large, the yolk divides slowly, or not at all. This was expressed long sinee by Balfour in the formula, ‘ The Velocity of segn'1enta,tiQn in ‘ FIG. I5.—' Segnientation Oi‘ the

, . Lro0“s one under the influence any Part 01 the ovum 15) roughly of atbeentliijfiigal force (from Korspeaking, proportional to the eon- sc_he1t and Heider, after 0. Hertcentration of the protoplasm there ; ;l(')1(’l'2l'_l111 gzﬁlcolilmllsitvsijl 3131: and the size of the segments is (yolk-syneytiuin) 2 I.-h,bln.stocoel; inversely proportional to the eon- ”” y°lk'““°1°17 "’ y 011"’ centration of the protoplasm.’ ‘ The rule has been vindicated by (). I-[ertwig experimentally. If the egg of the Frog be centri ‘ ("omp. Emb. i. C. 3. 30 CELL-DIVISION AND GROWTH II. I

fugalized with suﬂicient force the yolk is driven still more towards the vegetative pole, while the protoplasm is accumulated in the animal half of the egg. Such eggs segment meroblastically, a cap of cells or blastoderm being formed lying on the surface of a nucleated but undivided yolk. The yolk-nuclei, moreover, are enlarged, as in megalecithal ﬁsh eggs (Fig. 15).

The rule is, of course, only applicable to telolecithal eggs, and for many of these it holds good, notably for Vertebrates. In other classes there are, however, exceptions, which are best known in those \vhose segmentation has been most carefully studied, the ‘ spiral’ eggs of Turbellarians, Annelids, and Molluscs. Large cells, in these ova, often divide more quickly than small ones; the second quartette of micromeres, for instance, is formed before the first quartette divides in 07-epitlula, Uuio, Limaw, Troc/ms, Aplg/sia r/epilans, Discocelis, and the cells of the third quartette before the first products of division have had time to divide again in L[maa', Umbrella, and A/2/ysia limacina. 411 is often formed before the corresponding cells in the other quadrants (in Iﬁzio, for example), but in Crepirlula this is in accordance with the rule, since 4 a, 4 /2, and 40 contain more yolk than 4« cl. In Arem'coZa, though the yolk is uniformly distributed, the cells are still unequal. Other exceptions are to be found in the continued unequal division of teloblasts, in the formation of the micromeres in Echinoids, and in the unequal division of the blastomeres in the third and fourth phases in Ctenophors. According to Ziegler the formation of the micromeres in Ctenophors cannot be due to the presence of yolk, since they are still formed when part of the vegetative hemisphere is removed, as Drieseh and Morgan have also found.

Ziegler indeed puts forward another hypothesis to account for unequal division; he supposes that the centrosomes are heterodynamic. So far there appears to be little evidence in support of this view. It is quite true that in many cases of unequal division the asters—not the centrosomes——-vary in size with the size of the cells. This occurs, for instance, in the division of the first micromeres and of the first somatoblast in Nerei.9, in the formation of the ﬁrst and second quartettes, and in the division of the ﬁrst somatoblast in Uuio, in the division of the cell CD in /laplauc/ma, II. 1 CE LL-DIVISION 31

and in the division of the pole cells of Annelids (Wilson and Vejdovsky). It is doubtful, however, whether it is not the inequality in the cells that is responsible for the inequality of the asters, there being more room in a large cell for the outgrowth of the astral rays. At any rate, ‘there are many cases of unequal clcavage——in polar body formation~——where the asters are of the same size. Until evidence is brought forward of a difference in the size of the centrosomes the hypothesis is no more than a conjecture.

Before quitting this subject we should refer to a rule which Zur Strassen has found to hold good for the rate of segmentation of Ascaris megalocep/zala. The cells do not all divide at the same rate, but in certain groups of cells division is found to occur simultaneously. These cells are related, descended from some one cell, and the more nearly related the cells are, the more nearly together do they divide. Coincidence in time of division depends therefore on the degree of cell-relationship.

The direction of division of the cell depends upon that of the nucleus, since, speaking generally, the division occurs in the equatorial plane of the spindle, or, in other words, the plane of division is at right angles to the direction of elongation of the spindle or separation of the ccntrosomes. The latter again depends on the relation between the nuclear spindle and centresomes on the one hand, and on the other the cytoplasm and its contents, more particularly the yolk. The relation between the (resting) nucleus and the cytoplasm has been expressed by O. Hertwig in the following empirical rule: ‘The nucleus always seeks to place itself in the centre of its sphere of activity.’ The sphere of its activity being not the inert yolk but the cytoplasm, we ﬁnd, in accordance with this rule, that the nucleus places itself in the centre of the egg where the yolk is uniformly distributed (isoleeithal), nearer the animal pole but still in the axis where the yolk is on one side (telolecithal). Examples of the former condition are to be found in Eehinoids (the fertilization nucleus is nearly, but not quite, central) and large-yolked Arthropod ova, of the latter in the eggs of Vertebrates, Molluscs,

and many others. The nucleus, however, may wander from this position, as occurs, 32 CELL-DIVISION AND GROWTH 11.1

for instance, in the egg of Eehinoids after the expulsion of the polar bodies and before fertilization. Apart from such exceptions, due very likely to some temporary alteration in the relations of yolk and cytoplasm, the rule is a reliable one.

The relation between the dividing nucleus, the spindle and centrosomes and the cytoplasm has been stated by O. Hertwig in his second empirical rule ‘that the two poles of the division ﬁgure come to lie in the direction of the greatest protoplasmic mass ', by Pﬂiiger in the l'o1-mula, ‘the dividing nucleus elongatcs in the direction of least resistance.’

The objection that has been urged to this latter expression, that in a fluid the pressure is equal in all directions, may be set aside. For though the cytoplasm is fluid it is an extremely viscid ﬂuid, and the presence of the suspended yolk granules

FIG. 16.——Diagr-am oi the segmentation of the Frog's egg (after 0. llertwig, from Korschclt and Heider). A, first (meridional); B, third (latitudinal) phase of segmentation; p, superﬁcial pigment of animal hemisphere; pr, protoplasm; y, yolk; sp, spindle. must certainly oﬂ'er a greater resistance than the ﬂuid itself, and greater in proportion to their number and size. Pﬂiigcr’s formula, therefore, if not merely a truism, resolves itself into a restatement of Hertwig-’s rule. This rule certainly holds good for a large number of cases, for it explains, for instance, the two ﬁrst meridional divisions of all spherical telolecithal and radially segmenting eggs, the third, latitudinal (in smallyolked eggs 1), and possibly also subsequent meridional and latitudinal divisions (Fig. 16). It will not, however, in the present state of our knowledge, explain the obliquity of the spindles to the egg-axis in spirally dividing ova, nor cases of bilateral division ;

1 ln Sycon the third is meridional, the fourth latitudinal. II. I CELL-DIVISION 33

here, it is evident, other factors must come into play, in the second case probably a bilateral symmetry in the constitution of the cytoplasm. These exceptions may, however, ultimately prove to be special cases of Hertwig’s rule.

A very striking conﬁrmation of the rule is to be found in the division of the egg of Arcane m7_//roreuoxa (Figs. 17, 18). The

- FIG. 17.- Four stages in the fertilization of the egg of A.»-z-(n-is m'_qroveuosa. (After Auerbach, from Korschelt and Heider.)

egg of this worm is ellipsoid. At one end (that turned towards the upper end of the ovary) the polar bodies are extruded, and here the female pronucleus is placed. The spcrmatozoon enters at the

Fm. 18.-—Tl1ree diagrams of the rotation of the fertilization spindle in the egg of Ascaris nigrovenosa. e, s, the directions in which the female and male pronuclei approached one another in A; 1, 2, 3, successive positions of the spindle. (From Korschelt and Heider, after 0. Hertwig.)

muxisson D 34 CELL-DIVISION AND GROWTH II. I

opposite end. The line of union of the two pronuclei therefore lies in the long axis of the egg. Nevertheless the fertilization spindle is not formed in the minor axis of the ellipsoid as one might expect. The two pronuclei rotate together through 90°, the spindle is developed, as usual, in a direction at right angles to their line of union, that is to say the axis of the spindle lies in the major axis of the egg, and the rule is conﬁrmed. There is a similar rotation of the fertilization spindle in the egg of another Nematode, Diployaster (Ziegler), and in the Rotifer Airplane/ma the spindle, at ﬁrst oblique, becomes later coincident with the long axis of the ovum (Jennings).

Curiously enough, this rotation of the pronuclei does not occur in another ellipsoid egg, that of the Rotifer Callidiua. According to Zelinka, the polar body is formed at one end of the long axis, but the fertilization spindle lies in the minor axis, the ﬁrst division includes the major axis, and the law is disobeyed. After the division, however, the cells rotate, and the plane of contact is then, as in Ascaris m'groveuo.m, transverse.

Again, all polar divisions violate the rule, as also does the ﬁrst division of the fertilized egg of Away-is megalocep/zala, and the division of the cells of the germ-bands of Crustacca parallel to their length (Bergh).

On the other hand, Ilertwig has brought forward experimental evidence in support of his generalization. In the eggs of the Frog the directions of some of the divisions were altered by compression between glass plates. The eggs were just allowed to assume their normal position with the axis vertical. They were then placed between glass plates and compressed.

In the first series of experiments the plates were horizontal. In such eggs the ﬁrst furrow was meridional and vertical, the second meridional and vertical and at right angles to the first. So far, therefore, division was as in the normal egg. In the third division the furrows were, however, not latitudinal and horizontal, but nearly vertical, being parallel to the first furrow above, to the second furrow below. The surface of contact, therefore, formed by the furrows of this phase must pass through a meridional position in the interior of the egg. The fourth furrows are latitudinal. Born has repeated the experiment and confirmed this result (Fig. 19). He adds, however, that the furrows of the third division pass II. I CELL-DIVISION 35

towards the vegetative pole below, or may even remain parallel to the first furrow throughout. The fourth furrows, Born says, are parallel to the second. I have myself observed that this division may be either parallel to the second, or latitudinal, even in different

I will Ill l [ill ?

5' 3 3

/ / ' / FIG. 19.—- Segmentation of the Frog's egg under pressure. The compression is in the direction of tlie axis. A. v1ew of the egg between horizontal plates; the animal part IS

islraxled. II, (I, I), first (1), second (2), third (3), and fourth (4) divisions

as seen from the animal pole. (Alter Born, from Korsehelt and lleider )

FIG. 20.—-The first four divisions (I, II, III, IV) in a F10g’s egg compressed between horizontal plates in the direction of the axis. The third furrow is more or less meridional and vertical in three quadrants, horizontal in the fourth, and this a smaller quadrant. 'l‘he fourth furrow is meridional in this quadrant, horizontal in the remaining three.

quadrants of the same egg (Fig. 20). It will be observed that the quadrant in which the third furrow is latitudinal is smaller than the others. It is of great interest to observe the striking similarity between the direction of the third and fourth furrows in these eggs and the corresponding divisions in the Teleostean egg where the blastodisc is compressed by the chorion.

In the second series of experiments made by Hertwig the glass 36 CELL-DIVISION AND GROWTII II. 1

plates were vertical, the eggs, therefore, compressed not, as before, in, but at right angles to, the axis.

The first furrow was meridional, and therefore vertical, and at right angles to the plates. The second was latitudinal and horizontal, and also at right angles to the plates. The furrows of the third phase were parallel to the first, those of the fourth, in the four upper animal cells, parallel to the plates. Born again

,1

/ 1 7

F1G.2l.—Segmentation of the Frog's egg under p1'ess1u'e. The pressure is at right angles to the axis.

A, view of the colnpressed egg. The piglnented animal portion is shaded.

If, C, 1), views of the egg from the animal pole after the first (1), the second (2), and the third (3) divisions.

E, 11‘, 0, views of the egg from the compressed side after the first (1), the second (2), the third (3), and the fourth (4) divisions. The ﬁrst furrow may pass as (1') in E.

(From Korschelt and Heider, after Born.)

conﬁrms this account (Fig. 21). The direction of the furrows of the third phase is, however, variable ; it may be not parallel to the first, but perpendicular to it. In this case it may be parallel to the second, or so oblique to it as to become nearly parallel to the glass plates. The direction of the fourth division depends on that of the third, to which it is at right angles. It may, therefore, be either oblique and nearly parallel to the plates, as described by Hertwig, or parallel to the second furrow and perpendicular to the plates. In a third series of experiments I-Iertwig placed the plates II. I CELL-DIVISION 37

obliquely, at 45°. In these eggs the yolk sinks slightly from the upper to the lower side, while the cytoplasm rises in the opposite direction; in other words, a bilateral symmetry is conferred upon the egg by the combined action of pressure and gravity. The plane of this symmetry is midway between and parallel to the plates. The first furrow is at right angles to the plates and to the plane of symmetry.

VVe are indebted to Drieseh for a similar series of experiments on Echinoderm eggs. Drieseh compressed the eggs of 15?:/ri/u/.s~ under a cover-glass supported by a bristle. The direction of the egg—axis with regard to the pressure was not known, but the

FIG. 22.——I9'rIu')ms: segmentation under pressure.

(1, preparation for third division (radial); b, preparation for fourth

division (tangential); 11', after fourth division; 0, another form of the 8-cell stage (third division pa1'allcl to ﬁrst); «I, the same after removal of the pressure. (After Driesch, 1893.) Echinoid egg is nearly isolecithal. VVhen the egg membrane remained intact the ﬁrst two furrows were vertical, that is, in the direction of the pressure, since the slide and cover-glass were horizontal, and at right angles to one another.

The spindles for the next division are again horizontal, and usually tangential, sometimes, however, radial. The eight-celled stage consists, therel'ore,o[’ a ﬂat plate of cells. At the next division the formation of mieromeres —which would ordinarily occur at this moment——is suppressed; the spindles are horizontal and radial, the furrows, therefore, vertical and tangential (Fig. 22 a, /1, /2’).

In certain cases cell-formation is wholly or partially suppressed. When the pressure is less (in those eggs which lie nearer the bristle) the micromeres may be, but generally are not, formed. The spindles are no longer horizontal. Similar results are obtained when the eggs are released from strong compression.

In another experiment the eggs were first deprived of their membranes. The ﬁrst and second furrows are vertical and generally at right angles to one another. Sometimes,-however, 38 CELL-DIVISION AND GROWTII II. I

the second is parallel to the first, or one blastomere may lie apart from the other three. Should the eggs be now released from the pressure, each blastomere becomes rounded off, and——after two more cleavages—-the sixteen-celled stage consists of two plates of eight eells lying over one another. But if the pressure is maintained, the spindles are horizontal and the blastomeres lie all, or nearly all, in one plane (Fig. 22 c, (7).

Flu. 23.--Segnieiitatioii of the egg of E(‘7li1lII8 micI'oh¢bcrcuIulus under pressure. (After Ziegler, 1894.)

(V, 8 cells in one plane; 1;, 16 cells, the last division having been tangential ; c, (I, 16-32 cells: the direction of the spindles in c is shown by the line: it is in the greatest length of each cell; c, 64 cells: a cross signiﬁes a vertical or oblique division, a line a horizontal division.

Ziegler has followed the segmentation of the compressed eggs a step further (Fig. 23). As the figures show, the first two divisions are at right angles to one another, while the furrows of the next two phases are, roughly, parallel to the first and second. In the next division-—sixteen to thirty-two cells—the outer cells divide radially, the inner more or less tangentially, these divisions being, like the previous ones, at right angles to the compressing plates. In the following phase, some cells (those marked with a line) still divide in the same direction as before; but in others (distinguished by a cross) the spindle is perpendicular to the II. 1 JELL-DIVISION 39

plates and the division horizontal. Ziegler points out that, in the former cases, the cells have greater dimensions in the horizontal plane than in the la.tter. This, however, may be the eﬁeet, not the cause, of the direction of the spindle-axis.

Two other pressure experiments maybe mentioned here. In Nereis Wilson produced a ﬂat plate of eight equal cells by applying pressure in the direction of the axis. The formation of the first quartette of micromeres was thus suppressed. On relieving the pressure eight micromcres were formed. For the Ctenophora (]ierb'z') Ziegler has shown that the normal inequality of the third and fourth divisions is not altered by pressure.‘

The foregoing experiments all agree in demonstrating the perfectly deﬁnite eft'cct produced by pressure upon the segmenting cgg. The nuclear spindles place themselves at right angles to the direction of pressure, the divisions fall at right angles to the compressing plates. This holds good for the ﬁrst three or four divisions, at least, and sometimes for later phases still. In all these cases, therefore, the nuclear spindle elongates in a direction of least resistance, and, in the normal uncompressed egg, we may argue, with Ilertwig, the least resistance is offered by the greatest protoplasmic mass.

Even in the compressed eggs, however, the greatest extension of the protoplasm, or the least extent of the yolk, is a factor which must in some cases come into play. When the egg of the Frog is compressed between vertical plates, the nuclear spindle does not elongate in any direction at right angles to the pressure, but in one only, a horizontal ; and this is the direction of the greatest protoplasmic mass, since the egg-axis is vertical.

Speaking generally, therefore, experiment has upheld Hertwig’s contention that the direction of nuclear division, and therefore of cell-division, is determined by the relation between the nucleus with its centrosomes and the cytoplasm with its yolk.

There are one or two experiments which do not support Hertwig’s view. Boveri stretched the eggs of the sea.-urchin St7'o2z_9;yloceul/'0!/1.9 in the direction of the axis. The fertilization spindle lay in the usual equatorial position, occupied, that is, the

minor axis of the ellipsoid.

‘ I have recently had occasion to notice that when the egg of Anfedm is compressed in the direction of the axis the third division is meridional instead of latitudinal. 40, CELL-DIVISION AND GROWTH II. I

Again, Roux observed that Frogs’ eggs sucked up into a tube with a narrow bore became elongated either parallel or transverse to the length of the tube, the axis of the egg lying in each case lengthways. In the first case the division was at right angles to, in the second usually parallel to, the tube in accordance with the rule; but exceptionally, in the transversely stretched eggs, the division was not perpendicular to, but coincided with, the extent of the greatest protoplasmic mass.

However important a factor the disposition of the yolk may thus be in deciding the direction of cell-division, it is certainly not the only factor. In the eggs pressed between horizontal plates there are many—an inﬁnite number—of' directions of least resistance. In one of these the segmentation spindle elongates, and at right angles to this the first furrow falls. This is probably determined-——-as it is determined in the normal Frog and Sea-urchin egg-——by the point of entrance of the spermatozoon, or at least by the direction of the sperm-path in the egg. The second division is at right angles to the first, and here the direction may very possibly be decided on lIertwig’s principle. But why, in the next phase, should the furrows be at right angles to the second rather than to the first, for the extent of the protoplasmic mass is as great in each of the four cells, in a direction parallel to the first as to the second furrow? Here, it is clear, some other reason must be found for this succession of divisions at right angles to one another. The cause is probably to be sought for in the direction of division of the centrosomes ; for these divide——frequently soon after the telophase—at right angles to the axis of the previous ﬁgure. VVe thus gain a new expression for Sachs’ Law.

The original direction of divergence of the centrosomes is,

however, by no means always the ultimate one, for the growing ‘

spindle may be twisted out of its original position. Conklin has made a careful study of this phenomenon in C’)'¢j[Ii(]l(ltl, in which egg he ﬁnds that vortical movements are set up in the cytoplasm by the escape of nuclear sap at the beginning of mitosis. The movements are in opposite directions in sister cells, centre in the spindle poles, and often carry both nucleus and spindle into a fresh place. These currents, which had been noticed previously by other observers (by Mark in Limam and by Iijima in Ale]//zelis), may II. 1 CELL-DIVISION 41

thus play an important part in the production of the cell pattern. We shall see elsewhere that they, and other protoplasmic movements, are also of the very greatest signiﬁcance in diﬁerentiation.

There remains now to be noticed another principle, which is especially applicable to plant-cells with ﬁxed walls, though it may possibly be used for the phenomena of animal segmentation as well. Berthold has pointed out that when a newly formed cell-wall places itself perpendicular to the previously existing walls it is——at least in a good many instances—simply obeying tlie laws of capillarity, it merely conforms to the principle of least surfaces formulated by Plateau. This principle is as follows: ‘ Homogeneous systems of fluid lamellae so arrange themselves, the individual lamellac adopt a curvature such that the sum of the (external) surfaces of all is under the given conditions a minimum.’

A fluid lamella, of soap solution, for example, placed across the interior of a hollow, rigid cylinder, or parallelepiped, or cube, is, with the ﬁlm coating the internal surface of the vessel

-in which it lies, a special case of such a system of lamellae, and,

in obedience to the principle, the lamella places itself at right angles to the walls of the cavity and transverse to the long axis.

In the ease of the plant-cell, the cell-plate, formed by solidiﬁcation of the spindle fibres in the equator of the mitotic ﬁgiii'e, represents the soap-lamella, and like the latter in its parallelepiped, the cell-platc, or new cell—wall, places itself perpendicular to the old one, and transverse to its length.

There are very numerous cases in which the law is obeyed, but it is not so in all. Under certain conditions the. lamella should be not at right angles, but oblique to the wall of the chamber across which it is stretched. If, to take a concrete case, the lamella be made to move (by abstracting air) towards one end of its receptacle (a cube or parallelepiped), it will reach a critical position in which the principle of least surface can only be satisﬁed by its occupying an oblique position. The

point at which this occurs is when the lamella is distant 3 from 7r

the end, where a is the length of the side of the cube (short side of the parallelepiped). The lamella new forms the fifth side

to a wedge-sliaped space (quadrant of a cylinder, whose radius

. 4 . . . . is 1- = 9-a), but as more air is abstracted, and it moves still 4-2 CELL-DIVISION AND GROWTH II. I

further toward the end, it comes to another critical position when it must lie across one corner, forming so the base of a pyramid, or octant of a sphere. This position is deﬁned by the equation 2-, =11, where 1', is the radius of this sphere. It is impossible, therefore, for a very ﬁat cell, or short cylinder, to be divided in conformity with the principle parallel to its longest side, and yet this occurs, as, for instance, in cambium cells.

It will also be noticed that this principle does not explain why one particular direction is selected when many are apparently equally possible.

We turn now to a consideration of the remaining factor which assists in determining the shape of the cells and so the geometrical pattern of segmentation ; this is the movement of the cells upon one another.

That such movement does occur we have already seen; the question which immediately suggests itself is whether in taking

up their new positions the cells obey the laws of capillarity as enunciated for systems of fluid lamellac such as soap-bubbles by Plateau in his principle of least surfaces.

This principle, as we have seen, demands that the sum of the external surfaces should be, under the conditions, a minimum, or, expressed in physical rather than in geometrical language, that the total surface energy should be minimal. In accordance with this doctrine of minimal surface energy a drop of fluid floating in a fluid medium assumes, as need hardly be said, the form of a sphere. In a system of drops contact surfaces will be formed between the drops, provided that each possesses a coating film which has a positive energy with the media it separates; a film, that is, of such a nature that the total surface energy

would be diminished by apposition, without, however, involving ‘

the disappearance of the separating ﬁlm and fusion of the drops. In other words, the ﬁlm must be insoluble in both the external and the internal media. A simple example of this is afforded by the behaviour of the spheres of jelly covering the eggs of the Frog, when taken from water and floated between chloroform and benzole. Two or more such drops of jelly cohere by their coating ﬁlms, and form systems of lamellae —the films, that is, at the external surfaces and between the opposed surfaces of the II. I CELL-DIVISION 43

drops———in which the principle of least surfaces is obeyed. Soapbubbles form similar systems. But where this condition is not fulﬁlled, as in oil-drops ﬂoated, for instance, between alcohol and water, the drops either unite or separate, each retaining its spherical form.

The geometrical analysis of such systems given by Plateau is as follows. In a system of two bubbles the curvature of the

surface of contact is given by the equation r = £7, where 7' is the radius of that surface, p, p’ the radii of the larger and smaller bubbles. Since the pressure varies inversely with the radius, the surface of contact is convex towards the larger bubble. When p = p’, 7- = a, and this surface is plane. Since there is equilibrium the external surfaces of the bubbles and their common surface meet at angles of 120°.

In a system of three bubbles there are three contact surfaces;

these meet in one line and make angles of 120° with one

.another. When there are four bubbles, however, the four con tact surfaces cannot meet in one line except for an inappreciable instant; they immediately shift their positions in such a way that two opposite bubbles meet and separate the other two from one another. There are thus ﬁve surfaces of contact, and these make angles of 120° with one another as before. This is the arrangement when four bubbles—whether equal or unequal is no matter—are placed side by side in the same plane. When, however, one bubble is placed in a different plane to the remaining three, four surfaces are formed and disposed in such a manner that the four lines, each formed by the intersection of three of these surfaces, meet in one point, making with one another angles of 109° 28’ 16", the angles at the centre of a tetrahedron. In short, the four are now tetrahedral] y arranged. The systems of drops of jelly alluded to above arrange themselves as do soap-bubbles under similar circumstances. What holds good of four holds good of an assemblage of any number of bubbles. The size of the bubbles is a matter of indifference, except to the curvature of the surfaces of contact, and, to a certain extent, to the arrangement. Thus, if four equal bubbles be placed in a plane, they will form together ﬁve surfaces of contact, one of which will be between two opposite 44 CELL-DIVISION AND GROWTH II. I

bubbles. If these two be now diminished, or the opposite two enlarged, the surface of contact will be between the opposite pair of larger bubbles. On the other hand, it is possible to bring smaller opposite bubbles into contact, while the larger ones remain apart. Again, on four bubbles lying in one plane, four small ones may be superimposed in such a fashion that while two lie at either end of the surface of contact, the other two lie over between the two opposite large bubbles below. If now the two latter small bubbles be enlarged, they will displace the other two until all four come to lie not over but between the

FIG. 24.—Diugra1ns of systems of soap-bubbles.

A-0, four small bubbles superimposed on four large ones. In A and B the bubbles are not compressed ; in C the lower bubbles have been circumscribed by a. cylindrical vessel. In B the upper bubbles are small enough to show the surfaces of contact between each and the two adjacent large bubbles below. These surfaces are invisible in A and C.

D is a system of eight bubbles in one plane, four forming a cross in the centre.

In all figures notice the ﬁfth contact surface or ‘polar furrow ’.

bubbles below, the usual arrangement when four are superimposed on four (Fig. 24 A—C).

The ﬁnal disposition must depend, therefore, not merely on the principles of least surfaces, but also, provided that the conditions of that principle are fulfilled, on the sizes and initial arrangement of the bubbles.

It will hardly need pointing out that very many ova adopt the form which presents the least external surface, that of a II. I CELL-DIVISION 45

sphere, when placed in a fluid medium, and it is also a familiar fact that after the first (and subsequent) divisions the blastemeres are ﬂattened against one another (Cytarme, to use Roux’s term), and that whether they are compressed by an egg membrane or not (examples of the second alternative are to be found in Unio, .D1'eis3eu8z'a, Umbrella, C'7'q2i(I'/(la, /lp/yxia limecimz, /late;-iae), though the surface of contact is not always curved when the cells are unequal. The two cells, however, often become rounded of and partially separated from one another prior to the next division. Such a separation (Cytochorismus) has also been observed by Roux in the ease of cells of the Frog’s egg, which, having been isolated in albumen or salt solution, have subsequently reunited.

That the cells flatten against instead of repelling one another, as free oil-drops would do, suggests that they, like soap-bubbles, are provided with an insoluble coating-ﬁlm, while their subsequent separation may be provisionally explained by supposing that this coating-film becomes temporarily dissolved under the action of some substance formed in the cell. This idea is borne out by a striking experiment of Herbst’s, who found that in sea-water deprived of its calcium the blastomeres of the seaurchin egg came apart and resumed their spherical shape. At the same time the surface membrane underwent a visible alteration, becoming radially striated. It seems reasonable to conclude that there is a membrane by which contact is normally effected, and that this is soluble in sea-water devoid of calcium. On the addition of calcium the cells eohere again.

It may be mentioned that when systems of drops of jelly, ﬂoating in a medium of oil and united by their coating-ﬁlms of water, are removed to alcohol, in which both oil and water are soluble, the ﬁlms disappear and the drops separate.

In the next stage (four cells) the type of segmentation in which the laws of capillarity are most strictly obeyed is obviously that which we have distinguished above as the spiral or tetrahedral type, and Robert has been able to show that successful imitations of the four-, eight-, twelve-, and sixteen-celled stages of the egg of 2’7'oc/we may be made with soap-bubbles.

Four equal bubbles were placed in a porcelain cup, which held them together in the same way that the actual cells are held 46 CELIi-DIVISION AND GROWTH II. I

together by the vitelline membrane. Five surfaces of contact were formed, that between two opposite bubbles representing the cross furrow or polar furrow in the egg. In the fl’/'oc/ms egg, however, the polar furrows need not be parallel at the animal and vegetative pole; they may be at right angles to one another, and this tetrahedral arrangement of crossed polar furrows may be imitated by lifting up one of the bubbles and bringing it into contact with its opposite, one pair of bubbles being new in contact below, the other pair above. This arrangement is, however, unstable whilc the four bubbles remain in one plane, the two bubbles soon coming into contact both above and below. When the bubbles are not conﬁned within a cup the instability of the ‘ crossed-furrow ’ condition is extreme.

By reducing the volume of the bubbles that are in contact the other two may be brought together; as the polar furrow changes positions there is at least a temporary condition when they are crossed.

As we have already pointed out, both conditions—the ‘ parallel furrows’ and the ‘ crossed furrows ’—-are met with in the eggs at the four-celled stage of Molluscs, Annelids, and marine Turbellarians. Whether both opposite pairs or only one opposite pair of blastomeres are in contact does not, however, appear to depend upon whether the vitelline membrane is close to and compresses the egg or not. In most cases of crossed furrows the membrane ﬁts, it is true, quite closely (Nereis, Io/moo/titou, Porlar/cc, Lcpidouotus, Jjiscocelis, P/(yea, and possibly Li)/may and Planorbis, if there is in these two, as in P/13/ea, a very fine membrane between the albumen and the ovum); so also, speaking generally, where the furrows are parallel the membrane is absent (Umbrella, ./ljzlysia, Dreisseueia, Crepirlu/a), but in Am];/aitrile and C/gmzenella it is lightly applied to the egg.

It is remarkable that when the furrows are crossed, it is the A and C cells which meet at the animal pole, the B and D cells at the vegetative (except only in U/tio), and this must depend on other properties of the cells than their surface tensions. But it may be very plausibly suggested that the explanation of the fact that it is the cells B and D which meet to make the ‘parallel’ furrows is to be looked for simply in the large size of D.

Robert has indeed shown that by simply altering the sizes of II. 1 CELL-DIVISION 47

the bubbles the conditions observed in the four-celled stage of other types—Nereis, A/-euicola, Uuio, zlplysia, I)isc-ocelz'.s°-may be faithfully copied.

It only remains to be added that the contact surfaces of the cells, like those of the bubbles, make angles of 120° with one another.

Robert has also imitated the eig-ht—eelled stage (the four micromeres alternating with the four maeromercs), the stage of twelve cells (division of the micromeres), and that of sixteen cells (second quartettc formed). The bubbles of the second quartette may be made to slide in between the maeromeres and so rotate the whole first quartette, as happens in the egg. The division of the micromeres in the egg results in the arrangement of four cells crosswise in the centre, four others occupying the spaces between the arms of the cross. The bubbles behave in the same manner.

In the eigl1t—celled stage the micromeres alternate with the macrorneres. In the case of the bubbles this is not necessarily so; the two sets of bubbles may be superposed if the ‘polar furrow’ in one tier is at right angles to that in the other, or if, as pointed out above, the upper bubbles are small. Otherwise superposition is a very unstable condition.

It would appear then that many of the patterns exhibited by eggs with a spiral cleavage are explicable by reference to the laws of surface tension. The principle of least surfaces may be extended to other cases. The first four blastomeres of Ophiuroids and Asteroids form a perfect tetrahedron, though this arrangement is subsequently discarded for one which could not be imitated with soap-bubbles (we may notice in passing that in the ﬁrst case the egg is tightly invested by its membrane, in the second it is perfectly free). In zlscarzlv megalocep/aala the four cells come to lie, as do four bubbles, in one plane, and polar furrows have been seen in many eggs which belong to another type of segmentation (in Coelenterates (llyrlractiuia), Sponges (Spongillu), Crustacea (Brauc/z2'pu.v, Luci/‘er, 0rc/Iestia), Vertebrates (Petra/1z_yzo:2, Rana), Ascidians, and Am/2/u'oama).

The principle of least surfaces——not more than three surfaces meeting in a line, not more than four lines meeting in a point— is, however, not of itself suflicient to explain the whole of the phenomena even in this most favourable tetrahedral type; 48 CELL-DIVISION AND GROWTH II. I

other factors must intervene, just as other factors intervene in a mass of soap-bubbles—their size and initial arrangement~— in the determination of the actual pattern. These other factors are the direction of cell-, that is of nuclear, division, and the magnitude of the cells; and these, as we have seen, in turn depend upon the relation between the nucleus and the cytoplasm with its included yolk. Thus it is the direction of the spindles which determines whether the mieromeres of the first quartette shall be given oﬁ laeotropieally or dexiotropically ; the direction of division, oblique to the egg-axis, again determines that the mieromeres shall alternate with the macromeres and not be superimposed upon them ,- the size of the cells and the direction

FIG. 25.-— Mitotic division with elongation of the cell-body in a protozoon,Acanthor_1/stis aculeuta. (After Schaudinn, from Korsehelt and Heider.)

of division may determine the position of the polar furrow, while the rate of division will also not be without effect, since the whole arrangement at any stage depends in part on the disposition at the stage before.

There is one other point that is worthy of notice. The mitotic spindle possesses considerable rigidity, and is able as it elongates to materially alter the shape of the cell. This may be seen in many cases in Annelid, Mollusean and other eggs—the division of the first mieromeres in Nereis is an instance—and in the Protozoa (Fig. 25). Another interesting case is the Rotifer Asplcmc/ma, where, preparatory to the fourth division, the shortest axis of the cells——in which the spindles are placed-—becomes by the elongation of the spindles the longest. This alteration of shape is itself an important factor in deciding the positions to be taken up by the daughter cells. II. I CELL-DIVIS ION 49

In the other types——radial and bilateral——thc principle of least surfaces is obviously disobeyed, for here four or more surfaces meet in one line and at angles other than 120°.

Roux (1897) has, however, shown that if a certain condition be imposed on the system of lamellac, ﬁgures may be produced which very closely resemble the patterns presented by radially and bilaterally segmenting ova. This indispensable condition is that the system shall be surrounded by a rigid boundary, as the eggs themselves are by a membrane. Roux’s system was made by dividing into two, four, and eight a drop of paraflin oil suspended in a closely ﬁtting cylindrical vessel between alcohol and water. To this medium was added calcium acetate to prevent the drops reuniting. The drop was divided with a glass rod.

C9®f® ®a@

Fm. 2(5.~ltoux's oil-drops. A and B, the drop divided equally; U and

D, unequally. Each of the two equal drops divided equally in E, unequally in 1". (From Korschelt and Heider, after Roux.)

When the two drops formed by the first division were equal the surface of contact was ﬂat, when unequal convex towards the larger one, in accordance with the rule (Fig. 26 A—l)).

When the second was also equal, four drops were formed with four surfaces of contact meeting in one line, or enclosing between them a small ‘segmentation’ cavity. If the division of the two equal drops was unequal, and the smaller cells adjacent, they pushed into the larger ones; the result, in fact, was the same

Jicnxnsnv E 50 CELL-DIVISION AND GROWTH II. 1

as would have been produced by an equal following on an unequal division, the four surfaces meeting in one line as before (Fig. 26 E, F). The appearance presented is like a side view of a radially

Fm. 27.—-Arrangement of four oil-drops produced by unequal division of two equal drops, the small and large drops alternating. The first division is shown by I: the second (II) may pass as in a or in b, but the result is always as in c, the two large drops meeting in a polar furrow and excluding the small drops from the centre; the system is symmetrical (iso~bilateral) about the dotted lines in c. (After Roux, from Korschelt and Heider.)

segmenting egg after the third division. When, however, the smaller drops were 11ot adjacent, but opposite, five surfaces of con ” I 3 /. tact were formed,

a polar furrow

appearing between

A A the two larger and

I L 1 joining the centres \ /7 W of mass of the two smaller drops,

I I whether these are unequal or not.

C’ . . The direction 1n

ﬁ which the division

of the drops is per” V 3’ formed isirrelevant; the ﬁnal result is 4% always the same.

/ Should two adjacent

_lfIG. 2S.-— A_and B are diag1':1111s of an oil—drop drops be equal, the divided into four and eight to explain Roux’s 1 f _ - - 1 notation. C is a ﬁgure of the oil-drop divided into P0 M m row 15 Sh]

eight equal parts. (From Korsclielt and Heider.) formed by the union of those two which have together the larger mass (Fig. 27). II. I CELL-DIVISION 51

The length of the polar furrow varies directly with the size of the drops which unite to form it; its direction makes an angle with the plane separating the first two, which varies

FIG. 29.~Arrangenicnts of six oil-drops. In all cases A = B = a = b. In A, rt’ = a", b’ = b". In B, a’ > a", b’ > b”. In C, a’ < a”, b’ < b”. I, ﬁrst furrow; II, second furrow. (From Korsclielt and Heidcr, after

Roux.)

FIG. 30.-—Various arrangements of eight oil-clrops, all bilaterally syiiiinetrical about the ﬁrst furrow (1). In all cases the ﬁrst division has been equal. In A and B the second division (II) has also been equal, but in C 0, b are smaller than A, B. In A, a”, la”, A”, B" < rs’, b’, A’, B’. In B and C, a”, b" < a’, b’, but A", B” = A’, B’; hence a”, I)" < A", B". (From Korschelt and lleider, after Roux.)

_Ei_G. 31. —~ Arrangement of six (A) and eight (13) oil-drops, after iineqnal division of four equal drops (A = B = a = 1;), the smu.llcr and linger drops regularly alternating. (From Korschelt and Hcider, after Roux.)

E2 52 CE LL-DIVISION AND GROWTH II. 1

inversely with its length, so that when all the drops are equal the cross furrow lies in the same plane with the first division, and so disappears. .

By another division it is possible to make a ring of eight drops whose surfaces of contact all meet in one line, or in a ‘segmentation’ cavity (Fig. 28). To realize this condition, however, it is necessary that the division should be equal, and its direction accurately radial. If unequal, the larger drop invariably passes towards or wholly into the inside. If oblique or tangential the inner drop passes into the segmentation cavity (Fig. 32).

FIG. 3'Z.——'l‘hree stages in the passage of a large drop (a") into the centre of the system. The ﬁist stage extremely unstable. (From Korschelt and Heider, after Roux.)

Unequal division of all four equa.l drops produces very interesting patterns, some of which recall the appearance of bilaterally segmenting ova, when the divisions are corresponding-ly unequal on each side of the ﬁrst or second division (Figs. 29, 30), while others resemble certain phases of ‘ spiral’ division when small and large cells regularly alternate (Fig. 3]). It is a rule for the smaller of the two drops to go to the periphery, while the larger assumes an oblong or wedge shape, passing towards the centre if it does not slip entirely inside. The latter occurs with clean oil, when the large drop is flanked by small ones on both sides.

It is also possible to divide four equal drops horizontally into two tiers. The upper drops, however-—-unless absolutely undistnrbed~quiekly come to alternate with the lower.

In these systems of drops the ﬁnal arrangement is due to, ﬁrst, the principle of least surfaces; secondly, the circumscribing boundary; thirdly, the size of the drops; and fourthly, in some cases, the direction in which they are divided.

It only remains for us to consider, with Roux, to what extent II. I CELL-DIVISION 53

the cells of a radially segmenting egg, such as that of Rana fusca, are governed by the same inﬂuences as determine the pattern of the drops.

The resemblances, it will be conceded, are often very close. There are also important differences. The polar furrow, which is often present in the Frog’s egg, is not necessarily between the cells with the greatest mass. Again if, in the four-celled stage, with no polar furrow, one of the cells be diminished by puncture, a polar furrow does not always appear, as it would with oil-drops, nor, if it does, is it always formed by the union of the larger cells. 01', if when a polar furrow is present between the larger cells, one of these is diminished by puncture until it, together with its opposite, is less than the other two, the polar furrow nevertheless retains its position.

In the sixteen-celled stage the animal cells together form a ring of eight around the axis. The cells are not necessarily equal, and a small cell may be compressed by, instead of compressing, adjacent large ones, while they, not it, move away to the periphery.

Other differences are that large cells bulge into small, that cells are elongated tangentially instead of radially, that there are amoeboid processes at the inner ends of the cells, and intercellular spaces between them.

Further, Roux has examined the behaviour of the isolated cells of the Frog's egg in the morula stage. The cells were separated in a medium of albumen, or salt-solution, or a mixture of the two. They first approach and then ﬂatten against one another (Cytarme), as do the blastomeres in the egg, completely or incompletely. The contact surface is generally symmetrical to the line joining the centres of mass of the two cells ; it may be concave towards either the small or the large cell. More than two cells may unite to form rows or heaps. The angles made by the surfaces of contact may be 120°, or have other values. Four surfaces may meet in one line ,- at other times the arrangement is tetrahedral. In a 1-25 Z solution of salt the cells are elongated, and united end to end in long branching strings. The pigment, diffused through the cell, later returns to its original position at the surface, or usually to the middle of the free surface of each.

The cells may also move over one another (Cytolisthcsis) by 54- CELL-DIVISION AND GROVVTH II. I

sliding or rotation, or both. Even two cells will glide on one another, as two soap-bubbles will not. In complexes two threesurface lines may unite to form one four-surface line, a behaviour the very opposite of that exhibited by soap-bubbles.

It appears, then, that in the living egg of the Frog (and other radial and bilateral types) there are factors which overcompensate, to use Roux’s expression, the purely physical factors by which the behaviour of the oil-drops is governed. These organic factors are that division is slow, and begins on the outside ; that the direction of division—determined by the yolk—is persistent; that the cell contents are neither perfectly ﬂuid nor perfectly structureless ; that the cells being different, their surface tensions may be of dilferent magnitudes, and the whole system, therefore, not homogeneous; and that the cells possess a more or less solid rind or membrane, the rind which becomes wrinkled transversely to the furrow when the cell divides.

It would seem that this rind is an important factor, for if Roux’s experiments be repeated with drops of albumen suspended between xylol and oil of cloves, to which a little alcohol has been added, it will be seen that each drop gets a su1:erficial membrane, and that by these membranes adjacent drops adhere. In fact, such drops behave more like the cells of the egg‘ than do the oildrops. Thus, a small cell goes towards the inside, or the outside, according" to the way in which the division is made, and, after a horizontal division of four equal cells, the upper remain superimposed upon the lower.

At the same time, it is apparently because the cells have this surface ﬁlm, which the oil-drops have not, that they are able to flatten against one another as soap-bubbles do; while, on the other hand, it is because the ﬁlm is solid that the cells are unable to move upon one another and adopt the geometrical arrangement seen in systems of soap-bubbles.

There is still another kind of cell-movement to which brief reference must here be made, since it is found in one type of segmentation at least. In the segmenting eggs of some Platyhelmia (Triclads), Ascidians (Salps), Echinoderms (Asteroids), and Coelenterates (0ceam'a), the blastomeres have been seen to completely separate from one another, afterwards reuniting. Roux II. I CELL-DIVISION 55

has observed a similar reunion of the artiﬁcially isolated cells of the Frog’s egg. This Cytotropism, as Roux calls it (Cytotaxis would be a preferable term), is noticed when the slide is kept perfectly horizontal and streaming movements of the medium (albumen) are rigidly excluded. The cells become rounded, and then approach one another in, more or less, a straight line, oscillating slightly backwards and forwards. The cells must not be too far apart, not further than a radius of small, or less of large cells. Groups of two or more cells behave in the same way.

The movement may be simply a surface-tension phenomenon, or, as Roux suggests, more complex, of the nature of a response to a mutual ehcmotactic stimulus.

These various kinds of cell-motion are also an important feature in such processes of differentiation as the union of cells to form muscles, tendon, epithelia, and so forth.

A review of all the facts thus leads us to conclude that while some of the phenomena of segmentation-—-the ﬂattening of cells against one another, the pattern made by the cells in cleavage, especially of the spiral type—are largely referable to the action of the purely physical laws of surface tension, there are many cases, the radial and bilateral types, and the radial and bilateral periods of spirally segmenting eggs, in which the operation of these laws is restricted and conﬁned by other causes. But in any case those laws can only co-operate with other factors, which are to be looked for in the rate and direction of division, and in the magnitude of the cells, factors which themselves are dependent on the relation between the cell and its nucleus.

Before concluding this section we have to call attention to some experiments which may possibly throw some light on an event of fairly frequent occurrence in ontogeny—the division of the nucleus without the division of the cell,‘ as in the formation of coenocytia such as striated muscle ﬁbres and the trophoblast of the placenta ; or the fusion of distinct cells into a syncytium, as in the trophoblast again; or the secondary union of yolk-cells.

‘ In the Alcyonaria the nucleus may divide three, four, or ﬁve times before the egg simultaneously breaks up into eight, sixteen, or thirty-two

cells. See especially E. B. Wilson, ‘On the development of Renilla,’ Phil. Trans. Roy. Soc, clxxiv, 1883. 56 CELL-DIVISION AND GROWTH II. I

Driesch has observed that in the egg of Ea/Linus cell-division may be wholly or partially suppressed by pressure, and also by diluting the sea-water. Nuclear division continues (Fig. 33).

Morgan has found that the egg of another sea-urchin (Arbacia) will not segment in a 2 Z solution of salt in sea-water; on replacing the eggs in sea—water, however, the nucleus divides with great rapidity several times, and this is followed by celldivision. So Loch notices that the eggs when treated in this way, and brought back to their normal medium, divide simul taneously into four. The egg

es of the fish Cteuolabm.s (accord O“ o«° ing to the same author) behaves in a similar fashion when first f I deprived of, and then restored

,, 1, to oxygen. . FIG. 33.—Echinus: suppression of lmf’ again’ has Seen the re cell-division by 1)1'essure, I), and by union Of sister cells and nuclei

heat, a. Nuclear division continues. ' . ' . (After Driesch, 1893.) in the eggs of Avbacza leleased

from pressure.

Three distinct agencics——mechanical pressure, increase of osmotic pressure, and decrease of osmotic prcssure—arc all capable of effecting this interesting change in the usual relations of cell and nucleus. We can only guess at the real cause, and surmise that it will be found in an alteration of internal and external surface tensions.

LITERATURE.

No'rE.—For a complete bibliography of segmentation the well-known textbooks of 0. Hertwig and Korsehelt and Heider must be consulted. The literature of ‘spiral’ segmentation is given by Robert (quoted below).

F. M. BALI-‘OUR. Comparative Embryology, London, 1885.

G. BERTHOLD. Studien ﬁber Protoplasmameehanik. VII. Theilungsrichtungen und Thcilungsfolge, Leipzig, 1886.

G. BORN. Ucber Druckversuche an Froscheiern, Anat. Anz. viii, 1893.

T. BOVERI. Die Entwiekelung von Asca;-is megalorephala mit beson— derer Riicksieht auf die Kernverhaltnisse, Fesfschr. Xupﬂkr, Jena, 1899.

E. G. CONKLIN. P1-otoplasniic movement as a factor of differentiation, Woods Hell Biol. Lech, 1898. II. I CELL-DIVISION 57

H. DRIESCH. Entwicklungsmechanische Studien, IV, Zeilschr. wiss. Zool. lv, 1893.

H. DRIESCH. Entwicklungsinechanische Studien, VIII, Mitt. Zool. Stat. Neapel, xi, 1895.

A. FISCIIEL. Zur Entwicklungsgeschichte der Echinodermen. I. Zur Mechanik der Zelltheilung. II. Versuche mit vitaler Fiirbung, Arch. Ent. Mech. xxii, 1906.

J. H. GEROULD. Studies on the Embryology of the Sipunculidae, Mark Anniversary Volume, New York, 1903. _

A. GRAF. Eine 1-iickgitngig gemachte Furchung, Zool. Anz. xvii, 1894.

C. HERBST. Ueber dais Auseinandergehen von Furchungs- und Gewebezellen in ka.lkfrciem Medium, Arch. Ent. M¢'c7z. ix, 1900.

O. HERTWIG. Die Zelle und die Gewebe, Jena, 1893.

O. HERTWIG. Ueber den Worth der ersten Furchungszellen fiir die Organbildung des Embryo, Arch. nu'l.-r. Anal. xlii, 1893.

O. HERTWIG. Ueber einige mu befruehteten Froschci durch Centrifugalkraft hervorgerufene Mcclnmomorphosen, S.-B. Kimigl. prcuss. All-ad. 11733., Berlin, 1897.

J. LOEB. Investigations in physiological morphology, Joum. Morph. vii, 1892.

J. LOEB. Untersuchungen ﬁber die physiologischen Wirkungen des Sauerstoffnmngels, I7l:2ger‘s Arch. lxii, 1896.

J. LOEB. Ueber Kerntheilung ohne Zellt-heilung, Arch. Ent. Mech. ii, 1896.

T. H. MORGAN. The action of salt solutions on the unfertilized and fertilized eggs of Arbacia and of other aniimtls, Arch. Ent. Mech. viii, 1899.

E. PFLi'IGER. Ueber die Einwirkung (ler Schwerkra.i't und andere Bedingungen nuf die Richtung der Zclltheilung, 1_’ﬂc'«'ger‘s Arch. xxxiv, 1884.

J. PLATEAU. Statique des liquides, Paris, 1873.

A. RAUBER. Der karyokiuetische Process bei erhohtem und vermin(lertexn Atmosphiirendruck, Vcrs. Deutsch. Naimf. u. Aerzte, Magdaburg, 1884.

A. ROBERT. Rec-herchcs sur le développement des Troques, Arch. Zool. E.1'p. et G6». (3), x, 1902.

W. Roux. Ueber die Zeit der Bestimmung der Hauptrichtungen des Froschembryo, Leipzig, 1883, also Ges. Abh. 16.

W. Roux. Ueber den ‘Cytotropismus’ der Furchungszellen des Grasfrosches (Rmmfusca), Arch. Ent. Mech. i, 1894.

W. ROUX. Ueber die Selbstordnung (Cytotztxis) sich ‘beriihrendcr' Furchungszellen des Froscheies durch Zellonzusammenfiigung, Zellentrennung und Zellengleiten, Arch. Ent. Mech. iii, 1896.

W. Roux. Ueber die Bedeutung ‘geringei-' Verschiedenheiten der relativen Grﬁsse der Furchungszellen fur den Clim-akter des Furchungsschemas, Arch. Ent. Mech. iv, 1897. 58 CELL-DIVISION AND GROWTH II. 2

G. SMITH. Fauna und Flora. des Golfes von Neapel: Rhizocephala,

Berlin, 1906. O. ZUR STRASSEN. Embryonalentwicklung des Ascarisnmgalorephala,

Arch. Em‘. Jlfcah. iii, 1896. E. B. VVILSON. (_1leava.ge and mosaic work, An-7:. Ivlul. lilo:-72. iii, 1896. H. E. ZIEGLER. Ueber Furchung unter Pressung, Vcrh. Aunt. Gesell.

viii. 1894. H. E. ZIEGLER. Untersuehungen ﬁber die eisten Entwicklungsvor gétnge der Nematoden, Zeifschr. W1'.s-s. Zool. lx, 1895. H. E. ZIEGLER. Experimentelle Studien ﬁber die Zelltheilung, Arch. Elli. llferh. vii, 1898.

2. GROWTH

Following Davenport we deﬁne growth as increase in size or volume. Since, therefore, growth is increase in all three dimensions of space, it is most accurately measured not by increase in some one dimension—such as stature——but by increase of mass or weight.

Growth depends upon the intake of food and the absorption of water and exhibits itself in the form of increase in the amount of living matter or of secretions of watery or other substances, organic or inorganic, intra-cellular or extra-cellular, such as ehondrin, fat, muein, cellulose, calcium phosphate, and the like.

That growth depends———in later stages at least—upon the intake of food is obvious. That it is due to the absorption of water has been demonstrated effectively by Davenport for the tadpoles of Amphibia (zlmtlys/oma, Rana, I311/2)). The method employed was to weigh known numbers of the tadpoles at diﬂ:'erent ages, desieeate and weigh again. The results of the investigation are shown in the accompanying ﬁgure (Fig. 34-), from which it will be seen that the percentage of water rises with remarkable rapidity———from 56% to 96% during the first fortnight after hatching. After that point the amount of water present slightly but steadily declines.

The same result is brought out by an analysis of the terminal buds and successive internodes of plants. It is found in I/ele7'oceutrou (Kraus) that the percentage of water rises rapidly from the terminal bud to the first internode, more slowly from the first to the second internode, and then remains constant. II. 2 . GROWTII 59

It would thus appear that during the period of most rapid growth, growth is eﬁfected by imbibition of water rather than by assimilation, since the weight of dry substance in the tadpole during this period does not increase at all.

In later development the proportion of water slowly falls. This may be seen not only in Davcnport’s table of the growth of Frogs but in the data furnished by Potts for the Chick and by

FIG. 34.— Curve showing change in percentage of water in Frog tadpoles from the first to the eighty-fourth day after hatching. Abscissae, days; ordinates, percentages. (After Davenport, from Korschelt and IIe1der.)

Fehling for the human embryo. These data. are given in the accompanying tables (Tables I, II). The percentage of water, at first high, slowly falls in both cases; conversely, the percentage of other substances increases.

‘ These results indicate that during later development growth is largely effected by excessive assimilation or by storing up formed substance’ (Davenport).

There are other external agencies by which growth may be affected in various ways—-such as heat, light, and atmospheric pressure. These will be discussed in another chapter. For the present let us conﬁne our attention to certain features which are characteristic of growth in general, of the growth of the animal organism under normal conditions. These are the changes that 60 CELL-DIVISION AND GROWTH II. 2

take place during growth in the rate of growth itself, in the variability of the organism and in the magnitude of the correlations between its various parts.

TABLE I

Showing the percentage of water in Chick embryos at various stages up to hatching. ( From Davenport, 1899 (2), after Potts.) ’1_‘he table also shows the hourly and daily percentage increments of weight.

Absolute Hourly Daily

H f . . P r entage

48 0-06 83 54 0-20 0-14 38-3 919-2 90 58 0-33 0-13 16-0 384-0 88 91 1-20 0-87 7-9 189-6 83 96 1-30 0-10 1-7 40-8 68 124 2-03 0-73 2 0 48-0 69 264 6-72 4-69 1 6 38-4 59 TABLE II

Showing the percentage of water in the Human embryo at various stages up to birth. (From Davenport, 1899 (2), after Fehling.) The table also shows the weekly percentage increments of weight.

Age in Absolute weight \Vec-kly per- Percentage

weeks. in grammes. Increase‘ ccntnge increment. of water.

6 0-975 97-5 17 36-5 35-525 331-2 91-8 22 100-0 63 5 34-8 92-0 24 242-0 142-0 71-0 89-9 26 569-0 327-0 67 6 86-4 30 924-0 355-0 15-6 83-7 35 928-0 4 0 0-1 82 9 39 1640-0 712-0 19-2 74-2

VVe follow Minot and Preyer in measuring the rate of growth by the percentage increments of weight (or of other measurements where weight is not available) during a given interval of time ; that is to say, by expressing the increase in weight during a given period as a. percentage of the weight at the beginning (or end) of that period. The change of rate, if any, is found by taking such percentage increments for successive equal increments of time.

As a first example let us consider the data furnished by Minot II. 9. GROWTH 61

himself for the rate of growth, after birth, of guinea.-pigs (Table III, Fig. 35).

TABLE III

Showing the change of rate of growth in male and female Guinea.-pigs. as measured by daily percentage increments of weight. (From Minot, 1891.)

Average daily per cent. Average daily per cent.

Ago in Age in

d“Y‘- Ma1§§°'°m§'§§$1cs. “‘°““‘s- Ma1§§.°r°m°§§:.a1..s. 1-3 0.0 2.1 8 0.05 0.2 4-6 5.6 5.5 9 0.3 0.2 7-9 5.5 5.4 10 0.1 0.1

10-12 4.7 4.7 11 0.04 0.1

13-15 5.0 5.0 12 0.1 0.05

16-18 4.1 4.3 13 -0.2 0.3

19-21 3.9 3.5 14 0.5 -0.03

22-24 3.1 1.7 15 0.2 0.00

25-27 2.3 1.9 16 0.07 0.2

23-30 2.3 2-6 17 -0.1 -0.02

31-33 1.9 1.3 13 -0.05 -0.2

34-36 1.7 1-6 19-21 0.006 -0.1

37-39 1.9 1.3 22-24 0.02 -0.05

40-50 1.2 1.1

55-65 1.3 1.3

70-80 1.2 0.3

35-95 0.9 0.9

100-110 0.7 0.3 115-125 0.6 0.5 130-140 0.1 0.2 145-155 0 4 -0.03 160-170 0.3 0.5 175-135 0.2 0.2 190-200 0.2 0.2 205-215 0.4 0.3

|||||||i| I'I_ ||||||| II it

15Il| I7 D E II N 75 I50

FIG. 3-'3.—Curve showing the daily percentage increments in weight of female Guine-.1-pigs. (From Mxnot, 1907 ) 62 CELL-DIVISION AND GROWTH II. 2

An inspection of tlie accompanying table and ﬁgure in which Minot’s results are reproduced will show at once that there is in both sexes, almost from the moment of birth, a. decline in the growth-rate. The decline is not, however, uniform. The rate falls rapidly between about the ﬁfth day (when it is from 5% to 6%) and the ﬁftieth, from the ﬁftieth day onwards more slowly, becoming eventually very small, zero or even negative. The younger the animal, therefore, the faster it grows; the more developed it is the more slowly it grows. The rate of growth in fact varies inversely with the degree of diﬁerentiation. A mammal, therefore, which is born in a less developed condition than is the guinea-pig ought to grow at first more rapidly still. The rabbit is such an animal, and Minot has been able to show that on the fourth day after birth the young rabbit adds 17 % to its weight. The curve also shows the same rapid decline in the growth-rate as was observed in the guinea-pig, followed by a period of gentle decrease.

Accurate observations on the prenatal rate of growth of these two mammals are lacking, but Henscn’s few observations (quoted by Preyer) on the weight of guinea-pig embryos show that the daily percentage increase descends from 220% on the twenty-first day to 116% on the twenty-ninth day, to 33% on the forty-third (lay, and again to 6% on the sixty-fourth day, that is just after birth, the moment at which Minot’s observations begin. Again, Minot has found, as a result of the investigation of the weight

TABLE IV

Showing the decrease in the rate of growth of the Human embryo before birth. Percentage increments calculated from the ﬁgures given by

Hecker, Toldt, and Hennig. (From Preyer.)

Average monthly percentage increments of Month. Weight. Length.

(Hecken) (Toldt.) (Hennig.) 1 _ _ __ 2 — 133-3 433 0 3 — 100-0 110-0 4 418-2 71-4 92-8 5 398-2 66-7 69-8 6 123-2 50-0 28-2 7 92-2 16-7 14-2 8 28 8 14-3 11-9 9 25-6 12-5 6-3 10 — 11-1 4 3 II. 2 GROWTH 63

of spirit specimens of rabbit embryos that the mean daily percentage increment is 704 between the ninth and ﬁfteenth days, but between the ﬁfteenth and twentieth days only 212.

The postnatal decline in the growth-rate is therefore only a continuation of a process which has been going on for some time, perhaps from the first moment at which growth began.

The human being forms no exception to this rule. Data of the growth of the human embryo before birth are somewhat meagre, but an inspection of the tables will show that whatever the discrepancies may be between the results obtained by Fehlin g (Table II) and Hecker (Table IV), they agree in this, that the growth-rate falls with great rapidity between the fourth and the sixth months,thereafter more slowly till the end of pregnancy. This is graphically represented in the curve (Fig. 36). It will be observed from the table (Table IV) that the rate of increase of stature also declines, but less abruptly. This is a poi11t to which we shall return. For the study of the postnatal growth of man very numerous data have FIG’ 36' —' Curve Sh°‘”il‘<‘=’

_ monthly prenatal percentage 1nbecn collected byvarious observers. cmments in Man. (From Minot, Measurements of the bed y weight 1907-) have been made on Belgians by Quetelet, on Boston school children by Bowditch, on the school children of Worcester, Mass., and Oakland, Mass., by Boas, and on English of the artisan and the well-to—do classes by Roberts. It is unnecessary to reproduce all these data here, for they all show the same decline in the growth Monnvso-zaascrogm 64 CELL-DIVISION AND GROWTH II. 2

rate, but Quetelet’s measurements for males, being the completest series, are given in the accompanying ﬁgure (Fig. 37). The ﬁgure shows that at the end of the ﬁrst year after birth the per.

2005‘;

any 71/‘ I Mwmtﬁaez;

500%

E-§

._.-.. YEMISI 2 3 0 5 6 7 8 0 Ion Iz I3I4I5 I617 uamzozuzzaaaozs

FIG. 37.——Curve showing the yearly percentage increments in weight of Boys. (From Minot, 1907.)

centage increment is as high as 200% (or nearly), but that then this increment drops to just over 20% at the end of the second year. From this point the decline is slow but sure, until at the thirtieth year the annual percentage increase is only 0-1 %. The change of rate of growth in females is practically the same as in males. The monthly percentage increment immediately before birth is about 20% according to Miihlmann’ s curve (Fig. 36) ; this represents an annual percentage increment of, say, 250 %, and the annual increase at the end of the first year is about 200 %. The postnatal deerease of growth is, therefore, as in other mammals, a continuation of the prenatal change. Further, there are two points at which the rate diminishes with great rapidity-—between the fourth and sixth months of pregnancy and between the ﬁrst and second years after birth. It would be of the greatest interest to discover the causes of these sudden decreases. Elsewhere the II. 2 GROWTH 65

diminution is gradual. A point of importance is that in both years there is a slight temporary rise in the growth-rate about the time of puberty (see the curve, Fig. 37'). This has been noticed by all observers, but the actual time of its occurrence ditfers in diEercnt cases ; the rise is invariably earlier in females than in males. A comparison of the growth of the three mammals considered is interesting. A Guinea-pig reaches 775 grammes in 43.2 days. A Rabbit ,, 2,500 ,, 395 ,, A Man ,, 63,000 ,, 9,428 ,, or the average daily increment is for a Guinea-pig L82 grammes. Rabbit 6-30 ,, Man 669 ,,

Hence ‘men are larger than rabbits because they grow longer, but rabbits are larger than guinea—pigs because they grow faster’. Minot, however, distinguishes between the ‘rapidity’ of growth, the average actual increment, and the ‘ rate’ of growth, the percentage increment. The average percentage increments for these mammals are

Guinea-pig 0-4-7

Rabbit 0-50

Man 0-02

The rate is, therefore, much slower in man than in the otln r two. These percentages Minot calls the coefficients of growth. Together with the duration of growth they determine the ultimate size of the organisms.

The progressive loss of growth-power Minot speaks of as ‘senescence’, and compares to the loss of the power of celldivision in the ‘senile decay’ of Protozoa. The same author has also brought forward evidence to show that during differentiation there is an increase in the amount of cytoplasm in the cell, a decrease in the size of the nucleus, and a decrease in the ‘mitotic index’, that is in the proportion, in any tissue, of dividing cells. During segmentation, of course, the reverse of this is taking place, since cell-division is rapid and the protoplasm per cell is being constantly diminished until a ﬁxed ratio between

umxixsox 1'‘ 66 CELL-DIVISION AND GROWTH II. 2

nucleus and cytoplasm is reached (Boveri) (see below, p. 268). Minot suggests that ‘ senescence ’ and differentiation alike depend on an increase in the protoplasm.

10 Z Aboral Arm-Length.

...... _- Oral Arm-Length. Body-Length.

Pluteus of Slrongylomrirotus lividus.

120 z 110 z 100 % 90 % so % 70 z 60 Z 50 z 40 z 30 z 20 z 10 %

Shell-length of Linmaea stagnalis.

Days. 10 15 20 25 30 35 40 45 50_ 55 60 65 70 75 80 85

_ FIG. 38.— Curves showing the change with age of the rate of growth In the larva of the sen.-urchin Strongylocentrotus (from Vernon's data), and the pond-snail Limnaea (from Semper's ﬁgures). The nbscissae are days, the ordinates percentage increments. II. 2 GROWTH 67

The decline of the growth-rate may also be seen in Pott's weighings of the Chick embryo before hatching (Table I) and Minot’ s Weights of young chickens. It appears from these that the daily percentage increment is 919 % at the beginning of the third day of incubation, 189% at the end of the fourth day; at this point there is a sudden drop to 40 Z, which is still the rate of growth after eleven days of incubation; eight days after hatching the ra.te is 9% in the male, not quite 9% in the female, and then comes a period of more or less gradual decline, until when the chicken is 342 days old it is able to add less than 0-5 % to its weight per diem.

Sempei-’s observations on the pond-snail, Limizaea, and Vernon’s on the sea-urchin, St1'0ngyloce7m'ot1m, are other examples which

FIG. 39.—Dai1y percentage increments of weight in tadpoles: the continuous line (a) gives the whole weight, the broken line (b) the dry weight. (After Davenport, 1899.)

may be mentioned. The results of these autl101's are shown in the accompanying charts (Fig. 38). Their measurements are of lengths, not of weights.

So far we have found no exception to the law of the decline in the rate of growth as development proceeds. Davenport's measurements of tadpoles will not, however, conform to the generalization. As the ﬁgure shows (Fig. 39), the daily percentage increments, whether of the whole weight or of the weight of dry substance only, ﬁrst rise abruptly, then descend and then rise again. An explanation of this anomaly may possibly be found in the fact that Davenport/s measurements are 68 CELL-DIVISION AND GROWTH II. 2

taken during that early period when growth is due in the main to absorption of water, the other measurements (as may be seen from Tables I, II) during the later period when the percentage of water has already begun to decline and growth is effected by other means. '

It is_, of course, a commonplace of embryology that the growth of all the organs of the body does not occur at the same rate.

50 % 40 X Chest-girth. ~ - - ~- Stature.

'—-——- Weight.

30 Z

20%

10%

Years. 1 2 3 4 5 6 7 8 9 1011l21314151617l8l920

FIG. 40.«—C1n-ves showing the alteration during the ﬁrst twenty years

of life of the rate of growth as measured by weight, stature, and chestgirth in the human being (males). (Constructed from Quetelc-t’s data.) The abscissae are years, the ordinates percentage increments. (The percentage increment of weight for the ﬁrst year could not be included in the figure. It is given in Fig. 37.) There are nevertheless few cases in which the exact difference in rate has been ascertained. From those few cases, however, it appears that the individual parts, though they do not grow with equal rapidity, still obey the same law as the whole.

Thus human stature exhibits the same loss of growth-power

as is shown by the weight of the whole body, with this 11. 2 GROWTH 69

difference, however, that the rate is not so high in early stages, the descent in later stages less abrupt. This will be seen in Table IV, in which such ﬁgures as are obtainable for the prenatal growth-rate are given, and in Fig. 40, in which the curve of change of growth-rate in human stature has been constructed from Q.uetelet’s data (male Belgians). The percentage

in % l l I I '. I -10 Z I[e:ul-Length. ll — — — — — —- Leg-Lengtln. I -—j—- Stature. I - - — - - -- Vertebral Column. I '3oz- ‘l I l I I 1 20 %-

“1jI—l"I“T"'T%—l'_‘ Y(':l1'S. I 2 3 4 5 I‘) 7 S 9 1011]2l314151(';1718l‘.)‘.’.0

FIG. 41.— Curves showing the alteration during the first twenty years of life of the rate of growth of stature, length of head, length of vertebral column, and length of leg in the human being (males). (Constructed from Quetelet’s data.) Ordinates, percentage increments; abseissae, years.

increment in the first year is only 39-6 as against 190-3 for weight, in the second year 13-3 us against 22-2 for weight. Thenceforward the rate slowly declines, until at the fortieth year it is zero, and after the ﬁftieth year increasingly negative. The rate of increase of stature is always slightly less than that of 70 CELL-DIVISION AND GROWTH II. 2

weight. Q.uetelet’s ﬁgures do not show the rise in rate about the time of puberty, but this phenomenon is apparent in the data furnished by Bowditch, Boas, and Roberts (see Fig. 4-2). The change in the growth-rate is practically the same in women as in men. As with weight, the rise of rate at the time of puberty is earlier.

The decline in the growth-rate of chest-girth is shown in the same ﬁgure (Fig. 40). It will be noticed at once that in this case the drop in the first year is very great indeed, from nearly 50 % to nearly 5 %, and that the rate is only diminished by another 24 ‘Z in the next nineteen years. The weight will depend upon the volume and the volume on both stature and girth; in. fact a rough weight-curve might be constructed from the measurements of stature and girth. It is evident that the sudden loss in the rate of total growth after the first year is due to the very rapid decrease in the percentage increment of girth.

It maybe mentioned that other measurements of girth—girth by the sternum, the waist, the hips, the neck, the biceps, the thigh——show the same exceedingly abrupt decrease, almost to the minimum rate, between the ﬁrst and second years.

In other cases——distance between the eyes, width of mouth, length of hand, length of foot, arm-length, leg-length, length and breadth of head, distance from the crown of the head to the first vertebra, length of the vertebral column—the change is more gradual ,- the rate of change, however, diifers in different cases. As an instance of this let us consider the measurements—— from the crown to the ﬁrst vertebra, the length of the vertebral column, and the leg-length——whieh together make up the total stature. The growth-curves of these three and of the whole stature are presented in the ﬁgure (Fig. 41), from which it will be seen that the growth of the leg is faster than that of the vertebral column (until the eighteenth year), and this than that of the head. Increase in stature takes place at nearly the same rate as that of the vertebral column , but is on the whole a little faster.

There are few cases—besides man—in which we possess information as to the growth of the parts. In the sea-urchin, S57'07I_q,I/[Odell/7'0tl(8, Vernon has shown that the growth-rate of the oral and aboral arms of the Pluteus diminishes rapidly from II. 2 GROWTH 71

the third to the fifth days, more slowly from the ﬁfth to the eighth days. After this the rate becomes negative, as the skeleton of the Pluteus is used up by the developing urchin. The curves of change of rate of growth-—as constructed from Vernon’s ﬁgures—-are shown in the chart (Fig. 38).

In Oarcirzm mamas a gradual decrease in the growth-rate of the frontal breadth can be ascertained from Weldon’s data.

We have next to consider another feature of growth, the alteration of variability. The facts at our command are derived from a study of Echinoid larvae (Vernon), Duck embryos (Fisehcl), Guinea-pigs (Minot), the Periwinkle (Bumpus), the Crab, C'arcz'mts (Weldon), and the human being (Bowditch, Pearson, Robeits, and Boas). Vernon has shown that in the Pluteus of Stronyy/ocentrolus the variability of the body-length increases regularly up to the fifth day, and then decreases regularly again to the sixteenth day. So Fischel’s measurements of Duck embryos seem to establish a greater variability in younger than in older stages. This is true of the whole length, the head (as far back as the first somitc), the hand and trunk together, and the total length exclusive of the primitive streak. The data are, however, too few to be treated statistically; the variability can only be roughly estimated from the extent of the limits within which the part varies at each stage.

Minot, who expresses the variability of guinea-pigs by the difference between the mean weight and the mean weight of the individuals above, and of those below, the mean, likewise ﬁnds that the range in variation diminishes with age, and further that, in the case of the males, there is a pcriod—-from about the fourth to the ninth months—when the variability is very much less than at any other time. No such sudden fall is observed in the female, only a steady diminution. ‘

A more satisfactory calculation of the altera.tion of variability may be made from the measurements taken by Bumpus of the ‘ventricosity’ (ratio of breadth to length) of the shell of the Periwinkle, Lit/orina lit/area. The series of observations is very large, and includes both British and American forms. In the accompanying table (Table V) the coeflicients of variability (the standard deviation expressed as a percentage of the mean) are 72 , CELL—DIVISION AND GROWTH 11.2

given for each age, as determined by length of shell, for the English and American periwinkles separately and also for the complete series. It will be seen that the variability increases slightly and then diminishes again. This is the case also in the American examples, where the fall at the end of growth is greater still, but in the British specimens there is only a slight fall, at 20-21 mm., followed by a considerable rise. The possible signiﬁcance of this diiference in the behaviour of the same species on the two sides of the Atlantic we shall discuss

in a moment. TABLE V Showing the alteration in the variability of the ventrieosity of the shell of the Periwinkle (Liltorina littorea.) during growth.

Coefficient of variability (E; x 100).

Length in mm. British. American. All. -15 2-77 3-27 3-25

16-17 2-94 3-41 3-34 18-19 3-02 3-39 3-35 20-21 2-93 3-03 3-13

22 3-25 2-84 3-02

In the meantime let us consider another case, the Crab, Cr/rcimc.v mocnas, the variability of the frontal breadth of which was examined by the late Professor VVelden. Weldon found that the variability, as measured by the quartile error , first increased and then suddenly diminished with age (as determined by carapaee length). If the variability is measured by the coefficient of variability (easily calculated from VVel(lon’s data) the result is the same. This will appear from the table

(Table VI). TABLE VI

Showing the change in the variability of the frontal breaulth with age in Curr-inns momms. (After Weldon.)

Cara ace Len th Q. 0'

E, ,,,m_ 3 51- x 100. 7-5 9-42 1-64 8-5 9-83 1-76 9-5 9-51 1-73 10-5 9-58 1-78 11-5 10-25 1-93 12-5 10-79 2-06 13-5 10-09 1-95

For the calculation of the variability at different ages in man data have been provided by Roberts, Bowditch, and Boas. Some II. 2 GROWTH 73

of these results are collected in the following table (Table VII), from which it may be gathered that the variability diminishes at first, then rises until it attains a maximum at about the time of puberty, and then diminishes again, reaching ﬁnally a value which is lower than the original. The values for the coeﬂicient obtained by the diifercnt investigators are fairly similar, and agree very well with those first given by Pearson (for male new—born infants, weight 15-66, stature 6-50; for male adults, weight 10-83, stature 3-66). It may be seen from the values for the newborn that the variability has already undergone adiminution before the age at which the other observations begin.

TABLE VII

Showing the change in the value of the coeﬁieient of variability in the male Human being during growth.

Coeffioieiit of variability (1% X 100).

Vtleight. Stature. Years. Boston Worcester, English Boston Toronto \Vorecstor, (Bowditch). Mass. Artisans (Bowditch \. (Boas). Mass. (Boas). (Roberts). (Boas) 4 14-00 5 11-56 11-48 4-76 4-82 6 10-28 12-04 10-08 4-60 4-34 5-40 7 11-08 11-87 10-29 4-42 4-35 4-24 8 9-92 11-83 10-78 4-49 4-58 4-32 9 11-04 12-29 10-85 4-40 4-41 4-30 10 11-60 12-92 11-06 4-55 4-68 4-44 11 11-76 14-45 11-90 4-70 4-53 4-51 12 13-72 15-56 11-48 4-90 4-85 4-49 13 13-60 18-07 11-76 5-47 5-36 5-21 14 16-80 16-80 12-74 5-79 5-64 5-43 15 15-32 18-28 14-00 5-57 5-71 5-19 16 13-28 13-95 12-95 4-50 3-92 17 12-96 11-23 11-55 4-55 3-32 18 10-40 12-18 3 69 19 10-29 20 9-03 10-50 10-92 12-04

Further, the variability does not merely diminish as the animal grows older. Its diminution accompanies the diminution in the rate of growth, and when—as at the time of puberty in man—that rate increases, the variability increases too.

The variability of such parts as have been examined for the 74 CELL-DIVISION AND GROWTH II. 2

purpose alters in the same way as that of the whole body. Besides weight and stature Boas has recorded measurements of height sitting, head-length, head-width, length of fore-arm, and hand-width.

Though the evidence, it must be admitted, is scanty, it is none the less a remarkable fact that in all the cases we have examined the variability, whether of the whole organism or of its parts, decreases with the decrease in the rate of growth. We seem to be in the presence of a phenomenon of general occurrence, though what the signiﬁcance of the phenomenon is is not at present clear.

As is well known, Weldon has argued that the decline in the variability of the older crabs is due to a selective death-rate, an argument which is supported by the same author’s observations on the snail Clausilia, since in this form the variability of the adult was found to be the same as the variability of the same individuals when young, but less than the variability of the general population of young. It is possible that the marked decrease in the variability of the American as compared with the British periwinkles may also be attributable to the same cause, since this animal has only recently been introduced into America, and may, therefore, be subjected to a more severe struggle for existence in its new environment.

It is doubtful, however, whether this explanation will ﬁt all cases.

Vernon has suggested that at periods of rapid growth the effect produced upon the organism by a change in its environment must be much greater than at other times, and, since he has further shown that one of the effects of an adverse change of circumstances is an increased variability, he argues that an increase in variability would naturally accompany a high growthrate.

Lastly, Boas points out that the rate of growth is itself a variable magnitude, and this ‘ variation in period’ may, with other causes, be a factor in producing the actual variation at each stage. Should that be so, the variability would necessarily increase and diminish with an increasing and diminishing growthrate, since those that are above the mean would tend to remove II. 2 GROWTH 75

themselves further from the mean than those that are below could approach it, and the more so the faster they were growing, and conversely.

We have ﬁnally to consider very brieﬂy what little is known of the alteration with growth in the value of the correlation between various organs. Such data as we have indicate that, like variability, this value rises and falls with the growth-rate.

Boas has ascertained the correlation coefﬁcicnt (p) in man between weight, stature, height sitting, length and width of head at diiferent ages. Some of these results are tabulated below (Table VIII) ; from this table it will be evident that the value of p decreases, increases, and decreases again. The values are for girls, and the period of increase is earlier than that found for boys. In the chart (Fig. 42) are given the successive values of growth-rate (stature), variability (height), and correlation coefficient (height sitting and head-length) for boys; the three magnitudes rise at about the time of puberty, and subsequently decline together. Boas urges that if the actual variability is in part the eﬁ’cct of variation in period, this eifect will be greater during periods of rapid development. It follows from this that if the various organs of the body are equally affected by a change in the growth-rate, correlations would be closer during periods of rapid growth than at other periods.

TABLE VIII

Values of the correlation coefficient, p, during growth for four diﬂbrent correlations. Girls, Worcester, Mass. (Boas).

Years. Stature and Stature and Stature and Stature and Weight. Height sitting. Length of Head. Width of Head. 7 -73 -74 -30 -21 8 -76 -79 -36 -15 9 -80 -82 -35 -16 10 ~83 -83 -37 -16 ll -81 -84 -37 -25 12 -77 -82 -38 -27 13 -73 -83 .38 -37 14 -67 -82 -30 -25 15 -6!’) -79 -26 -22 16 -60 -74 -25 -10

It will be noticed that the value of p for the diﬁ'erent organs is different, being greater between axial organs—stature and height sitting, stature and length of head-—than between longi76 CELL-DIVISION AND GROWTH II. 2

tudinal and transverse parts, such as stature and width of head. The correlation between stature and weight is liigli.

To whatever cause it may be due this diminution of correlation with age is of the greatest interest, since it points to an increasing power of self-diﬁerentiation in the parts of the body. From other sources also there is evidence of a. progressive loss

.10 Correlation (p). -30 Height sitting and length of -90

head. .10 6 5 Variability 4 (I — 100 (M X ) 3 Stature. 0

Rate of Growth 4 A (Percentage 3 7 increments of ° stature). 3 Z

12

l "T |‘"’l""'l'jl—‘__l'_—‘l—""lj'l Years. 7 s 9 10 11 12 1:; 14 15 16

FIG.42.-—'Figu1'e to show how the rate of growth (percentage increments of stature), the variability (of stature) and the correlation coefficient (between height sitting and length of head) rise together at the time of puberty in man and then fall together. (Constructed from

the tables of Boas.)

of totipotentiality of the parts, of an inereasing- independence of the parts, of a tendency to be increasingly governed in their development by factors that reside wholly within themselves. But this evidence must be discussed elsewhere. II. 2 GROWTH 77

LITERATURE

F. BOAS. The growth of’ Toronto children, U.S.A. Report of the Commissioner of Ed'ucatz'on, ii, 1897.

F. BoAs and C. WISSLER. Statistics of growth, Unitecl Slates Education Commission, i, 1904.

H. P. BOWDITCII. The growth of children, Massachusetts Sfale Board of Iloalih, 1877.

II. C. BUMPUS. The variations and mutations of the introduced Littorina., Zool. Bull. i, 1898.

C. B. DAVENPORT. The role of water in growth, Proc. Boslou Soc. Nat. Hist. xxviii, 1899 (1).

C. B. DAVENPORT. Experimental morphology, New York, 1899 (2).

A. FISCIIEL. Ueber Varhibilitiit und VVa.ehstl1um des embryonalen Kéirpers, lllurph. Jahrb. xxiv, 1896.

G. KRAUS. Ueber die Wa.sse1-vertheilung in der Pﬂmize, Fests-rhr. Fem‘ Iuuulertjiilu-. Best. Natmf. Ges., Halle, 1879.

C. S. MINOT. Seneseence and rejuvenation, Jouru. I’h_2/s. xii, 189].

C. S. MINOT. The problem of age, growth and death, Pop. Sci. Jllonthlg/, 1907.

K. PEARSON. Data. for the 1)l‘Ol)1el1l of evolution in man. Ill. On the magnitude of certain coeﬂicients of correlation in mam. 1’rac. Roy. Soc. lxvi, 1900.

W. PREYER. Spezielle Physiologie (les Embryo. VIII. Dns embryona.le Wachsthum. Leipzig, 1885.

A. QUETELET. Anthropometric, Bruxelles, 1870.

C. ROBERTS. Manual of a.nthropomet1'y, London, 1878.

K. SEMPER. Animal Life, 5th ed., London, 1906.

H. M. VERNON. '1‘he effect of environment on the development of Echinoderm larvae : an experimental enquiry into the causes of variation, Phil. Trams. Roy. Soc. elxxxvi, B, 1895.

II. M. VERNON. Variation, London, 1898.

W. F. R. WELDON. An attempt to measure the death-ra.te due to the selective destruction of (larcinus mamas with respect to a, pu.rticul:|.r dimension, 1’roc. Roy. Soc. lvii, 1894-5.

W. I‘‘. R. WELDON. A first study of na.tura,l selection in (L'luu.x-iliu Imm'nalu, Biometrilca, i, 1901-2. CHAPTER III EXTERNAL FACTORS

1. GRAVITATION

IN the large majority of cases there is no definite relation between the vertical and either the axis of the egg, the planes of its segmentation furrows, or the position of the development of the embryo in it. Thus the eggs of insects are laid with the axis making any angle with the vertical, and the same may be said of Crustacean ova. In such eggs as develop freely in the sea (some Mollnsca, for example, Polyehaeta, Coelenterata, Ctenophora) the axis and the planes of segmentation undergo a perpetual change of position, and Oscar Hertwig has shown that in the eggs of Echinoderms there is no necessary ﬁxed relation between the direction of the planes of segmentation and the vertical. In these cases it is clear that the features of development referred to cannot depend upon the force of gravitation.

There are, however, instances in which it seems possible that the directions of the planes of segmentation—bearing as they do a constant relation to the axis of the egg-—may depend upon gravity, since the axis is normally vertical. It was Emil Pﬂiiger who in 1883 first brought forward experimental evidence to show that this was indeed the case.

It is well known that the yolk of the hen’s egg always turns over so that the germinal disc is uppermost, and the egg of the Frog, free to rotate inside its jelly membrane, invariably takes up a position with the black pole uppermost, the white pole below.

This property of the Frog-’s ovum is exhibited alike by the ovarian, the coelomic, the uterine, and the freshly laid egg, by the living egg and the dead egg, by the whole egg, and by portions that contain both kinds of egg-substance, the yolk and the cytoplasm, as Roux showed by ﬂoating eggs or fragments in a medium of the same speciﬁc gravity. It is simply due to the fact that in the spherical telolecithal egg the heavier yolk III. I GRAVITATION 79

is placed mainly on one side, while on the other the lighter protoplasm is more abundant, the yolk granules far smaller and more sparse. The distribution of these two substances determines indeed the axis about which the egg has a ‘ rotation structure ’ or is radially symmetrical. The symmetry is further marked by the disposition of the pigment and the position of the nucleus. The pigment is placed in a thick superficial layer in the protoplasmic portion, it extends over rather more, sometimes considerably more, than a hemisphere, for there is much variation in this respect, and its boundary is a circle whose plane is at right angles to the egg-axis—the line which passes through the centre of the egg, the centre of the pigmented portion or animal pole and the centre of the unpigmented portion or vegetative pole. There is also an axial, less-deeply pigmented plug in the animal hemisphere. The nucleus is placed axially, b11t excentrically, very much nearer the animal than the vegetative pole, in a pigment-free spot or ‘ fovea germinativa ’.

The egg is invested with a layer of jelly (muein), inside which it becomes eventually free to rotate. This, however, is not possible when the egg is ﬁrst laid, for the jelly is at that time closely adherent to it. In water, however, the jelly swells up, and a narrow cavity is formed in about three hours between it and the egg, and the egg then turns over until its axis is vertical. The formation of the cavity is much more rapid if fertilization (insemination) has taken place; in this case the egg turns over in half an hour. The rapid formation of the perivitelline ﬂuid is the first effect of insemination, and is due to some substance secreted by or accompanying the sperm, since the spermatozoon does not reach the egg for another quarter of an hour (0. Hertwig). A second effect is an alteration in the viscidity or cohesion of the egg-contents ; for while in the ovarian or uterine egg no alteration occurs apparently in the disposition of yolk, cytoplasm, and pigment, although the egg-axis may make any angle with the vertical, such an alteration is undoubtedly produced by gravitation (see below) after fertilization has occurred. Another effect noted by Roux is that fertilized (not merely inseminated) eggs turn over more rapidly in a medium of like speciﬁc gravity than do unfertilized. 80 _ EXTERNAL FACTORS III. I

The most important ﬁrst result of fertilization is, however, the replacement of the radial by a bilateral symmetry. About two or three hours after insemination a certain portion of the border of the pigmented area, crescentic in shape and extending; over about half its periphery, becomes grey by retreat of the pigment into the interior (Roux). The egg can now be divided into similar halves by only one plane, the plane of bilateral symmetry, which includes the axis and the middle of this grey crescent (Fig. 43).

Fm. 43.—Formation of the grey crescent in the Frog's egg (IE. femparuriu). A, B from the side; 0, D from the vegetative pole. In A, C there is no crescent, in B, D a. part of the border of the pigmented area has become grey.

The middle of the grey crescent is always diametrically opposite to the point of entry of the sperm (Roux and Schulze) ; the crescent has hence been held by Roux to be directly caused by that entrance.

The plane of symmetry, as we shall see in another connexion, becomes in most cases the sagittal plane of the embryo, since the dorsal lip of the blastopore arises in the region of the grey III. 1 GRAVITATION 81

crescent. This side becomes the dorsal side of the embryo, while the animal pole marks, approximately, the anterior end.

By complete disappearance of the pigment the grey crescent becomes added to the white vegetative area of the egg.

The foregoing account applies in particular to Rana temporaria and R. fusca; I{.11aZusm'n appears to be similar, but in R. escaZenta it is stated that the egg—axis is eventually not vertical but oblique (Fig. 44). It seems, however, doubtful whether this obliquity is not rather apparent than real. The grey crescent has apparently not been recognized as such—-the pigment is not so deep as in the other species—but included, nevertheless, in the white area, with the result that the centre of this, the definitive

FIG. 44.——Egg of Rana esculenta after fertilization, in its normal position with the axis oblique (‘?). A, from the side; B, from above; an’, egg-axis ; mm, plane of ﬁrst furrow. (After Korsehelt and Heider.) white area, has been confounded with the centre of the original unpigmented area or vegetative pole of the vertical egg-axis. (Compare Fig. 44 A with Fig. 43 B.)

As is well known, the planes of division during the ﬁrst few regular phases of segmentation bear a perfectly deﬁnite relation to the axis. The ﬁrst two, at right angles to one another, are meridional and therefore also vertical, the third furrows are parallel to the equator, therefore also horizontal; the furrows of the fourth phase are again meridional, and hence vertical, those of the ﬁfth once more latitudinal and horizontal. It is this obvious relation of the planes of cleavage to the direction of gravity which has raised the question whether there is not a causal connexion between the two, the question which Pﬂiiger attempted

mnxmson G 82 EXTERNAL morons 111. 1

to answer by experiments, performed, however, not on the eggs of the Frog but on those of a Toad, Bombinator igneus.

The close adherence of the unlaid egg to the glutinous jelly, which in its turn could be easily ﬁxed to some object, provided a simple method of keeping the egg in any desired position. The eggs were removed from the uterus, attached with the axis at various angles to the vertical to watch glasses and fertilized with just enough sperm-water to allow of development, but not enough to permit of the formation of the pcrivitelline space and rotation of the egg into the normal position with the axis vertical. In these forcibly inverted eggs it was found that the furrows of segmentation bore the same relation to the vertical as in the normal egg; that is to say, the first was vertical, the second vertical and at right angles to the ﬁrst, the third horizontal and nearer the upper pole, whatever the inclination of the egg-axis to the vertical, except in the extreme case where the white pole was exactly uppermost (180° of inversion), when segmentation did not occur at all. There was, however, no deﬁnite relation between the plane of the first (and therefore of subsequent) furrows and the original axis of the egg; the angle between this axis and the plane of the first furrow, as also that between the first furrow and the plane including the original and the actual vertical axes of the egg, might, it was found, have any value.

Except in a few cases, and where the white area was nearly exactly uppermost, these eggs gave rise to normal embryos. The upper smaller cells divided more rapidly than the lower ones, whether pigmented or unpigmented, and the blastula stage was reached; the dorsal lip of the blastopore appeared on one side a little below the (actual) equator, and the lower surface was covered over by the blastoporic fold in the ordinary Way. Only in the failure of the whole egg to rotate after the closure of the blastopore (owing to the close adherence of the jelly) and in the irregular pigmentation (according to the original degree of inclination) did these embryos differ from the normal. One other point is worthy of notice. In the majority of cases the dorsal lip of the blastopore, marking the sagittal plane, appeared on the unpigmented side and lay in the plane including the III. 1 GRAVITATION 83

original, now inclined, and the actual axis, or vertical line of intersection of the first two furrows. While, therefore, the cleavage planes are deﬁnitely related to the vertical but not to the original axis of the egg, the median plane of the embryo appears to be jointly determined by both.

From these experiments Pﬂiiger drew remarkable and farreachingconclusions. He conceived of the eggas beingmeridionally polarized, composed of a large number of rows of molecules placed meridional] y with regard to the original egg-axis. Each row is equivalent developmentally to every other row, but within the limits of each the molecules are of diﬁerent value, since one end, for example, is anterior, the other posterior. .Which of these equipotential rows shall lie in the sagittal plane of the embryo is decided by gravity, and by gravity alone. Similarly the vertical direction of the first two furrows, the horizontal direction of the third, is due to the operation of some general, though at present unknown, law, in accordance with which ‘ (lie Schwerkraft die Organisation beherrseht ’. 1

The original structure of the egg, on the other hand, has no deﬁnite relation either to segmentation or to the symmetry of the embryo, except, of course, in so far as the original axis together with the actual vertical axis determines its sagittal plane, the white side its dorsal side.

‘. . . das befruchtete Ei gar keine wescntliche Beziehung zu der spiiteren Organisation des 'l‘hieres besitzt, sowenig als die Sehneeflocke in einer wesentliehen Beziehung zu der Grosse und Gestalt der Lawine steht, die unter Umstiinden aus ihr sich entwickclt. Dass aus dem Keime immer dasselbe entsteht, kommt daher, dass er immer unter dieselben iiusseren Bedingungen gebracht ist.’ 2

It will certainly be agreed that so sweeping and revolutionary a dogma as this is in need of very substantial support; and though the facts as stated by Pﬂiiger are incontrovertible, as the repetition of his experiments has shown, it is unfortunate that he did not also take into consideration the internal changes occurring in his forcibly inverted eggs. The deﬁciency has been made good by Born. Like Pﬂiiger, Born found that in the

’ 1. C. i11fl'a, XXxii- p. 24. 3 I. c. infra, xxxii. p. 64. G 2 84 EXTERNAL FACTORS III. I

forcibly inverted eggs the cleavage planes had the normal relation to the vertical, but not to the egg-axis; he observed, however, that the first furrow was usually in or at right angles to the streaming meridian, the plane, that is, including ‘the original and the secondary vertical axes. The recent examination of 215 cases by the writer has shown that the ﬁrst furrow tends to lie either in, or at right angles to, or at an angle of 45° to this plane. Subsequent development was normal, and the sagittal plane coincided with the streaming meridian. The dorsal lip appeared on the white side, which is thus anterior (antc1'o(l0rsa.l). The examination of sections, however, showed that in these

FIG. 45.-—Seetions through forcibly inverted Frog's eggs. In A the egg has just been inverted, in B the streaming of protoplasm upwards and yolk downwards has begun. Both sections are in the streaming meridian or gravitation symmetry plane, including both the axis (a a’) and the vertical. bD, pigmented animal protoplasm; wD, unpigmented vegetative yolk. ct, animal pole; a’, vegetative pole; 1', pigment-free clear area; A-, nucleus; 1), superﬁcial pigment. (After Born, from Korsehelt and Heider.)

inverted eggs there had been a redistribution of the contents, the heavy yolk sinking to the lower side, the lighter protoplasm, with the pigment and the nucleus, and the spermatozoon rising to the upper side (Fig. 45). The movement is rotatory, the cytoplasm and yolk ascending and descending in opposite directions ; and it also takes place naturally parallel to, and in a similar manner on each side of, the plane in which the primary and secondary axes lie, hence known as the streaming meridian. That end of this plane towards which the protoplasm moves in its ascent, the end, that is, marked by the primary vegetative pole, III. I GRAVITATION 85

is anterior (more correctly, anterodorsal), for it is here that the dorsal lip of the blastopore appears; the opposite end is posterior (more correctly, postero-ventral).

The pigment moves with the cytoplasm; it is, however, unable to completely displace that yolk which remains at the upper surface in consequence of the greater viscosity of the superﬁcial rind, and here a ‘white plate’ or ‘grey patch’ is formed. Similarly, at the lower surface the pigment is not necessarily wholly displaced by the descending yolk.

There is one special case that may be noticed. When the inversion is complete (l80°) the yolk flows radially and peripherally away from the upper pole while the cytoplasm ascends in the axis.

Born’s observations make it perfectly clear that gravity rearranges the eontents of these inverted eggs, and so confers upon them a secondary structure like that of the normal, and symmetrical about a secondary axis which is likewise vertical. To this secondary axis the direction of elongation of the karyokinetic spindles, and consequently the cleavage planes, bears the same relation as in the normal egg; and there is certainly no more need to explain these directions by reference to gravity, to suppose, in fact, a causal connexion between the two, in the one case than in the other. The planes, indeed, may fall where they do simply because the mitotic ﬁgures elongate in the direction of least resistance (Pﬂiiger) or (O. Hertwig) in that of the greatest protoplasmic mass, or may be related, in some similar way, to the structure of the egg alone.

The point can only be determined by an experiment in which the directive inﬂuence of gravity is eliminated. This experiment has been made by Roux. The eggs were fastened in small vessels, at distances of from one to eight centimetres from the centre, to a wheel rotating continually about a horizontal axis, but so slowly (one revolution in from one to two minutes) that the centrifugal force developed was insullicient to make the eggs turn with the white pole outwards, and therefore negligible. The direction of the force exerted by gravity upon them from moment to moment was thus not constant. Of the eggs some were free to rotate inside their jelly, others were ﬁxed. To anticipate the objection that the plane of rotation, 86 EXTERNAL FACTORS III. I

the plane of the wheel, is constant, a third set were packed loosely in test-tubes, and so able to roll over one another in all directions as they fell from one end of the tube to the other with each revolution. The first furrow appeared in all these eggs at the normal time and it was meridional, as in the normal egg; similarly the second was meridional, the third latitudinal; but the egg-axes exhibited no deﬁnite relation either to the vertical or to the plane of the wheel. The eggs were allowed to continue their development on the apparatus, and gave rise to normal tadpoles.

From this experiment Roux drew the conclusion that it is not gravity which determines the direction of the planes of cleavage, and that gravity is not an indispensable necessity for the normal development of the egg of the Frog.

Incontrovcrtible though this conclusion appears to be on the evidence, it has nevertheless been disputed by certain embryologists, Sehulze and Moszcowski, the controversy between whom and Roux upon the subject has new extended over many years.

Schulze has urged (1) that eggs placed on such a machine do not develop normally, and that the rotation of the eggs in their jelly exactly compensates for the rotation of the wheel. With regard to the latter point Roux has replied that on this supposition the egg‘-axes ought to be, at any moment, vertical, which is not the case. To the first objection it is a sufficient answer that not only Roux, but subsequent investigators (Morgan, Kathariner and the present writer) have been able to produce normal tadpoles from such rotated eggs.

It may be noticed here that Kathariner has repeated Roux’s experiment with a slight variation. The eggs were kept constantly rotating, not in one but in an indefinite number of planes by a stream of air-bubbles passing through a glass vessel filled with water. Development was normal. This result does not differ materially from that obtained by Roux with the test-tube eggs referred to above, which has indeed been also independently corroborated by Morgan.

The criticisms of Moszcowski take a different form. This author urges that gravity always exercises an inﬂuence upon the egg in determining the bilateral symmetry of both egg and III. 1 GRAVITATION 87

embryo. The grey crescent which appears soon after fertilization and is regarded by Roux as a direct effect of this process, is supposed by Moszcowski to be produced by the action of gravity upon the egg-contents during the short interval before the perivitellinc space is formed and the egg able to turn over, to be comparable, in fact, to the grey area or white plate described by Born in his forcibly inverted eggs. Every normal egg, therefore, has a ‘gravitation’ plane of symmetry which later on becomes, as in inverted eggs, the median plane of the embryo ; nor are the eggs on the rotatory apparatus exempt, for it is held that the work of gravity can be accomplished on them even in the few moments before they are placed on the machine.

With regard to the latter point both Katharincr and Morgan have demonstrated that eggs kept in a state of perpetual rotation in all directions, from the very moment of insemination develop into perfectly normal, bilaterally symmetrical embryos, while Roux has replied to the first part of the criticism by pointing out that the grey area observed by Moszcowski was not the normal grey crescent produced by the entering spermatozoon, but the ‘white plate’ of Born due to the incomplete rearrangement of yolk and cytoplasm in an egg which had been quite unintentionally prevented from assuming its normal position. The grey crescent, indeed, Roux argues, could not possibly be due to gravity in a normal egg, for it does not appear until some time after the axis has become vertical.

There seems, therefore, to be little room for doubt that Roux’s original contention, that gravity does not determine the symmetry of the egg and embryo in the Frog, is correct, although it remains a result of considerable importance that this external factor may be made artiﬁcially to induce a bilaterality in the egg which is sullieiently strong to persist as the symmetry of the embryo.

There is one other matter of interest in this connexion. It is obvious, and has been experimentally shown by O. Hertwig, that a centrifugal force can replace gravity. On a. wheel rotated with sufficient velocity the eggs turn with their axes radial, their white poles outermost. If the velocity is great enough (145 revolutions a minute, radius from 24- to 32 cm.) the yolk is driven 88 EXTERNAL FACTORS III. I

inside the egg towards the vegetative pole, and the distinction between it and the protoplasm accentuated. The segmentation of such eggs is mcroblastic ; a cap of small cells is formed, a blastederm, resting upon an undivided, though nucleated, yolk, and these yolk-nuclei are large and irregular, resembling the giant nuclei of the large-yolked eggs of Elasmobranchs and other forms (Fig. 46). An experimental conﬁrmation is

FIG- 46-—S8gm6nt9:ti911 Of We thus aiforded of Balfour’s hypoFmgis egg under the Inﬂuence thesis tf 'v' rd on com arative

, P11 0).‘: d I)

of a centrifugal force (from KorSchelt and Heldelu after 0- Hert- grounds, that it is on the varying

wi ). The egg consists of a blas- - - _ togerm and an undivided yolk quantity of yolk that differences

(yolk-syncytium): Ich,blastocoel; in the segmentation of eggs prim, yolk-nuclei; d, yolk. marily depend_

If removed from the centrifuge in time, such eggs may continue to develop, though they frequently give rise to monstrosities

(Spina biﬁzla).

LITERATURE

G. BORN. Ueber den Einﬂuss der Schwerc auf das Frosehei, Arch. mikr. Auat. xxiv, 1885.

0. HERTWIG. Welchen Einﬂuss iibt die Schwerkraft auf die Teilung der Zellen ‘P Jen. Zeitschr. xviii, 1885.

0. HERTWIG. Ueber einige am befruchteten Froschei durch Centrifugalkraft hervorgerufene Mechanomorphosen, S.-B. lci1m'g. preuss. All-ad. Wiss. Berlin, 1897.

L. KATHARINER. Ueber die bedingte Unabliiingigkeit der Entwicklung des polar differenzirten Eies von der Schwerkraft, Arch. Em. Mech. xii, 1901.

L. KATHARINER. Weitere Versuche ﬁber die Sclbstdifferelizirung dcs Froschcies, Arch. Ent. Mech. xiv, 1902.

F. KEIBEL. Bemerkungen zu Roux’s Aufsatz ‘ Das Niclitiietliigsein der Schwerkraft fiir die Entwicklung des Froscheies ', Amzf. Anz. xxi, 1902.

T. H. MORGAN. The dispensability of gravity in the development of the Toad’s egg, Auat. Anz. xxi, 1902.

T. H. MORGAN. The dispensability of the constant action of gravity and of a. centrifugal force in the development of the Toad‘s egg, Anat. Anz. xxv, 1904.

M. MOSZCOWSKI. Ueber den Einﬂuss der Schwerkraft auf die Entstehung und Erhaltung der bilateralen Symmetric des Froscheies, Arch.

milcr. Anal. 11:, 1902. III. I GRAVI'1‘ATION 89

M. MOSZCOWSKI. Zur Analysis der Schwerkraftswirkung auf die Entwicklung des Froscheies, Arch. milcr. Amzt. lxi, 1903.

E. P1«‘LiiG1«:R. Ueber den Einﬂuss der Schwerkraft auf die Teilung der Zellcn, Pﬂiigeris Amh. xxxi, xxxii, xxxiv, 1883.

W. Roux. Ueber die Entwicklung des Froseheies bei Aufhebung der riehtenden Wirkung dcr Schwerc, Breslau drtz. Zeitschu, 1884 ; also Ges. Abh. 19.

W. RoUx. Bemerkung zu O. Schu1ze's Arbeit iiber die Nothwendigkeit, etc., Arch. Ent. Mech. ix, 1900.

W. RoUx. Das Nichtnothigsein der Schwerkraft fiir die Entwicklung des Froscheies, Arch. Ent. Mech. xiv, 1902.

W. ROUx. Ueber die Ursachen der Bestimmung der Hauptrichtungen des Embryo im Froschei, Auat. Auz. xxiii, 1903.

O. SCHULZE. Ueber die unbedingte Abhiingigkeit normaler tierischer Gestaltung von der Wirkung der Schwerkraft, Verh. Anat. Ges. viii, 1894.

O. SCIIULZE. Ueber die Nothwendigkeit der freien Entwicklung des Embryo, Arch. 1m'kr. Amtt. lv, 1900.

O. SCHULZE. Ueber das crste Auftreten der bilateralen Symmetric im Verlauf der Entwicklung, Arch. milrr. Anal. lv, 1900.

2. MECHANICAL AGITATION

The necessity of perpetual and violent agitation for the very numerous pelagic ova which are ordinarily exposed to the stress of wind and weather is well known to every zoologist who has attempted to rear such forms in an aquarium, and need not be further insisted on.’

There are also other eggs which require a small amount of movement. The Hen turns her eggs every day, and the operation has to be artiﬁcially performed in an incubator. Its omission leads to serious consequences, for, as Dareste has shown, the allantois sticks to and ruptures the yolk-sac in unturned eggs, the ruptured yolk-sac cannot be withdrawn into the abdomen, and the Chick cannot hatch out. Death may ens11e at an early stage.

A violent agitation of the IIen’s egg, on the other hand, is equally fatal.

Dareste subjected the unincubated eggs to violent shocks at the rate of 27 a second for varying periods (from %' hour to 1 hour). The percentage of monstrosities observed after three or four days of incubation was very high indeed, except when

‘ It seems probable that the principal value of the mechanical agita tion to the larvae is to prevent the Diatoms and Algae, of which their food consists, from sinking to the bottom. 90 EXTERNAL FACTORS III. 2

the eggs were placed vertically with the blunt pole uppermost, the blastoderm therefore resting against the shell membra.ne.

Marcacci has exposed the eggs, inside the incubator, to continual rotation for 48 hours. The eggs were fastened to horizontal and vertical wheels rotating 40, 80, and 60 times a minute. At the last mentioned rate of revolution, the direction of rotation was reversed half-way through the experiment.

Many of the eggs actually hatched out, but the chickens were feeble and liable to disease, and exhibited malformations of the muscles or skeleton. Others, however, died before hatching, in some cases at an early stage, and death seems to have been due to rupture of the vitelline membrane; this was always fatal. The vertical motion was, on the whole, more harmful than the horizontal, owing to the perpetual see-saw.

It may be noted here that Féré has succeeded in producing retardation and abnormality of development in the Chick by means of short exposures to sound-vibrations.

Mathews has shown that mechanical agitation —violent shaking in a test-tube—is sufficient to provoke development (artificial parthenogenesis) of the unfertilized eggs in As/arias, but not in /lrlacia (see, however, below, p. 124).

LITERATURE

C. DARESTE. Rccherches sur la production des monstruosités par les secousses imprimées aux ueufs de poules, Comptcs Rendus, xcvi, 1883.

C. DARESTE. Sur le role physiologique du retournement dcs oeufs pendant l‘incubation, Comptes Remlus, c, 1885.

C. DARESTE. Non velles rccherches concern-ant Pinﬂuence des secousses sur le gei-me de 1'wuf de la. poule pendant la période qui sépare la pontc de la mise en incubation, Comptes Remlus, ci, 1885.

C. DARESTE. Note sur 1‘évolution de l’embryon de la poule soumis pendant Pincubation in. un mouvenient de rotation continu, Comptes Ilumlus, cxv, 1892.

C. F1~':R£':. Note sur les differences des effets des vibrations mécaniques sur l'évo1ution de l’embryon de poulet suivant 1‘époque oil elles agissent, C. R. Soc. Biol. (10) i, 1894.

C. Fﬁné. Note sur Pincubation dc l'oeuf dc poule dans la position verticalc, C. R. Soc. Biol. (10) iv, 1897.

A. MARCACCI. Inﬂuence du mouvement sur le développement (les ceufs de poule, Arch. Ital. Biol. xi, 1888.

A. P. MATHEWS. Artiﬁcial parthenogenesis produced by mechanical agitation, Amer. Jomw. Phys. vi, 1901-2. 91

3. ELECTRICITY AND MAGNETISM

An external agent, to which all eggs are inevitably exposed, is the natural magnetism of the earth. No evidence has, however, as yet been brought forward that this agent exercises any directive inﬂuence upon them, although their development may be distorted by excessive exposure to it.

Thus Windle placed a number of Hens’ eggs between the poles of a large horse-shoe magnet. Over 50 ‘A of these, when incubated, gave rise to abnormalities, the area vasculosa being affected in most cases. No relation could be detected between the position of the egg in the magnetic ﬁeld and the kind of monstrosity produced.

In the case of Trout ova similarly treated a very high deathratc was observed, but this was attributed by the experimenter to the action of the electric currents set up by the running water between the poles of the magnet. VVealc electric currents had less effect.

Silkworms’ eggs, however, suffered no harm.

The effects of the electric current upon the eggs of Amphibia and Birds were tested by some of the older observers. Rusconi, Lombardini, and Fasola all found that the development of the Frog's egg could be accelerated by weak currents. Lombardini produced monstrosities in the ease of the Chick by this method. More modern experiments are due to VVindle, Dareste, Rossi, and Roux.

Windle observed a fairly high death-rate amongst Trout eggs exposed to the action of the current. Dareste has found a large percentage of monsters among embryos developed from Hens’ eggs subjected for from one to three minutes to the electric spark (12 cm. long from Bonnetty’s machine, 3-35 cm. long from a Rhumkorif coil). Development was, however, normal in the case of eggs placed for an hour in a Tesla’s solenoid traversed by a discharge of 500,000 periods a second. Rossi employed a continuous current passing through the eggs (of Salm/lamb-iua perspicilla/a) in the direction of the axis. Both yolk and pigment became aggregated at the animal pole, leading to the formation there of a grey raised area surrounded more or less 92 EXTERNAL FACTORS III. 3

completely by a furrow. When segmentation occurred the ﬁrst two blastomeres were unequal and detached; the vegetative hemisphere was hardly segmented at all in later sta.ges, the previous divisions having disappeared. The nuclei were affected in various ways, and the directions of the cleavage spindles altered. The capacity for resistance to these evil effects was noticed to increase as development advanced.

The polar area produced in these experiments recalls the polar areas observed by Roux in Frogs’ eggs exposed to a horizontal current, at right angles, therefore, to the axis. Alternating currents of 50 and 100 volts were employed. The eggs were fertilized two or three hours before the commencement of the experiment. In from fifteen to thirty seconds after exposure two polar areas appeared in each egg. The polar areas were turned towards the electrodes. They were marked, dotted in various ways, and flocked with white extruded drops of yolk, and separated by furrows from a middle or ‘equatorial’ zone, the width of which varied directly with the distance of the egg from the electrode, inversely with the strength of the current and the duration of exposure.

Unfertilized ova were found to react in the same way. So also eggs in which segmentation had begun, and in those cases where the furrow cut the equatorial zone obliquely, the two halves of the latter turned away from one another.

The polar areas appear too in eggs which are exposed in the ‘ morula ’ stage, each cell having in addition a polar area of its own. The latter, however, do not appear in enfeebled eggs, but only the former.

In the gastrula and later stages the reaction occurs, but less markedly.

None of the eggs which have been exposed to the current develops any further. They stick to the jelly, and consequently lose their power of rotation.

Similar results were obtained by the use of the continuous current (43 volts), but the anodic and the kathodie areas usually differed from one another in certain details.

It is important to notice that neither in these experiments, nor in another in which the eggs were placed inside a glass III. 3 ELECTRICITY AND MAGNETISM 93

tube surrounded by a coil, could any deﬁnite relation be satisfactorily made out between the direction of the ﬁrst furrow and that of the current. Indeed, though intrinsically interesting, the experiments throw no particular light upon the problem of development. Rather should they be classed with the investigations of Verworn and others upon the behaviour of Protozoa in the electric current, investigationswhich promise to contribute to the understanding of the structure and movements of living substanee. It may be noted here that Roux has himself produced these polar areas on such structures as the heart and gall-bladder of the Frog and other vertebrates.

LITERATURE

C. DARESTE. Rechcrches sur l‘iniluence de l’électi-icité sur 1'4.’-volution dc l‘embr_yon de la poule, Comples Bemlus, cxxi, 1895.

U. ROSSI. Sull‘ azione dell‘ elettricita nello sviluppo dclle uova (legli Anfibi, Arch. Ent. Mech. iv, 1897.

W. RoUx. Ueber die morphologisehe Polarisation von Eiern uml Embryonen durch den electrischen Strom, sowie ﬁber die Wirkung des eleetrischen Stroms auf die Riehtung (ler elsten Teilung des Eies, S’.-B. Icais. Alrud. IViss. Wien, ci, 1891, also Ges. Abh. 25.

B. C. A. WINDLE. On certain early ma.lformn.tions of the embryo, Jouru. Aunt. and Phys. xxvii, 1892-3.

B. C. A. W1NDLE. The effects of electricity and magnetism on development, Journ. Aunt. and Phys. xxix, 1895.

4. LIGHT

As Roux pointed out long ago in the case of the Frog, light exercises no directive inﬂuence upon the development of the ovum. Blane, indeed, has attempted to prove that the direction of the embryonic axis in the egg of the Hen may be made to depend upon the direction of the incident light-rays, but the experiments are hardly conclusive. The method employed was to blaeken the shell of the horizontally placed egg with the exception of one spot to right or left of the blastoderm. On this spot a beam of light was kept directed during incubation. In some cases, but not in all, the embryonic axis was found to deviate from its normal position at right angles to the long axis of the shell. Further, the head of the embryo might be turned towards or away from the source of light. There was 94 EXTERNAL FACTORS 111. 4

no relation between the amplitude of the deviation and the length of the exposure.

Nor are the processes of growth and differentiation necessarily affected in any way at all by the presence or absence of light, or by the kind of light to which the eggs are subjected.

Thus Driesch, who has experimented with the eggs of I9’o/Mme, P/auorbis, and Rana, maintains that neither red, yellow, green, blue, nor violet light has the slightest eifect upon the eggs during the early stages of segmentation and gastrulation, in what he calls the organ-forming period of development; and Loeb has asserted that the development of the embryos of the lish Fzmthzlus is as rapid in darkness as in the light, except that on the yolk-sac (not in the embryo) far fewer pigment-forming cells are produced.

Yung, on the other hand, has brought forward evidence to show that in later stages, at any rate, the embryos of the Frog react diﬁerently to lights of various wave-lengths, some of which are harmful, others, apparently, beneﬁcial.

Yung obtained his colours from solutions of fuchsin (red), potassium bichromate (yellow), nickel nitrate (green), bleu de Lyon (blue), and viole de Parme (violet). The colours, it may be noticed, are not absolutely monochromatic.

Freshly laid eggs of Rana tem17omria were placed under the inﬂuence of these lights. After one month, samples of the tadpoles were measured, with the following result in millimetres : TABLE IX Red. Yellow. Green. Blue. Violet. White. Length 2158 25-91 18-83 26-83 29-66 25-75 Breadth 4-83 5-58 4-16 5-75 6-83 5-25

The mortality in the green light was great. After two months the dimensions were as follows :—

TABLE X Red. Yellow. Green. Blue. Violet. White. Length 26-25 31-83 All ‘ 33-50 41-30 31-00 Breadth 6-00 7-50 dead. 8-00 10-16 7-33

All the tadpoles in the red light eventually died. White and yellow light gave the greatest number of perfect frogs, but, as will be seen, those in the violet were larger. They III. 4 LIGHT 95

were, however, less differentiated, for they did not acquire their hind legs so soon as did those in the white light. It may be mentioned, however, that when the tadpoles reared under these conditions are replaced in ordinary light and starved, those from the violet exhibit a. greater power of resistance.

Experiments with Rana ésczzlenta gave the same result. In this case the effect of darkness was also tried and found to be distinctly unfavourable. Thus after one month the lengths in darkness and white light were respectively 19-66 mm. and 23-10 mm., the breadths 4-66 mm. and 5-50 mm. ,- after two months the difference was intensiﬁed, the lengths being 21-50 mm. and 32-16 mm. , the breadths 7-16 mm. and 7-66 mm. The deathrate in the dark was exceedingly high.

The eggs and embryos of the Trout were likewise found by Yung to be highly sensitive to green and red light, while the larvae reared in violet hatched out rather more quickly than those from yellow, blue, or white light.

In an experiment on the eggs of I/inmaca stayizalie, due to the same investigator, the effect is measured by the time required for the young to hatch out, as the following table shows :—

TABLE XI

Light. Time to hatching in days.

Red . . . . . . 36

Yellow . . . . . . 25

Green . . The heart is formed, then death occurs. Blue . . . . . . 19

Violet . . . . . . 17

White . . . . . . 27

Dark . . . . . . 33

Green light is evidently fatal; development is retarded in red light, less so in darkness ; yellow has about the same clfcet as white light, while there is a considerable acceleration in blue and violet.

The relative effect produced by the various lights is as in the preceding experiments.

The results obtained by Vernon for Eehinoid larvae are, however, not quite consonant with this, as may be seen in the table (Table XII), where the colours are arranged in the order of the eﬁect they produce. It will be observed that yellow is 96 EXTERNAL FACTORS III. 4

more harmful than red, while green exerts about the same effect as blue (copper sulphate). The author states, however, that in two other experiments the larvae were entirely killed off by the green light though developing perfectly in the white. He also adds that in violet light no development was possible owing to the swarms of bacteria.

TABLE XII Percentage change of size.

Semi-darkness . . . . . + 2-5 ‘ Absolute darkness . . . . . — 1-3*

Blue (copper sulphate) . . . . -46

Green . . . . . . . -4-8

Red . . . . . . . -6-9

Blue (bleu de Lyon) . . . . —-7-4

Yellow . . . . — 8-9

Almost within the limits of experimental error.

In the Pluteus Vernon found that both the oral and the aboral arm-length decreased in darkness, green and blue (bleu de Lyon) lights, while blue (copper sulphate), yellow, and red light exerted little inﬂuence on this magnitude.

It only remains to be added here that Blanc and Féré have brought forward some not very satisfactory evidence to show that white light is favourable to the development of the Chick. Féré has also stated that red and orange lights are more harmful than white, while violet has about the same elfect. The experiments are, however, vitiated by the fact that the eggs were not turned over.

LITERATURE

L. BLANO. Note sur l‘influence de la lumiere sur l'o1-ientation de l'embryon dans l’oeuf de poule, C. R. Soc. Biol. (9) iv, 1892.

L. BLANC. Note sur les effets tératogéniques de la lumiere blanche sur l’oeuf de poule, C. R. Soc. Biol. (9) iv, 1892.

H. Dnmscn. Entwicklungsmechanische Studien II, Zeitschr. wiss. Zool. liii, 1892.

C. Feat}. Note sur l'inﬂuence de la lnmiere blanche et de la lumiere colorée sur l’ineuba.tion des oeufs de poule, C’. R. Soc. Biol. (9) v, 1893.

'1‘. LIST. Ueber den Einﬂuss des Lichtes auf die Ablagerung von Pigment, Arch. Ent. Mech. viii, 1899.

J. LOEB. A contribution to the physiology of coloration in animals, Journ. Morph. viii, 1893.

J. Lorna. Ueber den Einﬂuss des Lichtes auf die Organbildung bei Thieren, Pﬂ12ger's Arch. lxiii, 1896. III. 4 LIGHT 97

H. M. VERNON. The effect of environment on'the development of Echinoderni larvae: an experimental enquiry into the causes of variation,

Phil. Trans. Roy. Soc. clxxxvi, 1895. E. YUNG. De 1’inﬂuence des milieux physiques sur les étres vivants,

Arch. Zool. Exp. et G'e'n. vii, 1878. E. YUNG. De Pinﬁuence des lumieres colorécs sur le développement

des aniinaux, Mitt. Zoo]. Stat. Neapel, ii, 1881.

5. HEAT

As is very well known, those activities by which every organism maintains its speciﬁc form can only be carried on within certain deﬁnite limits of temperature. So also a certain degree of heat is necessary for the due performance of the functions of growth and diﬁerentiation ; above or below certain limits—more or less definite for each organism, but varying in different organisms—-development is unduly accelerated or retarded, or brought to a standstill, while its form is frequently distorted as well.

’l‘o Oscar Hertwig we are indebted for a careful inquiry into the conditions of temperature under which the development of the Frog-’s egg takes place.

In the case of Ifanaﬂcsccz Hertwig has found the cardinal temperature-points to be as follows :—The normal is about 15°—16° C.; above this up to 20°—22° C. development is accelerated without being otherwise altered ; this temperature is therefore the optimum (Fig. 51). Above this point the form of development is altered, a11d at such a high temperature as 30° C. death follows very quickly. At low temperatures (6°-1° C.) there is considerable retardation, and at the zero-point a complete cessation of segmentation ; the eggs are often permanently injured.

At the high temperatures referred to—-from 23° C. upwards— it is the yolk—cells which are primarily affected. At from 29-6‘’ to 27-5" the yolk is unable to divide, though it is nucleated, and segmentation is conﬁned to the animal hemisphere, and soon ceases even there (Fig. 47). At 26-5° the first furrow indeed passes through the yolk, but subsequent segmentation is mereblastic, with the resulting formation of a cap of cells or blastoderm

lying upon and separated by a segmentation cavity from the

JEN xmson 11 98 EX'1‘ERNAL FACTORS 111. 5

nucleated yolk. The eggs then die. At lower temperatures25°-23°—the yolk is also affected, and many eggs die in the ‘ morula’ stage; such as do survive give rise to distortions or monstrosities (Figs. 48, 49). The injury to the yolk interferes

’9&o

, °\

A 11

Flo. 47.——Meridional sections of eggs of Rmmfusm developed (A) at 29-5° 0., (B) 26-5“ 0. Five hours ten minutes after fertilization. 7;, nuclei; p, pigment.

F100.’ 48.—Meridiona.l sections of eggs of Rama fusca developed at ._‘2 -5 C. Qne day after fertilization. /.-, nuclei; kk, blastocoel; 2, cells imbedded in unsegmented yolk.

FIG. 49.——Abnorma1 embryos of Rauafusca, produced by heat. A, embryo two days old developed at a. temperature of 24° C. ; 13, embryo three days old, reared at a. temperature of about 25° 0. br, brain; y.p, yo1k—plug; Lb, tail-bud ; t,ta.1l; s, sucker ; g, gill. III. 5 HEAT 99

with the proper closure of the blastopore ,- there is consequently a. large, persistent yolk-plug surrounded by a thickened blasteporic rim into which the separated halves of the medullary plate and notoehord are differentiated (spina biﬁda) (Fig. 50). The

FIG. 50.-~’l‘wo transverse sections through the embyro shown in Fig. 49 A. A, passes through the blastopore and yolk-plug; B, through the anterior end. (I, yolk-plug ; m.p, medullary plate; ch, notochord; mk, mesoblast.

front end of the arehenteron is, however, normally developed if the temperature is not too high, and in this case the anterior portion of the nervous system and notochord are undivided ; posteriorly, however, their right and left halves diverge round the blastepore, and are continued into the halves of the double tail when the latter is formed. Gill slits, protovertebrae, striated muscle—ﬁbres, the pronephros and its duct, and Fm.51.__Mel_idi0m1section

the tail {in may all be differentiated. of an egg of Rum, f,,_m,, de_

The development of the organs Veloped at 9: t01"Pe1‘°«t“1‘6 0f f t] t .d f t] b d . f_ 22° C. Six hours ﬁfty minutes 0 1e W0 S‘ es 0 1° 0 Y 1s 1c‘ after fertilization. kh, blaste quently unequal. coel.

At low temperatures segmentation and the closure of the blastopore take place very slowly, and at ()° cease altogether. The eggs are not, however, dead, but will resume their development when replaced under ordinary temperature conditions. They show abnormalities, however, due to injury of the yolk;

ll 2 100 EXTERNAL FACTORS III. 5

Morgan has similarly found that the fertilized (not, however, the unfertilized) eggs of an American species (If. palust/'18) which ve been subjected to a temperature of 1° C. and then allowed elop under normal circumstances exhibit spina bifida and

lze has also observed these abnormalities as the result of e cold. On one point, however, this author is not quite ement with O. Hertwig, for he states that eggs and s exposed to 0° in various stages do continue to develop,

thld of course very sl0\vly. ’.l‘hus, in the ease of eggs exposed sl8§ after fertilization, the blastula stage was only reached in

ays, while a month elapsed before the bl-astopore was formed. ie and Knowlton, however, state that in la’. rireecens and .4m0l_y.s*to1/ta liqrinum segmentation is totally inhibited at 0°. In another species of Frog (If. esculenla) which spawns much later in the year——in May and J une——the cardinal points were found by Ilertwig to be much higher, and the eggs endured a temperature of 33° C. without injury. They are, in fact, acclimatized to a higher temperature, and it is very interesting to notice that Davenport and Castle have succeeded in artiﬁcially acclimatizing the eggs of another Amphibian (Bag/‘o le7zli_qz'/zoszw) to a considerable degree of heat. Eggs were reared at 15° C. and 24°—25° C. After four weeks the heat rigor temperature was 40° C. for the former, 43-2° for the latter; and in another experiment the temperature was raised to 43-3’ by allowing the eggs to develop at 33°—34.«° for seventeen days.

A similar lowering of the minimum seems to have been observed by Lillie and Knowlton in the case of /lmM_y&lo1/za tigrimzi/I. In this form, which spawns much earlier than Rana vi7'c.vc¢m.v, there is considerably less retardation of development at 4°.

That temperature markedly inﬂuences the rate of development, or, as Hertwig puts it, that the quantity of developmental work performed per unit of time is a function of the temperature, is abundantly clear, and is well shown in the annexed diagrams (from Hertwig), in which the curves show the times taken to reach various stages at various temperatures (Figs. 52, 53).

It will be seen that as the temperature sinks the rate of development, or rather of differentiation, decreases, but at an The rate of growth, however, may increase at an increasing (or decrease at a) decreasing) rate, as Lillie and Knowlton found

FIG. 52.—Curves showing

development of the Frog (Ramtfusca). The abs '

ture in degrees Centigrade, the ordinates the

each of the stages I to ]X. I

dullnry folds closed, sucker

VII, operculum beginning; VIII, opercnlum closing; IX, rudiments of hind legs. (After 0. Hertw 98.)

s;I

EZIIIIIIIIIIIIIIIIIIIIIII .A.lIIIIIIIIIIIIIIIIIIII .. y . IIIIIIIIIIIIIIII EIIIII7 IIIIIIIIIIIII I

, g:1strula.; II, V. tail-bud ig, 18

I EEIEIWI%II%IIIIIIIIIIIIIII IIIIIIHIIIIIIIIIIIIIIIIIII IIIIIIIIIIHI I771 IEIIIIIIIII IIIIIHHIIIIIII IIIIIIIIIIKIIIHIIIIIHHIMII

IIEIIIIIIIIIIIIIIIHIIIIII II

ae giv ( '3 req inedullary plate; III,

V, tail

IIIIIHWMIIMHHIIDIIIIIIII II .I'ﬂIMﬂHIHIﬂﬂIIﬂIIIIII VIIIIHIIIHHIMIIHHMIII

1 HUIHIIIIIII "ii

the effect of temperature upon the rate of cuss e the tempera» <11) uired to reach

and gills; VI,

metail ﬁn ;

increasing rate. Lillie and Knowlton have made the same observation for the species investigated by them.

III. 5

HEAT

101 102 EXTERNAL FACTORS III. 5

for the tadpoles of Rana viresccns and Bufo lenliginoxzw. The same authors state that at low temperatures (below 3° in the case of the Frog, below 6° in the case of the Toad) growth was altogether inhibited, while at 2° there was an actual shortening in length in the ease of the Frog tadpole, due, it is suggested, to a diminution in the turgor of the cells.

The cardinal points have also been determined for the lIen’s egg. According to Kaestner normal development occurs only

FIG. 53.—Eﬂ'ect of teinperaturc upon the growth of the tadpole of the frog (Rana fusca). A, B, developed at a temperature of 14-5"--15" (1.; A, two days old, circular blastopore (Stage I in Fig. 52) ; B, three days old (Stage II in Fig. 52) ; C, 1), developed at a temperature of 20° C. ; 6', three days old (Stage V in Fig. 52); D, four days old (Stage VI in Fig. 52). (From Minot, 1907, after 0. Ilertwig, 1898.) between 95° and 102° F. (35° and 39°C.). The maximum, the temperature above which the embryo dies, is 43° C.; the minimum, at which development stands still, 28" C. Edwards, however, ﬁxes the minimum or physiological zero at 20°—21” C., for, as the annexed diagram shows (Fig. 54-),development may continue between 20° and 29°, though it is, of course, very much retarded.

Edwards has further made the highly interesting observation III. 5 HEAT 103

PERCENTAGE OF NORMAL DEVELOPMENT

FIG. 54.—~'1‘l1c index of; development (percentage of normal development) for the egg of the Hen at tenrperatures varying from 20°C. to 30-75"G. (After Edwards, 1902.)

AVERAG E

DIAMETEROF BLASTOIXRM IN MM.

_- p , ....I

 -%-Wﬁﬁ

‘ s=....'é.‘ns 2...

540 .ﬁt!m .... ,. III: ﬁ

p

E: 5.40 gig ii iii, Igga- UIIF I as? '*“-' 4.40 " 1......

RELATION OF TEMPERATURE T0 GROWTH OF BLASTOOERM

FIG. 55.—Growth of the blastoderm of the Hen’s egg independently of the appearance of the primitive streak, at low temperatures. (After Edwards, 1902.) 104 EXTERNAL FACTORS III. 5

that at tl1e low temperatures in question growth may occur without differentiation (Fig. 55). Thus in one series of experiments at 24-5° for six days the blastoderm increased in diameter from 4-4 mm. (the average diameter of the blastoderm in unineubated eggs) to 6-9 mm. The primitive streak was, however, not formed.

A temporary exposure to low temperatures often inflicts a permanent injury on the egg and leads to malformations. Kaestner, by subjecting the eggs at many different stages to temperatures of 15°—25°, 10° and 5°, has discovered that the capacity of resistance decreases as development proceeds (though not with absolute regularity). Thus the maximum exposure to 21° consonant with subsequent normal development was 192 hours for embryos of six hours, 96 hours for embryos of one day, 7 2.’ hours for embryos of two to six days, 4-9 hours for embryos of eight (lays, and 24 hours for embryos of 20 days.

At these low temperatures development is stated to be completely arrested, though the heart never ceases beating, irregularly and convulsively. The cooling process may be repeated over and over again without altering the capacity for future growth and diiferentiation, or reducing or increasing the maximum capacity of endurance of cold.

Malformations, as stated above, are of frequent occurrence, b.ut only in those cases in which the embryo has been exposed in an early stage, during the ﬁrst two or three days of incubation, and only after long exposures.

The Inedullary folds may remain unformed anteriorly, the two halves of the heart may remain widely separate, the head amnion fold may be absent and abnormal gill slits be formed; the heart and blood—vessels are often enormously distended, and hacmorrhages are frequent. Kaestner attributes these monstrosities not directly to the cold but to the pressure of the blastoderm against the shell, for in the cooled eggs, owing to some change in the speciﬁc gravity of the albumen or yolk, the latter rises up ; if the egg is placed with the blunt end uppermost, so that the embryo is pressed against the shcll—membrane, no monsters are produced.

Mitrophanow is another observer who has utilized low temperatures to cause malformations. High temperatures also give III. 5 HEAT 105

rise to abnormalities accompanied by acceleration (Féré, Mitrephanow).

The effects of extremes of heat and cold upon the ova and embryos of certain Invertebrates have been studied by Drieseh, the brothers Hertwig, Vernon, Sala, and Greeley.

FIG. .")6.—-The effect of heat upon the segmentation of the Echinoid egg. a, b, c, (1, four successive stages in the segmentation of the same egg of I9'¢'hinus; e, f, two successive stages in the division of the same egg of Sphaercchinus. (After Driesch, 1893.)

FIG. 57.— Suppression of cell-, but not of nuclear, division by heat (E:-hbms). (After Driesch, 1893.)

The ﬁrst-named observed that by subjecting the fertilized ova of Sp/zaerec/aiizus to a temperature of 30°—3l° (the normal is 19°) development was accelerated and segmentation abnormal (Fig. 56).

After the ﬁrst furrow—though not after subsequent divisions—— the blastomeres separated and sometimes remained apart, a fact which provided a means of watching the independent development of the ﬁrst two blastomeres. After the four-celled stage the direction of division became irregular, one spindle being 106 EXTERNAL FACTORS III. 5

perpendicular, instead of parallel, to the other three, or two perpendicular to two, or all irregular; in the next phase the formation of micromeres was partially or wholly suppressed. Nevertheless these abnormally segmented eggs produced perfectly normal Plutei.

It is also possible for nuclear division to continue while celldivision is suppressed, as a result of exposure to high temperatures (Drieseh) (Fig. 57).

FIG. 58.~—'l‘he effect of heat upon the development of S'pl1mu'e¢-/u'nu.s_r/ranulm'z's. a,exogu.strul-a; b, exog.1.strula.,wit.l1 tripartite gut; r, Pluteus, with tripartite gut; (d) Anenterion, with stoinodaeum, but no gut. (After Driesch, 1895.)

By exposing the blastulae to the same high temperature Driesch brought about a very interesting malformation, an Aneuterion (Fig. 58). The arehenteron was formed and constricted into the normal three portions, but it was evaginated instead of invaginated. Later on it shrank up and disappeared; the rest of the embryo, however, became a Pluteus, with a stomodaeum.

Vernon finds the optimum temperature for Echinoid larvae III. 5 HEAT 107

to be from 17-5° to 21«5° C. Exposure to high or low temperatures after fertilization, either for longer or shorter intervals or continuously, produced a decrease in body-length of the Plutei. The arm-length, however, increased with increasing temperature.

Vernon has also made the most interesting observation that the variability alters with the temperature. Eight-day larvae were measured, and the mean variability (Galton’s Q) of the body-length was found to be at 16° to 18°, 22-2, at 18° to 20°, 26-3, at 20° to 22°, 24-8, and at 22° to 24°, 24-0. Thus the variability is greatest at the temperature most favourable for development, and conversely.

It is also possible for the cell processes that occur during fertilization itself to be seriously affected by heat and cold, as the researches of O. and R. Hertwig have shown.

Moderate exposure (twenty minutes) of the eggs of Sta-ony_ylo— cenlrotmr to a temperature of 31° C. so weakens the cytoplasm that many spermatozoa are enabled to enter. Each sperm forms its own aster, and these combine with one another to form various irregular mitotic figures (triasters, tetrasters, and so on). The segmentation of such eggs is very irregular. With longer exposures the cones of entrance become feebly developed and the asters are not formed, while the numerous sperm-nuclei remain unaltered. Greater heat—over 40° C.— prevents the entrance of the spermatozoa altogether.

This pathological polysperm y may also be produced by cold; in this case also excessive exposure prevents the formation of the vitelline membrane, the cones of entrance, and the sperm-asters, while the spermatozoa remain in the peripheral layer of the egg. The eifect of a low temperature on eggs which have already been normally fertilized is seen in the reduction of the astral rays and spindle ﬁbres, though not of the spheres, and in the thickening and irregular aggregation of the chromosomes. At a normal temperature the achromatic figure reappears.

Very similar phenomena have been described by Sala. in A.9carz'8. This author kept the eggs (the females, that is, containing eggs in all stages of development) at low temperatures ——-from 3° to 8° C.—for from half an hour to ﬁve hours and longer. The eifect of a short exposure to a very low tem108 EXTERNAL FACTORS III. 5

perature is not so harmful as a longer exposure to a less degree of cold. The processes of maturation and fertilization were both abnormal. Granules of chromatin took the place of the tetrads and were unequally distributed to the spindle-poles ; or, if the chromosomes (tetrads) had been normally formed before the commencement of the experiment, their division was irregular, in extreme cases all passing to one pole and into the first polar body. Again, the formation of the polar bodies might be suppressed altogether, or abnormal, the second only being formed, or both as one, or the first polar body might be as large as the egg itself. 'l‘hc achromatic figure was also deformed, the spindle being split at one or both poles (pseudotriaster, pseudo-tetraster), and centrosomes appeared instead of the usual centrosomal granules. The cytoplasm became granular, the vitelline membrane was not formed, two or more eggs frequently fused together. Polyspermy, with consequent multiplication of asters and eentrosomes, was very noticeable, aml, in fertilization stages, a separate pronueleus may be formed from each female chromosome, or fragment of a chromosome.

Closely connected with the cytoplasmic effects brought about by these temperature changes is the phenomenon of artiﬁcial parthenogenesis, produced by Morgan and Greeley in Ar/mcia and .»1stc7-ins by lowering the tempe1'at111'e of the sea—water to the freezing-point. Greeley has also shown that a lowering of the temperature, like the raising of the osmotic pressure, results in a withdrawal of water, the cause to which, as is well known, Loeb attributed the development of unfertilized ova in his experiments.

Greeley has shown that by the combination cl‘ a low temperature with a chemical reagent :1. higher percentage of swimming blastulac can be obtained.

LI'l‘ERATURE

C. B. DAVENPORT and W. E. CASTLE. Studies in lnorphogencsis; III. On the acelilnatization of organisms to high temperatures, Arch. Em. Mesh. ii, 1896.

H. DRIESCH. Entwicklungsmeeh. Stud. IV: Experimentelle Veranderung des Typus der Furehung und ihre Folgen (Wirkungen von Warmezufuhr und Druck), Zeilschr. u-r'.vs. Zool. lv, 1893. III. 5 HEAT 109

H. DRIEscH. Entwicklungsmech. Stud. VII: Exogastrula und Anonteria, Mirth. Zool. Stat. Neapel, xi, 1895.

C. L. EDWARDS. The physiological zero and the index of development for the egg of the domestic fowl, Gallus do;msti('u.s‘, Amer. Journ.

Phys. vi, 1901-2. _

A. W. GREELEY. On the eﬂ‘ect of variations in the temperature upon the process of artiﬁcial parthenogenesis, Biol. Bull. iv, 1903.

O. HERTWIG. Ueber den Einﬂuss verschiedener Temper-aturen auf die Entwicklung der Froscheier, S.-I3. loom’;/. preuss. Al.-ml. Wis.»-. Berlin, 1896.

0. HER'rwI(}. Ueber den Einﬂuss cler Temperatur auf die Entwicklung von Rana fusca und esculcnta, Arch. mikr. Anat. li, 1898.

S. KAESTNER. Ueber kiinstliche Kitlteruhe von Hiihncreiern im Verlauf der Bebriitung, Arch. Anat. Phys. (Anna), 1895.

S. KAESTNER. Ueber die Unterbrechung der Bebriitung von Hiihnereiern a.1s Methode zur Erzeugung von Missbildungen, Verh. Amrt. GeseIIsr'h.,

1896. F. R. LILLIE and E. P. KNOWLTON. On the effect of tempemture on

the development of animals, Zool. Bull. i, 1898. L. SALA. Experimentelle Untersuchung ﬁber die Reifung und Befruehtung der Eier bei Ascaris megalocephala, Arch. milcr. Anat. xliv, 1895. O. SCHULZE. Ueber die Einwirkung niederer Temperatur auf die

Entwickelung des Frosches, I, Anat. Anz. x, 1895. O. SCHULZE. Ueber die Einwirkung niederer Temperatur auf die

Entwickelung des Frosches, II, Anat. Am. xvi, 1899.

6. ATMOSPHERIC PRESSURE. THE RESPIRATION OF THE EMBRYO

The respiratory exchange, which is so characteristic a function of adult organisms, is a necessity for the embryo also, and in some cases can be detected in very early stages indeed.

In the case of the Chick this need of oxygen is shown by the arrest or distortion of development, or the death of the embryo when the egg is placed in too conﬁned a space, or when the shell is varnished, wholly or above only (Mitrophanow, Féré), though a coat of varnish on the lower side has no effect according to the latter author; or again, when the egg is placed in an atmosphere of hydrogen, or when the pressure of the ordinary

atmosphere is reduced (Griaeomini). Giaeomini found that at a pressure of about 600 mm. the 110 EXTERNAL FACTORS III. 6

embryos were small and abnormal in respect of the medullary tube and amnion; the optic vesicles and cranial ﬂexure were absent, and there were serious disturbances in the area vasculosa, where, though the blood islands were present, the capillaries were either not formed or failed to reach the embryo. No haemoglobin was produced. Embryos exposed at a later stage (four days) nearly all died in two days of asphyxia, the blood being dark red and haemorrhages numerous. That these eﬁeets were due not to the reduced pressure but to the want of oxygen was shown by the complete normality of embryos reared in an atmosphere of pure oxygen at the same pressure (except in certain characters always exhibited by such embryos ; see below).

Similar methods may be employed to demonstrate the necessity of oxygen for the Frog’s egg, a necessity which is indeed patent to any one who has observed the inferior development, accompanied by spina biﬁda and open blastopore (Morgan) of the eggs in the middle of a mass of spawn.

Thus, according to Rauber, development is retarded at a pressure of % atmosphere of ordinary air, and the mortality high, while at pressures of 7}; or 7,1 atmosphere death very rapidly ensues. As a result of four days’ exposure to pure hydrogen or nitrogen (ordinary air from which the oxygen had been removed) Samassa observed retarded segmentation, and subsequently irregularities in development of the type already referred to. Carbon dioxide produced irregular segmentation and death in twenty hours.

Godlewski’s experiments are perhaps more thorough. The eggs subjected to ordinary air at a greatly reduced pressure (2 .mm.), as well as those kept in thoroughly boiled water, segmented but little, and cell-division was conﬁned to the animal hemisphere. In an atmosphere of pure oxygen at the same low pressure, however, development was, in many cases at least, neither retarded nor abnormal. Further experiments with pure oxygen, pure hydrogen, and an atmosphere composed of oxygen and carbon dioxide in equal parts, gave the same result, as the subjoined table shows (Table XIII). It is also clear that the absence of oxygen makes itself felt almost from the beginning, while pure oxygen accelerates development. III. 6 ATMOSPHERIC PRESSURE 111

TABLE XIII ~__ 1- Oxygen and : Hours Oxygen. Hydrogen. Carbon Controls. Dioxide. 3 First furrow in No furrow M-‘ No furrow seine 3% All but one with One - hall‘ Most with first first furrow with first furrow ' furrow , 4 All but one 4 cells The same, 8 All with 2 cells 2 cells ‘J3 5 All with 4 cells Most with 4 g Most with 21 cells; ce s 4» a few Wltl 4 17% Blastomeres smaller Blastomeres a, Normal than in controls smallerthan 3; in controls o 22} Blastomeres very Segmenta.- 7* Normal small tion ceases _ 47 Blastopore closed White) hemisphere visi le 73 Mcdullary folds Blastopore closed

This method has given similar results for the eggs of the fishes Cte7z0la6¢'u.s and Fmzriulus (Loeb). One or two points are, however, worthy of especial notice.

The former develops at the surface of the sea, and is more sensitive to a lack of oxygen than the latter, the segmentation of whose egg will indeed continue for twenty-four hours in pure hydrogen, though an embryo is never formed. In Cleuolabrus, on the other hand, segmentation never advances further than the eight-celled stage, and the cell-boundaries already formed subsequently disappear, though they can be restored on removal to pure oxygen. In Fvmrlulus the capacity for enduring a lack of oxygen decreases (or the need of oxygen increases) with the progress of development; the fatal exposure for a newly fertilized egg is four days, for a newly formed embryo thirtytwo hours, for an embryo with the circulation established twenty-four hours, and for the newly hatched larva. shorter still. Carbon dioxide is quickly fatal to both species.

The lack of oxygen has also a noteworthy eifect on the pigment cells which are found, especially round about the bloodvessels, on the yolk-sac of Fzmrlulus. These pigment cells are of two kinds, black and red, and when the embryo is deprived of oxygen the former disappear, the latter diminish only a little. 112 EXTERNAL FACTORS III. 6

It has been noticed elsewhere that this pigment is less abundantly formed in darkness than in light, and Loeb has suggested that light may promote oxidation.

The ova of Echinoids also require oxygen from the beginning of their development (Loeb). Without this element segmentation is impossible, or, if segmentation has already begun before they are deprived of it, the blastomeres swell up and fuse. According to Lyon the eggs of Arbacia are only sensitive to a want of oxygen for from ﬁfteen to twenty minutes after fertilization. Vernon has shown that water saturated with carbon dioxide and mixed in the proportion of 20 % or more with sea-water is fatal to the development of these forms.

Exact quantitative determinations of the oxygen absorbed and the carbon dioxide excreted have been made by Godlewski for the Frog and by Pott and Preyer for the Chick. The results are shown in the tables annexed (Tables XIV, XV).

TABLE XIV

Showing the result of one experiment on the respiration of the Frog's egg (Godlewski).

Day? anger A111ourz1llI;‘ilnpg§|;.air‘i)I(p:a‘:é;e:')f24 hrs. femhzatlom 0 absorbed. CO, excreted. 0-03908

1 __ 2 0-4502 0-0995 3 0-7033 0-2131 4 1-0539 0-4193

It thus appears to be the very general rule that the egg begins to respire at an early age. There is a case, however, A.s-crmlv, in which not only can the egg endure an atmosphere of nitrogen or carbon dioxide or nitric oxide for prolonged periods and still develop, but is actually killed by pure oxygen (at 2-} atmospheres) (Samassa). The adult worm, of course, is an endoparasite, and Bungc has shown that it can manage to produce carbon dioxide though denied access to free oxygen.

The eifect of pure oxygen has also been tried on various embryos. In such an atmosphere (at ordinary pressure) the development of the chick is normal, except that skin, allantois, limbs and amniotic fluid are all very red with oxyhaemoglobin; an excessive amount of carbon dioxide is produced. The amount of this gas III. 6

ATMOSPHERIC PRESSURE

113

excreted by the undeveioped though incubated egg in pure oxygen is, however, less than in air.

TABLE XV

Showing the oxygen a.bso1-bed and carbon dioxide excreted by the Hen’s egg during incubation. (After Pott and Preyer, from Preyer, Spez.

Phys.) Days of Amount in grammes per 24 hrs. of iIl0l1')Mi0l1- O absorbed. CO, excreted.

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According to Samassa. and Rauber the development of the Frog-’s egg in pure oxygen is normal, but Godlewski states, as

JENKINSON

I 114 EXTERNAL FACTORS III. 6

we have seen, that it is somewhat accelerated. At a pressure of 2% atmospheres, however, segmentation is arrested and death ensues (Samassa).

When the tadpoles, newly hatched, are exposed to its influence, the hyoid becomes immensely thickened and the branchial chamber completely closed; the internal gills a.re weak (Rauber), and the same author states that ‘ gastrulae’ subjected to three atmospheres of ordinary air had their development temporarily arrested, while later embryos, in the stage of the medullary folds, became small and immobile in air at twice the atmospheric pressure. This result seems to be due to pressure, not to the oxygen.

In the foregoing the general necessity of respiration for the life of the developing organism has alone been taken into consideration, but it should not be forgotten that oxygen may exert a stimulus on some part, the response to wl1ich results in a process of diiferentiation. Thus His has suggested that the growth of the blastoderm over the yolk is oxygenotropic, and IIerbst that the migration of the blastoderm-forming cells to the surface in Arthropod ova, and the migration of spiculeforming cells in Echinoid larvae are cases of deﬁnite reaction to oxygenotactic stimuli. Loeb, we may note, has found that the regeneration of the head of ’l'ubularia will only take place when the stem is supplied with fresh water, and the same author has suggested that the accumulation of the pigment cells round

the blood-vessels on the yolk-sac of Fmztlulus is also an oxygenotaxis.

LITERATURE

C. F1’-Jmé. Note sur l‘inllueuce des entluits particls sur l'ine-ubation dc l’u.:ul' do poule, C. If. Soc-. Biol. (10) i, 1894.

C. GIACOMINI. Inﬂuence de 1‘a.ir iarifié sur le dévcloppemcnt dc 1'ueufde poule, Arch. Ital. Biol. xxii, 1895.

E. GODLEWSKI. Die Einwirkung des Sauerstolfes auf die Entwicklung von Rana tempormia, Arch. Eut. Mesh. xi, 1901.

C. HERBST. Ueber die Bedeutung der Reizphysiologie fiir die cauale Auffassung von Vorgangen in der thierischen Ontogenese, Biol. Cmtralbl. xiv, xv, 1894, 1895.

J. LOEB. Ueber die relative Empﬁndlichkeit von Froschembryonen

gegen Sauerstoifmangel und Wasserentziehung in verschiedenen Entwicklungsstadien, Pﬂﬁgefs Arch. lv, 1894. III. 6 ATMOSPHERIC PRESSURE 1 15

J. LOEB. Untersuehungen ﬁber die physiologischen Wirkungen des Sauerstoffmangels, P_ﬂiige)"s Arch. lxii, 1896.

P. MITROPHANOW. Einﬂuss der veritnderten Respirationsbedingungen auf die erste Entwiekelung des Hﬁlmerembryos, Arch. Ent. Meclt. x, 1900.

R. Po'r'r. Versuche iibcr die Respiration dcs Hiihner-Embryo in einer Sauerstoffatinesphiire, If/h'iger’s 47071. xxxi, 1883.

R. Po'r'r und W. PREYER. Ueber den Gaswechsel und die chemischen Veri‘r_nderungen des Hiihnereies withrend der Bebrutung, Pﬂz'iger's Arch. xxvii, 1882.

W. PREYER. Spezielle Physiologic dos Embryo, Leipzig, 1885.

A. RAUBER. Ucber den Einﬂuss dcrTen1pc1-atur, des atinospliarischen Druckes nnd vcrschiedener Stoffc auf die Entwicklung thierischer Eier, S.-B. Nrmuf. Gas. Leipzig, x, 1883.

H. SAMASSA. Ueber die étusseren Entwicklungsbedingungen der Eier von Rana tomporaria, Verh. ])eutsch. Zeal. (v'cs. vi, 1896.

7. OSMOTIC PRESSURE. THE ROLE OF WATER IN GROWTH

That growth seems to depend in many cases on the absorption of water or a watery ﬂuid—in the swelling of the Echinoderm blastula, for example, or the enlargement of the Mammalian blastoeyst——has been noticed by several observers; in a. few instances experimental proof has been given of the relation between the two.

Although, as is very well known, the Hen’s egg loses weight daily throughout incubation by loss of water, this loss is due almost entirely to the slow evaporation of the albumen, and a humid atmosphere is necessary for development, as Pott and Preyer have found. Féré’s experiments with eggs incubated in desiccators demonstrated, during later stages, a slight reta.rdation accompanied by abnormalities and a high death-rate; in earlier stages, up to about the fourth day, there was on the contrary an acceleration of development.

Davenport has shown for tadpoles of various Amphibia (Amblystoma, Toads, Frogs) that increase in weight is very largely due to increase in weight of water. Known numbers of tadpoles, from which superﬁcial water had ﬁrst been carefully removed, were placed over sulphuric acid in a. desiccator. Re I 2 116 EXTERNAL FACTORS III. 7

peated weighings were made until a constant minimum was reached. The results are set forth in the accompanying table and ﬁgure (Table XVI, Fig. 59). It will be seen that the percentage of water rises very rapidly in the ﬁrst fortnight, from 56 % to 96 %, then decreases slightly, afterwards becoming nearly constant.

TABLE XVI Showing the rate of absorption of water by 'l‘u.«lpoles (after Davenport). Days after hatching. Percentage of water. 56

OOAH

ﬁx!-‘H>¢:~1U1L\:>v-4 to O‘

FIG. 59.-—Curve showing change in percentage of water in Frog tadpoles from the ﬁrst to the eighty-fourth day after hatching. Abscissae,

days; ordinates, percentages. (After Davenport, from Korschelt and Heider.)

A different, and a less satisfactory, method has been employed by Loeb, hypertonic solutions being used to prevent the absorption of water. While the newly fertilized eggs of Fimdulus III. 7 OSMOTIC PRESSURE 117

developed as normally in fresh water as in sea-water, only a blastoderm with occasionally a dwarf embryo was formed in a 5 % solution of sodium chloride in sea-water, and segmentation was arrested in the thirty-two-celled stage when the concentration of the salt was raised to 10 %. Older eggs were, however, far less sensitive, and after three or four days the embryos could he placed directly in a. 276 % solution without arresting their development, though the heart heat more slowly and differentiation was less rapid.

A I

FIG. 60.——A and 0, formation of ex-ovatcs in the egg of Arba('1'(( by dilution of the sea-water; ls, nucleus; m, egg-membrane; B and 1), hlastulae formed from A and C; If becomes constricted into two blastulae, each of which gives rise to a. Pluteus; D produces a single Pluteus. (After Loch, from Kolschelt and lleider.)

The eggs can nevertheless be aeelimatized to the salt. Removed from the 10% solution after the thirty-two cells had been formed to ordinary sea-water for eighteen hours, they were capable, when once more replaced in the strong solution, of giving rise to embryos which lived for a considerable time.

Similar experiments made on Arbacia showed that though cell-division is suppressed in the hypertonie solution (2 Z sodium 118 EXTERNAL FACTORS III. 7

chloride) nuclear division continues all the same, for when returned to seaewater the eggs divided at once into as many cells as had in the meantime been formed in tlie controls, a,result conﬁrmed by Morgan.

That the normal egg is in a condition of osmotic equilibrium with the sea-water is further shown by its behaviour in sea» \vater diluted to twice its volume ; in this experiment the egg

6‘

FIG. 61.—»Va.1-iations in the semnentation of Iu'¢-}u'nu.s- microtubm-culalns produced by dilution of the sea-water. a, tetrahedral four-eel] stage; II, eight cells, three premature niieromeres; 0, eight cells, two precocious microineres ; (I, the same egg afterthe next division, the precociolls micromeres have divided unequally, two normal micromeres have been formed. (After Driesch, 1895.)

(of /lrl/acia) absorbs water, swells and bursts its membrane and so produces a large ex-ovate which may develop independently of the rest of the ovum (Loeb) when replaced under ordinary conditions (Fig. 60). Driesch has produced irregularities of segmentation by the same means (Fig. 61).

Although, therefore, it seems reasonable to suppose that in the cases just quoted the observed eifects really are due to the III. 7 OSMOTIC PRESSURE 119

increased osmotic pressure of the medium and consequent withdrawal of water from, or prevention of imbibition of water by, the eggs, the weak point of the experiment, and of all such experiments, is our ignorance of the extent to which the ova or embryos are permeable to the substance employed, since the osmotic effect, or withdrawal of water, will obviously vary inversely with the permeability. The neglect of this possibly disturbing factor has indeed led in some cases to quite unwarrantable conclusions.

In 1895 O. Hertwig showed that certain abnormalities could be produced by growing the eggs of the Frog (IF. /‘urea and

FIG. 62.—Three sodium-chloride embryos of Rana fusca. df, yolkplug; hp, brain; Ici, gills; .9, margin of epidermic layer of ectoderm; sch, tail; m-, lip of blastopore, (After 0. Hertwig, from Korschelt and Heider.)

esculenta) and of the Axolotl in a solution of common salt. In stronger solutions (1 % to 0-8 %) segmentation was conﬁned to the animal hemisphere, though nuclear division went on in the yolk. Weaker solutions (0-6 %) allowed of further, but distorted, development; the yolk-cells were unable to move beneath the lip of the blastopore, so that the latter remained open with a persistent yolk-plug, and the mcdullary folds failed to close in the region of the brain, a condition recalling the abnormalities known in Human and Comparative Teratology as Hemicrania and Aneneephaly (Figs. 62, 65). The exposed region of the brain underwent a grey deg'enera.tion with dis— 120 EXTERNAL FACTORS III. 7

integration of the epithelium. Other organs were, however, normally formed, the front end of the gut by iiivagination, the notochord and mesoderin, protovertebrae, heart, pronephros, auditory vesicles, optic vesicles, infnndibulum, and liver, until the embryo died. The persistence of the yolk-plug has also been induced by Gurwitsch by means of halogen salts (sodium bromide and lithium chloride) and weak solutions of alkaloids (strychnine, caifein, nicotine) (Fig. 63), by C. B. Wilson in Hana, C/I0?'0j)/I27’IlS, and A7110/‘I/8/07ll(l F;G,53__Me,-idiom1sec. by means of sodium chloride and Egggruffilfﬁggélzff Ringei"s solution, and by Morgan ‘with 1,, blagtocoel. (After Gm-. Various lithium salts; and Bataillon, Witﬂchs from Korscllelt and who has used isotonic solutions of caneHeider.) . . sugar, sodium chloride, and a large number of other salts for the purpose, claims that in this case the results produced depend upon the osmotic pressure alone, and are therefore due to a withdrawal of water from the developing

embryo.

-B

' Fm. 64.—Sections of Frogs‘ eggs grown in solutions of, A, annnoniuin iodide (1-5%), and, B, urea (2-37,). In both cases segmentation is iiieroblastie, although in A there are a few lai'::,e. divisions in the yolk. In B the multinueleate cell masses of the animal hcniispliere protrude above the surface. The nuclei are large, lobed, and lioiiiogeneously clf'_Iroinal;ic in both cases. (Ammonia is probably present in the solution 0 urea.

Recent experiments made by the author do not, however, bear out this conclusion. In the first place, it is to be observed that isotonic solutions (isotonic with a O-625 % NaCl solution) do not III. 7 OSMOTIC PRESSURE 121

produce the same, but markedly diﬁerent effects. Some solutions arrest development at an early stage (during segmentation (Fig. 64), gastrulation, or the formation of the medullary folds); in others development proceeds but is distorted, the medullary folds remaining open in whole or in part, and the yolk-plug uncovered, or either of these malformations may occur without the other; in one case (dextrose) development is quite normal in form but very considerably retarded, while ﬁnally in urea development is normal both in form and rate (Figs. 66, 67). No legitimate deductions can be made from these experiments, how

FIG. 65.—1<‘rog embryos grown in a -625% solution of sodium chloride. A and B after ﬁve days, 0 and D after six days. In all the yolk-plug is fully exposed. In A the medullary groove is wholly open, in B and C it is closed behind, in D it is closed throughout. ever, until the permeabilities of the tissues to these solutions are ascertained. The tadpole requires water (Davenport), and the degree of shrinkage of the tadpoles in these solutions affords a means of determining the question ; it appears that they are perfectl y permeable to urea, more or less impermeable to cane-sugar, dextrose, and sodium chloride, the sllrinkage being rather greater in the first than in the other two. On the assumption that the permeabilities of the embryo are the same as those of the tadpole, it follows that the greater effect produced on the former by sodium chloride than by cane-sugar, or, still more, than by dextrose, cannot be set down to the osmotic pressure of the solution alone, 122 EXTERNAL FACTORS III. 7

a. result which is further corroborated by the constancy in the relative toxicities of the bases and the acids in the case of the

FIG. 66.——F1-og embryos grown in isotonic solutions of, A, sodium chloride (-625%); B, cane-sugar (6-6%); 0, dextrose (3-4%); and, D, urea. (1-14%). In A the medullary folds are closed but the blasto ore open; in B the medullary groove 18 open but the blastopore close ; in C development is normal, but retarded ; in D development is normal, both in form and rate, though the embryos die soon after the stage shown in the ﬁgure.

FIG. 67.—A. Longitudinal section of a Frog embryo grown in a -45% solution of lithium chloride. The medullary groove is open, except in front and behind. The notochord is bent in several ilaces and the gut roof much crumpled. B. Longitudinal section of a. ‘rog embryo grown in a. 6-6% solution of cane-sugar. The medullary groove is open, except in front, the cells in its ﬂoor degenerating. The gut roof is incomplete in part and there is an evident neurenteric canal.

monobasic salts. The observed deformities are therefore to be attributed to some other—chcmical or physical——property of III. 7 OSMOTIC PRESSURE 123

the solutions, though what this is is not known.‘ It may be added that in Gurwitseh's experiments the concentrations of the alkaloids employed were certainly far below those which would be isotonic with a -625% solution of sodium chloride. It also follows that during the closure of the blastopore the Frog’s eggdoes not need to absorb water from the outside; it may, in fact, be exposed to a very considerable degree of desiccation at this period without interfering in the least with the closure of the blastopore or of the medullary folds, a result which is all the more surprising in that the newly hatched tadpole imbibes water at so rapid a rate.

The experiments which have hitherto been considered relate to the need of water for normal development. There are, however, certain processes for which not the absorption, but, on the contrary, the abstraction, or at least the local abstraction, of water appears to be essential, the phenomena, namely, of fertilization. Cytologists have observed that the entrance cone and funnel, the mechanism by which the spermatozoon is swept into the interior of the egg, appear to be aggregations of a watery substance about the aerosome or apical body, and that the sperm sphere and aster are similarly due to the withdrawal of water by the centrosome in the middle-piece from the cytoplasm ,- in other words, that the stimulus whereby the spermatozoon restores to the egg its lost power of cell-division is essentially a process of local dehydration.

This inference is substantiated by the familiar experiments of Loeb, who has succeeded in rearing normal larvae from the unfertilized eggs of Echinoderms and certain worms by temporary immersion in certain solutions. In his earlier experiments he found that a mixture in equal parts of a 2,9 a solution of magnesium chloride and sea-water produced more Plutci than any other solution tried, and hence believed the result to be specific and attributable to the magnesium ion. Later, however, this artiﬁcial parthenogeuesis was successfully brought about by various isotonic solutions (chlorides of sodium, potassium and calcium, potassium bromide, nitrate and sulphate, cane-sugar

‘ In this view Stockard, as a result of experiments on Fmululus, concurs (Arch. Em. Mach. xxiii, 1907, and Journ. Exp. Zool. iv, 1907). 124 EXTERNAL FACTORS III. 7

and others). The increased osmotic pressure was, therefore, considered to be the cause of the phenomenon, and it was suggested that in ordinary fertilization the spermatozoon introduces a substance which has a higher osmotic pressure than, and is therefore able to withdraw water from, the egg.

Hunter has also shown that sea-water concentrated to 7 0% of its volume is sufficient to bring about the result. It must still be remembered that the permeabilities of the ova to the various solutions are not known; Sollmann, indeed, has proved the secondary swelling after the primary shrinkage of many eggs in hypertonie solutions, which must therefore enter and cause the dissociation of the cytoplasm.

Further, Delage has, as a matter of fact, denied that the increased osmotic pressure is solely responsible for the results. The French zoologist succeeded in making the ova develop in solutions hypertonie to sea-water, but found that isotonic solutions of different chlorides or mixtures of chlorides did not all give the same percentage of larvae. He holds, therefore, that other factors are involved. Other methods, as noticed elsewhere, are low temperatures and mechanical agitation.

Fischer has successfully demonstrated the phenomenon in the Chaetopods, Nereis and A7221;/zzhile, Bullot in 01)/(elm, and Bataillon in Vertebrates (lfamz. _/usca and I’eh-omyzon j/laueri); but in this last case segmentation did not continue for very long and the processes of nuclear division were highly irregular. An attempt made by Gies to incite development (of Echinoids) by means of extracts of spermatozoa was unsuccessful.

Although in brilliancy of conception and completeness of execution Loeb’s experiments are certainly pre-eminent over those of any other investigator, it should not be forgotten that about the same time Morgan had succeeded in inducing asters, and even the beginnings of segmentation, in the unfertili7.ed ova of sea-urchins and some other forms by the use of salts and other substances, and that the way for all recent work was really paved by the original labours of (). and R. Hertwig, to be described in the next section.

Loeb did not undertake an examination of the cytological changes, but Wilson has shown that ordinary nuclear division occurs with asters and centrosomes: a primary radia.tion III. 7 OSMOTIC PRESSURE 125

centring in the nucleus ﬁrst appears; this then fades away, and a deﬁnite aster with a centrosome is formed just to one side of the nucleus; this divides to form the ﬁrst amphiaster (cleavage-spindle). Asters also arise independently of the nucleus in the cytoplasm (cytasters) ; these contain centrosomes, and may divide, and the cytoplasm divide round them. The part played by the cytasters in development is, however, insigniﬁcant; their activity soon comes to an end. The number of chromosomes is one-half the normal number. This latter statement is conﬁrmed by Morgan, but denied by Delage, who asserts that, as in egg fragments enucleated and subsequently fertilized, the half number becomes doubled.

LITERATURE

E. BATAILLON. La pression osmotique et les grands problemes de la Biologie, Ar:-71. Ent. Mach. xi, 1901.

E. BATAILLON. Etudes expérimentales sur l‘évo1ution des Am-' phibiens, Arch. Em. Mecli. xii, 1901.

C. B. DAVENPORT. The rele of water in growth, Proc. Boston Soc. Nat. Hist. xxviii, 1897-8.

C. F1’«:Iu':. Note sur l’inﬂuence de la désliydratation sur le développement de Pembryon de poulet, C’. R. Soc. Biol. (10) i, 1894.

A. GURWITSCH. Ueber die formative Wirkung des verandertcn chemischen Mediums auf die embryonale Entwicklung, Arch. Eur. Mach. iii, 1896.

O. HERTWIG. Die Entwicklung des Froscheis unter clem Einﬂuss sehwiieherer und stéirkerer Kochsalzlesungen, Arch. milcr. Anat. xliv, 1895.

O. HERTWIG. Die experimentelle Erzeugung thierischer Missbild— ungen, Fcstsclzr. Gegenbaur, Leipzig, 1896.

J. W. J ENKINSON. On the effect of certain solutions upon the development of the Frog's egg, Arch. Ent. Mach. xxi, 1906.

J. LOEB. Investigations in physiological morphology, Journ. Morph. vii, 1892.

J. LOEB. Ueber die relative Empﬁndlichkeit von Fischembryonen gegen Sauerstotfmangel und Wasserentziehung in verschiedenen Entwicklungsstadien, Pﬂﬁger’.-c Arch. 1v, 1894.

J. LOEB. Ueber eine einfache Methode zwei oder mehr zusammengewaehsener Embryonen aus einem Ei hervorzubringen, P_/iiige;-'3 Arch. lv,

1894. LITERATURE ON ARTIFICIAL PARTHENOGENESIS

E. BATAILLON. Nouveaux essais do parthénogénése expérimentale chez les Vertébrés inférieurs, Arch. Ent. Mech. xviii, 1904.

G. BULLOT. Artiﬁcial parthenogenesis and regular segmentation in an Annelid (Ophdia), Arch. Ent. Mech. xviii, 1904.

Y. DELAGE. Etudes sur la mérogonie, Arch. Z001. E.I:p. et Gén. (3), vii, 1899.

Y. DELAGE. Etudes expérimentales sur la maturation cytoplasmique 126 EXTERNAL FACTORS III. 7

et sur la parthénogénese artiﬁeiclle chez les Echinodermes, Arch. Zool. Exp. et Gén. (3) ix, 1901.

M. A. FISCHER. Further experiments on artiﬁcial parthenogenesis in Annelids, Amer. Journ. Phys. vii, 1902.

W. J. Gms. Do spermatozoa contain an enzyme having the power of causing the development of mature ova ? Amer. Joum. Phys. vi, 1901-2.

A. W. GREELEY. Artiﬁcial parthenogencsis in the star-ﬁsh produced by lowering the temperature, Amer. Journ. Phys. vi, 1901-2.

A. W. GREELEY. On the analogy between the effects of loss of water and lowering of temperature, Amer. Journ. Phys. vi, 1901-2.

A. W. GREELEY. On the effect of variations in the temperature upon the process of artiﬁcial parthenogenesis, Biol. Bull. iv, 1903.

R. HERTWIG. Ueber die Entwicklung des unbcfruchteten Seeigeleies, Fesischr. Geyenbaur, Leipzig, 1896.

S. J. HUNTER. On the production of artiﬁcial parthenogenesis in Arbacia by the use of sea-water concentrated by evaporation, AlII('I'. Journ. Phys. vi, 1901-2.

J. W. JENKINSON. Observations on the maturation and fertilization of the egg of the Axolotl, Quart. Jam-n. Min-. Sci. xlviii, 1904.

K. KOSTANECKI. Ueber die Veriinderungen im Inneren des unter dem Einﬂuss von K01-Gemischen kﬁnstlich-parthenogenetisch sich entwickelnden Eis von Mactra, Bull. Inlrrn. Acad. Sci. Cracovie, 1904-5.

J. LOEB. On the nature of the process of fertilization and the artiﬁcial production of normal larvae (Plutei) from the unfertilized eggs of the sea-urchin (two papers), Amer. Journ. Phys. iii, 1899-1900.

J. LOEB. Further experiments on artiﬁcial parthenogem-sis, and the nature of the process of fertilization, Ame)‘. Journ. Phys. iv, 1900-1.

J. LOEB. Experiments on artificial parthenogenesis in Annelids (Chaetopterus), Amer. Journ. Phys. iv, 1900-1.

A. P. MATHEWS. Artificial parthenogenesis produced by mechanical agitation, Amer. Journ. Phys. vi, 1901-2.

T. H. MORGAN. The fertilization of non-nucleated fragments of Echinoderm eggs, Arch. Ent. Mech. ii, 1895-6.

'1‘. H. MORGAN. The production of artiﬁcial astrospheres, Arch. Em. Mech. iii, 1896.

T. II. MORGAN. The action of salt solutions on the unfertilized and fertilized eggs of Arbaciu, Arch. Ent. Mach. viii, 1899.

T. H. MORGAN. Further studies in the action of salt solutions and other agents on the eggs of Arbacia, Arch. Em‘. Me('h. x, 1900.

E. B. WILSON. A cytological study of artiﬁcial parthenogcncsis in sea-urchin eggs, Arch. Em. Mech. xii, 1901.

8. THE CHEMICAL COMPOSITION OF THE MEDIUM

By means of solutions of alkaloids and other substances the brothers Hertwig have been able to incite very remarkable cytological changes in the eggs of sea.-urchins (Stronyylocemfrotus). III. 8 CHEMICAL COMPOSITION 127

The effects of nicotine are perhaps the most striking (Fig. 68, a-e). Various solutions—-1 % and less of a concentrated extract——were allowed to act upon the egg for diﬁerent lengths of time (ﬁve to ﬁfty minutes) before fertilization; the ova. were then replaced in sea-water and fertilized. The cytoplasm is so paralysed by the

FIG. 68.~The effect of alkaloids and other poisons on the processes of fertilization and nuclear division in the egg of the sea—urchin, Sh'ong_:/Iocentrotus lividus. (After R. and 0. Hcrtwig, 1887.)

a. The egg was exposed to nicotine (one drop in 200 c.c. of seawater) for ten minutes, and then fertilized; drawn ﬁfteen minutes later.

b, c. The same for ﬁfteen minutes; drawn after one and a half hours.

d. The same for ten minutes; drawn after three hours, ten minutes.

2. The same; drawn after three hours. Only part of the complex ﬁgure is shown; the remainder lies in another plane.

f, g, h. Exposed to a 0-05% solution of quinine for twenty minutes one and a half hours after fertilization; drawn from one to two hours later.

is. 1-5, male pronucleus, 6, female pronucleus. Exposed to chloral (05%) one minute after fertilization; ﬁxed after 150 minutes.

I, m. Chloral 06% one minute after fertilization; fixed after six hours. Male and female pronuclei reconstructed and metamorphosing, in m the ‘fan ’ form with commencing division.

11,0. Placed in chloral 0-5% ﬁve minutes after fertilization ; preserved after ninety minutes.

n. Female pronucleus (four-rayed rosette), and male pronucleus .

(three-rayed rosette). 0. Fusion of pronuclei.

p. The same. Female pronucleus in the pseudo-tetraster forms. 128 EXTERNAL FACTORS III. 8

poison that the normal vitelline membrane cannot be formed and consequently many spermatozoa enter. In such eggs segmentation does not occur in the ordinary fashion by successive binary divisions, but many small cells are simultaneously formed. The resulting blastulae are abnormal, the segmentation cavity being ﬁlled with a solid granular mass (Stereoblastulac), and very few reach the Pluteus stage. The irregularities of segmentation are due to the complex mitotic figures and divisions which polyspermy entails. One, two, three or more of the spermatozoa fuse with the female pronucleus ; each has its own aster, which divides into two. Hence the most complex nuclear ﬁgures are formed.

In the case where two sperm-nuclei unite with the eggnucleus a tetraster is formed, that is four asters united by spindles in a. square or rhombus, or a triaster with an odd aster united to one angle of the system. The chromosomes are grouped in the equators of the four, or three, united spindles, as the case may be, and the egg divides simultaneously into four, or three.

The arrangement becomes still more involved when there are other sperms, whether these fuse with the female pronuclcus or not. Each amphiaster is united by one pole to the tri-, tetra-, or polyaster developed round the combination nucleus, or to the poles of other amphiasters; in one case there were nineteen spindles in all, not, of course, all in one plane. Each centrosphere receives half the chromosomes of the spindle attached to it, and each cell, when division occurs, contains one or more nuclei.

Hydrochlorate of morphine will produce similar effects, but only with longer exposures—a 0-4 % solution for from two to five hours. Strychnine, however, is poisonous in very weak doses (-005 % to -25 ‘Z), and quite short exposures are sufficient to call forth marked results. Other solutions successfully tried were chloral hydrate (from 0-2 % to 0-5 % for from one to four and a half hours), cocaine (from 0-025 Z to 1 ‘Z for ﬁve minutes), and sulphate of quinine (-05 % for ten minutes). In quinine (-05 % for thirty minutes) and chloral (o5 %) the entrance cone was small and no asters were formed, from which the III. 8 CHEMICAL COMPOSITION 129

Hertwigs argue that the contractility of the cytoplasm is impaired in these solutions. Chloroform dissolved in sea-water has the very interesting property of stimulating—without the addition of spermatozoa—the formation and separation of the vitelline membrane. The male generative cells are also sensitive to the action of these alkaloids, but not necessarily in the same measure. They can resist, for example, the inﬂuence of a solution of nicotine, which is ten times as strong as one necessary to evoke pathological changes in the ova. Though chloral hydrate (0-5 %) and quinine (0-05 %) are both temporarily fatal to the motility of the spermatozoa, sea-water restores the capacity for fertilization. Strychnine (0-O1 %) and morphine (0-5 %) are without elfect.

In the experiments just described the abnormalities seem to be directly due to the initial paralysis of the egg by the reagent and consequent polyspermy.

Should, however, the egg have been ﬁrst normally fertilized, the irregularities produced by the subsequent action of the poison are, though well marked, not of the same kind, for in this case the vitelline membrane has already been formed and only one spermatozoon has gained admittance. Chloral hydrate (Fig. 68, /:—p) was employed for ten minutes and at varying intervals after insemination (one, one and a half, ﬁve and ﬁfteen minutes). Exposure to the solution very shortly after insemination ﬁrst retards the progress of the sperm-head and the formation of its aster, and when later on the chromosomes are formed they lie heaped together in the centre of an achromatic ﬁgure described as a pseudo-tri- or pseudo-tetraster. This consists of three or four conical groups of ﬁbres, the bases resting on, and the ﬁbres connected to, the chromosomes, the apices outwardly directed and sometimes with, sometimes without, asters; in any case, however, they are not united by spindles, as is the ease in the complex ﬁgures observed in polyspermy. Isolated asters are also to be seen in the cytoplasm, and, which is perhaps more remarkable, the female chromosomes are themselves the centre of a unipolar (fan-shaped) or multipolar apparatus of the same kind. The reader will not fail to notice the similarity to the phenomena occurring in artiﬁcial parthenogenesis.

Jmrxxuou K 130 EXTERNAL FACTORS III. 8

Should the pronuelei unite—-which is only possible before these pseudasters have been developed, if the eggs have been subjected to the action of the poison immediately (one minute) after fertilization——the conjugation nucleus itself becomes the focus of a similar system. In eggs poisoned after a longer interval (ﬁfteen minutes) the male and female pseudasters may them selves unite. The nucleus—or nuclei—divide irregularly, the chromosomes

passing in unequal numbers to the poles of the ﬁgure. The several pseudasters and isolated asters, with which nuclei may possibly become secondarily associated, may be united by clear streaks of protoplasm, thus giving rise to a dendritie ﬁgure. Simultaneous and unequal division of the whole ovum follows.

Should the spermaster have already been developed-ﬁfteen minutes after insemination—it degcnerates. The subsequent changes comprise the formation of multipolar ﬁgures and irregular cell-division.

In later stages—when fertilization has been completed and segmentation is about to beg-in—the ova are almost or quite indiﬂerent to nicotine, strychnine, and morphine ; but chloral (0-5 %) destroys the asters which are already in existence and brings about a reconstitution of the combination nucleus with subsequent formation of a tetraster and quadruple division. In future mitoses, however, the spindles are bipolar. Cocaine and quinine (-05 %) (Fig. 68,./'—/1) have the same effect.

The importance of these experiments does not require to be emphasized. Not only do they throw a valuable light on the possible causes of those pathological mitoses that occur in malignant growths, they also contribute very greatly to the understanding of the normal processes of fertilization and karyokinesis.

Thus from the failure of the asters to appear in eggs treated with chloral before fertilization the brothers Hertwig argue that the contraetility of the cytoplasm is diminished by this substance, and from the failure of the pronuelei to unite in eggs which have been immersed in the solution shortly after fertilization they suggest that it is the contractility of the ovum which normally brings about the union of the pronuelei. Since, III. 8 CHEMICAL COMPOSITION 131

however, both male and female nuclei are able to divide, this division must be normally incited, not by their union with one

«another, but by the separate action of the cytoplasm on each, a view which is fully borne out by the phenomena. of artiﬁcial parthenogenesis and merogony (the development of fertilized enueleate egg fragments), whatever interpretation may eventually be put on the ‘ contractility ’ of the cytoplasm.‘

Another alkaloid which exerts an injurious influence on the ova of Echinoderms is atropine, the sulphate of which retards and dwarfs the development of /lxlericts and /lrbacia (Mathews). Pilocarpine, on the contrary, has an accelerating eﬁect, a result attributed by Mathews to its activity as an oxidizer, while atropine is regarded as a reducing agent, the property to which Loeb has also assigned the value of potassium cyanide in prolonging the life of unfertilized ova. The eggs of sea-urchins, when once laid, are only capable of fertilization and development within a certain deﬁnite limit of time, after the expiration of which they degenerate and die ,- after twenty-four hours, for example, they are only able, when fertilized, to reach the gastrula stage, and after thirty-two hours even fertilization is hardly possible. By treatment with an appropriate solution of potassium cyanide this limit may be considerably postponed. In the most successful series of experiments the ova were first

placed in a solution of KCN 7;?) in sea-water, and then

1'emoved successively every twenty-four hours to

7!: 7! 2500’ 3000' lengths of time, then removed to pure sea-water and fertilized. As the table shows (Table XVII), segmentation was still possible after 168 hours’ sojourn in the solution, but the greatest number of Plutei was obtained after only 66 hours’ stay.

It was also shown that better results could be obtained with artiﬁcial parthenogenesis if the ova were first kept in the cyanide solution. Loeb points out that in the higher animals

74 7! 1400’ 2000’ In the last solution they were kept for various

‘ Strictly speaking, only the division of the male chromosomes can be regarded as being stimulated by the egg cytoplasm. What exactly it is which excites the female nucleus to divide is not at all clear.

K2 132 EXTERNAL FACTORS III. 8

the eﬁects of this substance are due to its inhibition of oxidation; that this is the real cause of the prolongation of the life of the eggs is shown by the fact that when kept in‘ an atmosphere of hydrogen for thirty-eight hours they were still capable of being fertilized and developing intoswimming larvae.

TABLE XVII

Showing the effect of exposures of various length of Sea-urchin

eggs to a solution of KCN§(%—0- (After Loeb.) Length of exposure in hours. Result.

66 80 7; Plutei, vitelline membrane formed 90 30 ‘Z, Plutei, no vitelline membrane formed 99%‘ 20 Z Plutei) n 91 n n

112 Less than 20 Z Plutei

I20 Gastrulae, but no Plutei

139 A few blastulae

140 Blastulae, not swimming

Eight-celled stage only

Simultaneous lowering of the temperature to the freezing-point enhanced the value of the cyanide treatment.

In later stages, however, immediately after fertilization and subsequently, the action of potassium cyanide is by no means beneﬁcial ,- at this time, as we know, oxygen is a necessity (see above, p. 112); and Lyon has shown that the moment at which the ova are particularly sensitive to both KCN and the lack of oxygen is the same, about ﬁfteen minutes after insemination.

Chemical agents are also able to incite irregularities of growth and abnormalities in later stages of development.

In a long series of experiments Féré has shown that monstrosities can be produced by exposing the Hen’s egg to the unfavourable influence of a large variety of substances. Vapours of ether, alcohol, essential oils, nicotine, mercury, and phosphorus, injections of alkaloids such as morphine, nicotine, strychnine, and others, of bacterial toxines (those of tubercle, diphtheria), of peptones, dextrose, glycerine, several alcohols, certain salts (KBr, III. 8 CHEMICAL COMPOSITION 133

KI, S1-Brz), are all baneful, retarding and distorting the embryo to a. greater or less extent. Ammonia, it may be noted, is fatal at once.

It has already been shown (p. 123) that the malformations induced by sodium chloride in Amphibian embryos are to be set down to some other property than the osmotic pressure of the solution, and it is here only necessary to advert to

Fm. (S9.-—Gane-sugar‘ (6-6 2,). Two stages in the forma.tion of the notochord from the whole thickness of the roof of the archcnteron in the Frog.

Dextrose (3-4 7,). Secondary degeneration of the gut roof and ventral part of notochord.

some of the more interesting effects occurring in particular solutions.

Although the more poisonous salts (e.g. LiI, CaCl2, SrBr2, and others) inhibit altogether the formation of the blastoporic fold, a cause which normally assists in its production—the proliferation of small cells in the roof of the segmentation cavity--may continue to operate, with the result that that mof is thickened and thrown into puckers and folds. 134 EXTERNAL FACTORS III. 8

Again, the notochord may be formed from the whole thickness of the arehenteric roof (cane-sugar)recalling the mode of its development in Urodela and Petromyzon (Fig. 69); the solid medullary tube observed in potassium chloride and other salts reminds one of the rudiment of the nervous system in Teleostei and others, while the mode of closure of the medullary tube in, for example, some of the magnesium salts resembles that observed in Am];/ziomas ; the formation of notochordal tissue from the wall of the neural

FIG. 70.-—-Fornmtion of vaeuolated notochordal tissue in the medullary tube of the Frog embryo under the inﬂuence of urea (1-6%). Underneath the notochord is the subnotochordal rod.

tube and the roof of the arehenteron (Fig. 70) in strong solutions of urea (1-17' % to 1-56 %) shows that the prospective potentialities of these organs are not yet ﬁxed, while the development of an optic cup without a lens in urea, sodium chloride, and sodium bromide demonstrates that the formation of the former is independent of that of the latter of these two parts of the eye.

The grey degeneration of the exposed part of the medullary plate (due to the distribution of the pigment throughout the cell—body), the protrusion of cells (‘framboisia’ of Roux), and disintegration of the epithelium which is so characteristic in III. 8 CHEMICAL COMPOSITION 135

many of these solutions (cane-sugar, NaCl, LiCl, MgCl2, MgSO,), have been noticed by many observers (Roux, Hertwig, Morgan, Bataillon). All the more violent solutions attack the yolk-granules. In some cases the effect produced appears to be speciﬁc; thus in lithium salts the ectoderm is often pitted and wrinkled before any degeneration appears in the nervous system, and in ammonia salts, which are highly poisonous, the nuclei are much enlarged, lobed, highly chromatic, and homogeneous. The very similar appearance of the nuclei (Fig. 64) in those stronger solutions of urea which arrest development in an early stage suggests that ‘the ammonia set free is the toxic agent in this case. In solution isotonic with -625 % NaCl urea permits of normal development up to a certain point, when the embryos die.

In this connexion it is interesting to notice that Moore has found that sodium sulphate will act as an antidote to the poisonous effect of sodium chloride on tadpoles. Thus the

average length of life of tadpoles in a 3 NaCl solution was

four and a quarter days, but was prolonged to twenty-one days by adding from 4% to 8 X of Na2SO4. The poisonousness of sodium chloride, sodium nitrate, calcium nitrate, and magnesium chloride to Fmululus embryos and the value of other salts as antidotes has been shown by Loeb, while Lillie has noted that sodium is fatal but magnesium and calcium beneficial to the ciliary movement of /lrem.'c0/a larvae, a result ﬁrst obtained by Loeb for the Plutei of Ea/u'7m3; the muscular contractions of the larva, on the other hand, are inhibited wholly by magnesium, partly by calcium, while sodium is necessary for their continuanee. In an artiﬁcial solution which combines the three elements in the proper proportions normal development is possible. The nature of the part played by the ions——-whether toxic or antitoxic— is, however, a very open question.

Arguing from the fact that the evil eifects of such salts as sodium chloride and nitrate may be counteracted by calcium and magnesium salts, Loeb has suggested that toxicity and antitoxicity are functions of valency, and also of electrical charge, since it is further stated that toxicity increases with the valency 136 EXTERNAL FACTORS III. 8

of the anion, antitoxicity with that of the cation. Ions of the same valency are not, however, necessarily equally antitoxic (Loeb and Gies, Lillie, Mathews), and sodium sulphate, as we have seen, may act as an antidote to the chloride (Moore). Mathews has accordingly sought for the cause of toxicity in another physical property, the decomposition tension of the salt, and has certainly succeeded in showing that the poisononsness of solutions to the eggs of Fzmrlulzw varies inversely with the decomposition tension, and that a similar relation holds good in certain other cases.

Lillie argues that a physiologically balanced solution is necessary, one in which the electrolytes are in a state of chemical equilibrium with the necessary ion-proteid compounds in the tissues. Solutions which only contain some of these substances, or solutions (for example, non-electrolytes) which contain none, are poisonous, because they permit of the outward diffusion of the needful ions.

It must be pointed out, however, that this explanation will not ﬁt the cases where the embryo develops perfectly well in fresh water (lllzmtlulus) or in distilled water (the Frog), and that some other reason must be found for the poisonous effect of cane-sugar upon the latter. The whole question, however, is one which belongs more properly to the province of pharmacology.

Poisonous although these salts are, the embryo can still be acclimatized to them. C. B. Wilson placed the unsegmented eggs of Amlalyertoma, Rana, and 0/102-op/zilzcs in a 0-05 % solution of sodium chloride; after twenty-four hours they were removed to 0-1 %, and then successively to stronger solutions by increments of 0-1% until 10% was reached, a concentration which quickly causes death under ordinary circumstances. In this case, however, development was normal, and the larvae hatched out and lived for some time.

The distortions of development which solutions of salts and other substances call forth in Amphibian embryos find a parallel in the malformations which Herbst has produced in Eehinoderm larvae (I976/dmw, Sp/zaerec/ainus) by similar means; as in the former case, the results were at first assigned to the increased osmotic pressure of the media. III. 8 CHEMICAL COMPOSITION 137

When potassium salts are added to the sea-wa.ter—for example, a 7 % solution in sea-water of a 3-7 % Solution of K01 in tap-water——the egg gives rise to a Pluteus in which, though the gut is, as normally, tripartite, the skeleton is rudimentary and the arms suppressed (Fig. 71). Herbst suggests that the suppression of the arms is" due to the absence of a. stimulus normally exerted by the skeletal spicules. These abnormal forms may fuse together to form double monsters.

Such ‘ potassium ’ larvae are developed in sodium salts, but lithium has a more pronounced elfect (Figs. 72, 73). In this case

FIG. 71.-—Potassium larvae of Echinoids. a. Potassium larva of Sphaerechinus (1860 c.c. sea-wate1'+ 140 c.c. 3-7% KNO3). There is no skeleton. The rut is tripartite, and the mouth surrounded by the ciliated ring. I), c. otassium larvae of .E(‘h’i1‘l'llS (20% of 3% K01). Note the buttonshaped apical tuft of cilia, and, in c, the secondarily evaginated archenteron. (After Herbst, 1893.)

the blastula becomes constricted into two portions, a thin-walled gastrula wall provided with long cilia, and a thick-walled archenteron, which may be muscular and mobile, and is thickly covered with short cilia. The arehenteron has, in fact, failed to invaginate, and the larva is an ‘ Exogastrula ’. Occasionally there is an attempt at invagination at the end of the archenteric portion, and, after temporary exposure, the invaginated part may be divided into three, and a mouth formed. All the parts of the gut, however, remain in the same straight line. A middle section may be formed by further constriction of the archenteron (Ea/aimzs) or of the gastrula wall (Sp/zaerec/liuus). Double 138 EXTERNAL FACTORS III. 8

monsters sometimes arise by fusion of these larvae by their archentera.

A skeleton is not usually developed; if present it is abnormalin position, the spicules being placed near the animal pole and

FIG. 72.———Lithium larvae of Splmerechin us _qrmmlw'is. a. Larva. partially constricted into gastrula wall and archentcric portions, the former with lon , the latter with short cilia. (980 c.c. sea-wa.ter+2O c.c. 3-7% LiCl). b. imilar larva. to the last, but a neck or connecting piece has been formed from the ectodermal portion. 0, (1. Progressive diminution of the ectodermal gastrula. wall portion with increase in the quantity of Li.

the arms of the Pluteus formed under their inﬂuence near the mouth instead of by the side of the anus, in the number of the spicules, and consequently the number of arms (three, four, or

ﬁve, instead of two), and in the number of their radii (four or ﬁve, instead of three). III. 8 CHEMICAL COMPOSITION 139

The gastrula wall is often smaller than the archenteron, and, as the strength of the solution is increased, becomes still further reduced, until nothing of it is left but a small button at the

F10. 73.—~a. Larva with three skeletal spicules, and a. ‘ cell-rosette ’ at the end of the archenteron. Ir. Larva with skeleton—-—more than three spicu1es—and arms developed. The neck is invaginated into the ectodermal portion, the gut tripartite. c. Five-armed Plutcus with ﬁve skeletal rods. The gut is normally invaginatecl and tripartite. d. Larva. of Echirms microtuberculatus. There is a neck, and the gut is partly invaginated. In the blastocoel are aggregations of mesenchyme and pigment cells. (After Herbst, 1895.)

animal pole, which only indicates its real character by the longcilia which it carries. Such larvae Herbst terms ‘Holoentoblastia’. This nearly complete suppression of the ectodermal 140 EXTERNAL FACTORS III. 8

region can, however, only be realized when the salt is allowed to act at a stage in the blastula when the diﬁerentiation into the two primary layers is already beginning. Should the embryos be removed before this stage is reached, after twentyfour hours’ exposure to the solution, only ‘Exogastrulae’, not ‘ Holoentoblastia,’ can be obtained. Should, on the other hand, older blastulae, or gastrulae, or Plutei be placed in the solutions, they die without showing any signs of the characteristic abnormal development. From the fact that equimolecular solutions of monobasie lithium salts produced like effects (such solutions, it must be observed, are also chemically equivalent), Herbst concluded at first that the osmotic pressure was responsible for the abnormalities; but the permanent after-effects of temporary immersion just referred to subsequently convinced him that the ova were permeable to the lithium ions to which he now attributes the speciﬁc nature of the monstrosity. He suggests further that they act upon the endoderm cells by increasing their absorptive activity and their power of cell-division, while at the same time they inhibit the functions of those mesenchyme cells which are devoted to the formation of the skeleton.

As in other monstrosities, there is an alteration in the prospective potentialities of cells, elements which would normally be ectodermal becoming converted into endoderm, and additional mesenchyme cells being involved in the secretion of skeletal spicules.

It is only by lithium salts that the typical ‘ Holoentoblastia’ can be, produced ; but Exogastrulac can be reared in others, in sodium butyrate, for example; in this solution a stomodaeum is formed, but is, like the arehenteron, everted. Even lithium, however, is powerless to cause the ‘ holoentoblastic ’ reduction of the gastrula wall in the larvae of Aaterias, although exogastrulation "may, but need not, occur. A characteristic deformity is the absence of the pre-oral region, and the elevation of the mouth on a sort of hypostome. In Amp/ti0.'D7(8 and Ascidians it is impossible to obtain even exogastrulae by these methods. It is evident, therefore, that the speciﬁc morphological reaction depends not only on the nature of the substance employed, but also on the constitution of the reacting organism. III. 8 CHEMICAL COMPOSITION 14-1

Herbst has not omitted to point out the signiﬁcance of theseand indeed of a1l—monstrosities for the theory of the origin of those larger, discontinuous variations known as ‘sports’, or, in more modern phraseology, ‘mutations’; and Vernon has been able to show statistically that the degree of continuous variation may also be altered by changes in the chemical environment.

In all the foregoing experiments the effect is observed of the addition of some chemical substance to the medium in which the embryo is placed. We have now to consider a very remarkable series of investigations, for whose planning and execution we are indebted to the genius of Curt Herbst, investigations in which substances which are present in the normal environment of the larva are omitted, and an insight thus gained into the part they play, if any, in the normal development of the organism. Herbst has indeed succeeded in demonstrating in the most conclusive manner the necessity to the sea-urchin egg for the normal performance of this or that phase of developmental function of a large number of the elements present in sea-water.

The sea-water at Naples, where Herbst carried out his work, has the following c0mp0siti0n:~—— N aCl . . . . . 3 % KCI . . . . . - 7 % MgCl2 . . . . . ~32 % Mg-S04 . . . . . -26 Z CaSO4 . . . . . "-1 % CaHl.’O4

Ca3P208

CaCO,,

Fe2CO Si

Br

I

It may be said at once that silicon, bromine, and iodine are unnecessary, and that, though earlier experiments led Herbst to believe that phosphorus and iron were essential, he has since assured himself that phosphorus is certainly, and iron probably,

in small quantities.

ll 142 EXTERNAL FACTORS III. 8

not. All the other elements, however, can only be omitted under penalty of retardation, abnormality, or death (Figs. 74 A and B).

The method employed was a simple one. A series of artiﬁcial sea-waters was made up, from which, one by one, each of‘ the elements was omitted, another being substituted in its place. Care was taken to make these artiﬁcial media approximately

FIG. 74.——'1‘he necessity of substances contained in sea-water for the normal development of the larvae of sea-urchins.

a. Without OH. Ciliated stereoblastula of Sphaerechinus. b. KOH has been added. c. Normal blastula of Sphaeiw-hinus. d. Blastula in

a K-free medium. e. Reared in K-free and replaced in sea-water (Sphm=r- '

echinus). f. Larva from a medium devoid of Mg (Sphaerec-himls). g. Echinus Pluteus with tripartite gut, mouth and coelom sacs, but neither skeleton nor arms ; reared without CaC0, or CaSO,. h. Normal Pluteus of Echinus.

isotonic with sea-water, and so exclude a possibly disturbing factor, the alteration of the osmotic pressure. The réle of each of these necessary e1ements—or ions—will be considered separately and in some detail. Sp/2ae7'ec/Iinua and E0/timcx were the forms principally employed. III. 8 CHEMICAL COMPOSITION 143

i. S0,‘.

This is ordinarily provided by Mg-S04 and CaSO4 ; when the fertilized ova are placed in a solution in which MgCl2 is substituted for it (as, for example, in 3 % NaCl+-07% KCI + 5 % MgCl2 + CaHPO4 + CaCO3) then their development is retarded from the blastula stage onwards, the embryos are small

and degenerate without reaching the Pluteus stage (Fig. 74- B). The gut is straight instead of bent, and not divided into the

FIG. 74 B.

a. Normal position of skeletal spicules in Sphaerechimls. b, (1. Abnormal position and number after treatment with S0,-free medium. c. Larva of Echiuu.s- from a S-free solution. e. Pluteus of Sphaerechinus with three fenestrated skeletal arms, instead of two. ’l‘rea.ted with a S0,-free medium and replaced in sea-water. f. Normal Pluteus of Sphaerechinus. (After Herbst, 1897 and 1904.)

usual three parts ; in S11/zaerec/u'm¢s no mouth is formed, the gut is evaginated (Exogastrula). The endoderm is very thick, the cells dark and dense.

The sulphuric acid radiele (sulph-ion) is thus necessary for the proper development of the gut, and necessary from the very beginning, for in embryos which have been kept in S0,,-free water up to the mesenehyme-blastula stage and then replaced in sea-water the alimentary tract is still abnormal.

Deprived of S04, in fact, the gut remains radially symmetrical, 144 EXTERNAL FACTORS III. 8

and the same must be said of the skeleton. Normally there are two tri-radiate spicules, one to the right, the other to the left of the gut and some little way from it. Without the needful sulphate the spicules become placed near the gut, and may with the growth of the latter be pushed towards the animal pole. The number of spicules may also be diminished or increased to one, three, or four, arranged in a circle round the gut. On timely removal to sea-water, however, a secondary bilateral symmetry may arise by two of these outgrowing the rest and stimulating the development of the typical arms of the Pluteus. It seems that a sulphate is present in the calcareous skeleton of the Pluteus, as there is in that of the adult urchin.

A ciliated circum- oral ring is formed, but is abnormal in its position, at right angles instead of parallel to the long axis of the body. The pigment which should be secreted by the secondary mesenchyme cells (separated off from the inner end of the archenteron) remains in abeyance, and the apical tuft of cilia is hypertrophied. Other processes, however—fertilization, segmentation, and ciliary motion——arc independent of S0,.

The development of eggs which are allowed to remain in ordinary sea-water until the blastula stage is no better, whence Herbst concludes that no S04 is taken up during segmentation. During the early stages of gastrulation, however, they appear to absorb a store of it for future needs, for gastrulae reared in sea-water develop further in the SO,-free solution than do those embryos which have been kept in it since fertilization. S04 is equally necessary for the continued life of the Pluteus and of the Bipinnaria larva of Asterias, and without it the rate of regeneration of the head of Tubularia is retarded and the number of tentacles reduced, until eventually a completely tentaeleless head is evolved.

The necessary sulphate can be, to a certain extent, replaced by a thio-sulphate. The addition, for instance, of -35% Na2S2O,, to the SO,-free solution renders it possible for the larvae to reach the Pluteus stage, though the arms are short, the skeleton small, and the pigment reduced. The larvae die.

Selenium and tellurium are both poisonous in an early stage. III. 8 CHEMICAL COMPOSITION 145

ii. Cl.

A solution was made up in which the sodium chloride was replaced by sodium formate, the magnesium chloride by magnesium sulphate, the potassium chloride by potassium sulphate ; thus, NaCOOH 3- 5 % + MgSO4 -26 % + MgSO4 -4- % + KZSO4 -12 Z + CaSO4 -I Z + CaHPO4+ CaCO3.

The eggs did not segment, and even when KCl and MgCl2 were used in their ordinary proportions, segmentation did not progress very far. Nor did the substitution of Na2SO4 for NaCOOH give any better results. A considerable amount of chlorine appears therefore to be absolutely necessary for the earliest developmental processes, its function being, Herbst suggests, to transport certain necessary cations, the tissues being possibly more permeable to NaCl than to Na2SO4. Later stages ——blastulae, gastrulae, Plutci—all die in the Cl—free mixture.

Chlorine can be replaced in some measure by bromine. Plutei are formed, though with a distorted skeleton, Tubularia regenerates its head and the eggs of the ﬁsh Labrax develop as well as in sea-water. Iodine is, however, poisonous; so also are chloratcs.

iii. Na.

2-96 % of MgCl2 was added to a solution containing the usual amounts of KCl, MgSO4, CaSO4, CaHPO4, and CaCO3. In this the ova indeed segmented, but abnormally, the blastomeres being of unequal size. Death followed; nor was the addition of a certain small amount (-84: %) of NaCl suﬂicient to save them, though segmentation was normal and traces of an archenteron could be detected; with more NaCl (1-34» %) the gastrula stage was reached. The sodium which is thus necessary in the earliest period is also required later on; without it gastrulation is impossible to eggs which have been reared in sea-water even as far as the mesenchyme—blastula stage.

The part played by sodium is not clearly understood. It is known that it counteracts the evil eifeets of calcium and is necessary for the continuance of muscular contractions. Since calcium is necessary for the cohesion of cells (see below) Herbst opines that sodium may pull them apart ;. its action in that case is capillary.

Jnnxmson L 146 EXTERNAL FACTORS III. 8

Sodium cannot possibly be replaced by lithium, potassium, rubidium or caesium, all of which would at the necessary concentrations inevitably be poisonous.

iv. K.

In the artificial solution employed the small quantity (-07 %) of potassium present in sea-water is simply omitted.

Without it scgmentation—except the ﬁrst few phases-is impossible in Ea/cinzw. Sp/zaerec/limts, however, segments, but the blastocoel is reduced, the cells are opaque and not vacuolated, and the ova, though ciliated, are motionless and die (Fig. 74 A, rl, e).

Later stages are also sensitive to the want of potassium. Blastulae gastrulate, but are shrunken, with short archenteron, and in gastrulae the gut does not divide into three. Plutei, like all the others, die when deprived of it.

In the K-free medium spermatozoa temporarily lose their motility, and such spermatozoa cannot effect fertilization. The fertilization, however, of eggs which have been kept without potassium is possible ; in fact, at this earliest stage, no potassium is absorbed, for eggs fertilized in sea—water develop no further in the K-free solution than do those fertilized and kept continuously in it.

The absence of potassium i11 segmentation leaves its effect upon later stages. Two days’ exposure is not too long to prevent normal development on removal to sea-water, but ﬁve days’ exposure causes abnormalities of the skeleton (asymmetrical with several triradiate spicules round the gut) and alimentary canal (no mouth).

De Vries has shown the importance of potassium for the turgor of young plant-cells, and its function here is probably similar, to promote growth, as the subjoined table of measurements shows. Its absence also aifects the rate of development (Table XVIII).

Potassium can in a measure be replaced by rubidium and caesium. The use of lithium either has no effect or, in larger quantities, produces lithium larvae.

Other forms to which the lack of potassium was found to be fatal were the ova of Asterias and Uotﬂorﬁiza, and the adult III. 8 CHEMICAL COMPOSITION 147

Amp/iioxus. Potassium is also necessary for the contractions of muscles (umbrella. and tentacles of Obrlia).

TABLE XVIII Showing the effect of potassium upon the growth of Sea.-urchin blastulae. (After Herbst.)

‘Ratio (unit 2 Th mm.) of crossdiameter to long diameter.

After Without K. With K. 18 hours . 24 hours 45 hours . . .

Showing the effect of potassium on the rate of development of

Sea—urchin larvae. (After Herbst.) Artiﬁcial sea-water with

After -008 Z KCI. -016 Z KCl. .024 % KC]. 36 hours Small gastrulae. Nearly Plutei. Plutei. 60 hours No mouth ; gut not Small Plutei. Fully formed Plutei. constricted. v. Mg.

' The fertilized ova were placed in a solution which did not

include the -32 % MgCl2 and the -26 Z MgSO4 present in seawater.

Segmentation proceeds normally, but the blastula. is slightly smaller than when magnesium is present (the ratio of the diameters is §§). The skeleton, however, though bilaterally laid down, is retarded and deformed (magnesium is present in the skeleton of the sea-urchin and possibly in that of the Pluteus) and the gut is not properly diiferentiated, not tripartite and without a mouth (Fig. 74 A, f).

When the MgSO4 is replaced by an isotonic quantity of Na2SO4 the results are the same.

In the Mg-free solution cilia cease beating, development is retarded (Table XIX), and, though the spermatozoa retain their motility, the ova are so injured that fertilization is impossible unless they are restored to sea-water. The ova, in fact, seem to have a store of Mg which they lose in the Mg-free mixture. Fertilization can, however, take place without magnesium if the

L 2 148 EXTERNAL FACTORS 111. 3

eggs have been kept in sea-water, and such eggs develop, in Mg-free water, to precisely the same extent as those which have been fertilized in sea-water; at this period, therefore, the egg needs no external magnesium. The original store of this element apparently suffices also for the ﬁrst steps in the formation of the gut; for the lack of it is felt equally in the later stages of its diﬁerentiation and in the ﬁrst moments of its development; the alimentary canal is as abnormal in those larvae which have been kept without magnesium only up to the time when the mesonchyme and archenteric plate arise as in those which remain in the solution throughout.

TABLE XIX Showing the retardation of development of Sea-urchin larvae deprived of magnesium. (After Herbst.) Two solutions were employed, mixed in various proportions; one (1)

has no Mg; the other (2) contains Mg. Solution. Result after 24 hours.

1 Nearly motionless; archentcron formed. 1+ 10 Z of 2 Larger. 1 +20 Z of 2 Larger still. 1 + 30 Z of 2 Larger; gut constricted ; mouth i'or1ned.

1 % of 2 )1 n n n 7) 1 % of 2 H n H :9 n 2 H H n 7) n

It is, in fact, only after the gastrula stage that magnesium is absorbed. Eggs, blastulae and young gastrulae, reared in seawater develop far worse when placed in the Mg—free liquid than do gastrulae.

The formation of pigment and the contractility of muscles remain unaifected by the absence of Mg.

In Mg-free media tl1e blastomeres of .1.s-/erizw fall apart and the adult medusae of Obeliu degenerate.

vi. Ca.

The calcium salts present in normal sea-water are C-aCO_.,, Ca2SO4, CaHPO4, and Ca3(PO,)2.

When the carbonate only is absent the blastulae are crumpled and opaque; a few gastrul-ate but are markedly abnormal, the ciliated ring being crumpled, the gut ﬂattened on the oral side and the skeleton absent (Fig. 74 A, g). Should the skeleton have already been formed when the larvae are exposed it 4 III. 8 CHEMICAL COMPOSITION 149

becomes dissolved. When the sulphate only is omitted (magnesium sulphate being present) development is still inferior to the normal, inferior even to development in the presence of CaSO, but in the absence of MgSO4 ; CaSO4 may be replaced by CaCl2. It seems, therefore, that the sulphate is necessary as a calcium salt.

FIG. 75.——a—r. Separation of the blastomeres of Echimts microtubermr Iulus in a medium containing NaCl, 3-07%, KCI, 0-08%, MgSo,, 0-66%, MgHPO, and FeCO3, but no Ca. Note the radially striate border, which is the altered uniting membrane. (1. lilastuln disintegrating in the same medium. (After Herbst, 1900.)

If only the phosphate (Ca3(PO4)2) is present the egg dies during segmentation, though, if the other phosphorus compound is substituted for it, the effect is the same as when the carbonate alone is omitted.

Should, however, all calcium salts be removed the result is

more serious still (Fig. 75). The blastomeres are unable to 150 EXTERNAL FACTORS III. 8

cohere, and separate as fast as division takes place, swimming about independently for a time, and then dying. The same phenomenon is witnessed when later stages are placed in such a. calcium-free mixture.

On removal to sea-water division continues without separation, and should the egg membrane still be intact all the cells unite and a whole larva is formed. Even should the egg membrane be lost, a reunion of the cells is always possible so long as they remain in contact with one another. The separation is due to a. change in the surface-tension of the cells; a visible change takes place, in fact, in the superﬁcial layer which covers and unites the blastomeres ; it becomes ill-deﬁned and radially striated.

The lack of calcium also aifects the rate of development, and causes shrinkage, but leaves karyokinesis, ciliary motion, and pigment formation unaltered.

Calcium is not replaceable by magnesium, strontium, or barium.

vii. CO3.

As has just been pointed out, calcium carbonate is necessary for the due formation of the skeleton, although a beginning may be made without it.‘

Whether the crumpling of the larva, due to diminution of internal osmotic pressure, which is observed in the absence of calcium carbonate is attributable to the lack of CO3 or the lack of the hydroxyl ion is diﬁicult to determine, since, as Herbst points out, a carbonate necessarily introduces OH, while the latter can convert into carbonates the CO2 of the atmosphere and of respiration.

viii. OII.

The alkalinity of the sea-water——reckoned by the number of free hydroxyl ions—is provided by the calcium carbonate and calcium hydrogenphosphate. By the omission of these a solution —-neutral to litmus—may be obtained in which the ova give rise to thick-walled, opaque blastulae with granular contents, ciliated

1 CaCO,, is necessary for the formation of the skeleton of the larva of the sponge Sgcandra selosa (0. Maas, S.-B. Ges. Mozph. Phys. Miinchen, xx, 1905). In a medium devoid of all calcium salts the Amphiblastulae fall to pieces. III. 8 CHEMICAL COMPOSITION 151

but motionless, and doomed to eventual degeneration and death (Fig. 74 A, a, 6). Very occasionally a gastrula with a short gut is formed.

When the blastulae are immersed in the solution they give rise to small, opaque gastrulae.

A certain degree of alkalinity is necessary for fertilization. The spermatozoa are less sensitive to a want of alkalinity, more sensitive to excessive alkalinity than the ova.

By the addition of a small amount of sodium hydrate to the neutral medium development is accelerated, but an increase of the alkalinity of ordinary sea-water is unfavourable. Loeb, on the other hand, has found that the addition of from -006 % to -008 % sodium hydrate to sea-water accelerates the development of Aréacia.

The formation of pigment and the vibration of the cilia are other processes which depend on the presence of the hydroxyl ion. Plutei die without it and their skeleton is dissolved.

The function of the ion does not appear to be to neutralize any acids produced by the tissues, for these give a neutral reaction even in OH-free media.

Since aeration improves the development of eggs in these media, and the more so if the air is deprived of its carbon dioxide, Herbst has concluded that one function of the OH ion is to neutralize the CO2 and allow of the formation of the necessary carbonates. Another function is possibly, as Loeb suggested, the acceleration of processes of oxidation.

The experiments which we have been considering are unique of their kind, and it is impossible to exaggerate their importance. For, whatever may be the ultimate explanation of the facts, there can be no doubt whatever that the most complete demonstration has been given of the absolute necessity of many of the elements occurring in ordinary sea-water, its normal environment, for the proper growth and differentiation of the larva of the sea-urchin. Nor is this all. Some of the substances are necessary for one part or phase of development, some for another, some from the very beginning, others only later on. Thus potassium, magnesium, and a certain degree of alkalinity are essential for 152 EXTERNAL FACTORS III. 8

fertilization, chlorine and sodium for segmentation, calcium for the adequate cohesion of the blastomeres, potassium, calcium and the hydroxyl ion for securing; the internal osmotic pressure necessary for growth, while without the sulph-ion and magnesium the due differentiation of the alimentary tract and the proper formation of the skeleton cannot occur; the secretion of pigment depends on the presence of some sulphate and alkalinity, the skeleton requires calcium carbonate, cilia will only beat in an alkaline medium containing potassium and magnesium, and muscles will only contract when potassium and calcium are there.

The part played by each substance is therefore speciﬁc; for some particular part of the morphogenetie process it is indispensable. Not, of course, independently of internal factors, but in co-operation with them, it does, in fact, determine the production of organic form; and the relation between the embryo and the environment in which it develops is in this case, at any rate, of the closest and most intimate kind.

LITERATURE

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'1‘. H. MORGAN. Further studies on the action of salt-solutions and of other agents on the eggs of Arbacia, Arch. Ent. Mech. x, 1900.

T. H. MORGAN. The relation between normal and abnormal development of the embryo of the Frog as determined by the effect of lithium chloride in solution, Arch. Ent. Mech. xvi, 1903. III. 8 CHEMICAL COMPOSITION 155

H. NEILSON. Further experiments on the antitoxic effects of ions, Amer. Joum. Phys. vii, 1902.

S. R1NG1=.n and A. G. PH!-JAR. The inﬂuence of saline media. on the tadpole, Jam-n. Phys. xvii, 1894-5.

J. RITCHIE. The relation of chemical composition to germicidal action, Trans. Path. Soc. 1, 1899.

W. ROUX. Beitritge zur Entwieklungsmechanik des Embryo : I. Zur Orientierung ﬁber einige Probleme der embryonalen Entwicklung, Zeifschr. Biol. xxi, 1885.

D. RYWOSCH. Ueber die Bedeutung der Salze fiir das Leben des Organismus, Biol. Centralbl. xx, 1900.

T. SOLLMANN. Structural changes of ova in anisotonic ‘solutions and saponin, Amer. Journ. Phys. xii, 1904-5.

C. B. WILSON. Experiments on the early development of the Amphibian embryo under the inﬂuence of Ringer and salt solutions, Arch. Ent. Mech. v, 1897.

W. D. ZOETI-IOUT. The effects of potassium and calcium ions on striated muscle, Amer. Jom-n. Phys. vii, 1902.

9. SUMMARY

In the numerous experiments which we have been considering the eﬁect is observed upon the development of the embryo of certain alterations in the constitution of that embryo’s normal environment. Either some factor which is not usually present is added to the environment, or else some factor which is customarily found there is altered by increase or decrease, or removed altogether.

In some cases development remains undisturbed by this treatment, in others it may be merely generally retarded or accelerated, in others again it may be altered not merely in rate but in form, with the production of an abnormality or monstrosity, and if its eifeet is too intolerable the death of the embryo may ensue.

Throwing light as they do on the causes of the formation of natural monsters, such experiments are no doubt of the highest interest from a general teratological point of view. The mere possibility of the occurrence of such malformations is, however, itself a fact of the deepest morphological signiﬁcance. A monster is an organism in which the development of some part or parts has either exceeded or fallen short of its normal limitations, and any such phenomenon points indubitably to a certain mutual 156 EXTERNAL FACTORS III. 9

independence of the parts in the growth and differentiation of the organism; while some pursue their normal course, others deviate from it. It follows that when such deformations are due to changes in the external conditions the parts are not equally sensitive to the unusual inﬂuence to which they are exposed. Thus in the Frog embryos which exhibit persistent yolk-plugs and open brains when grown in solutions of various kin(ls, the yolk and the medullary folds are alone susceptible to the action of the poison, other parts are unaffected and continue their development as though under normal circumstances: or, again, a sea—urchin passes through the early stages of segmentation and gastrulation unchanged when placed in a sea-water from which magnesium has been removed, but the subsequent differentiation of the gut and the formation of the skeleton are abnormal ; magnesium is necessary for these, though not for the earlier processes. A means is thus afforded of watching the behaviour of one or more parts independently of others, as, for example, of the animal cells in the gastrulation of the Frog's egg when the yolk-cellspare injured, and the most valuable information contributed, often quite unexpectedly, to our understanding of the events of normal ontogeny.

Quite apart from this such experiments have already contributed, and will probably contribute still more in the future, to the study of variation. Between conspicuous monstrosities and those milder abnormalities which are termed ‘sports ’ or ‘ mutations ’ there is every intermediate gradation, just as there is, on the other hand, no sharply deﬁned limit between these discontinuous and those far smaller continuous variations to which the term has been often exclusively applied. The embryo is particularly sensitive to a change in its environment and reacts to such change by a variation in its form of greater or less degree. And not only that ; as Vernon has shown, these changes can produce also an alteration in the variability of the species ; and so provide greater opportunities for the operation of natural selection.

At the same time teratolugy is not the main inquiry with which the experimental emhryologist is concerned. The problem that confronts him is to determine the part played by each factor of the external environment in the processes of III. 9 SUMMARY 157

normal, speciﬁc growth and differentiation, and for the solution of this problem only those experiments, of course, are of avail in which such factors are either altered or removed.

By this means, as we have seen, it has been shown that a certain constitution of the physical environment, ﬁxed within certain limits, is needful for the embryo; to these conditions it is closely adapted; those limits it can only transgress under pain of abnormality or death.

Every factor, or nearly every factor, is necessary for this or that phase or part of the process, some for the whole. Light of a certain wave-length will accelerate development, light of another kind, or in some instances darkness, will retard it, or stop it altogether; a certain degree of heat is indispensable ; oxygen is required for respiration, water for growth ; some eggs demand constant agitation, others comparative rest; fertilization, or segmentation, or gastrulation, or some one or other of the later phases of development may depend absolutely on the presence of some particular chemical element ; remove the factor in question, whatever it may be, and that particular process will not occur, and the speciﬁc, typical end which is reached in normal development will not be attained. Nevertheless, the achievement of this end does not depend wholly upon extrinsic forces, for the ovum is no completely homogeneous ‘isotropic’ substance in which the complex circumstances of its environment conspire to produce heterogeneity and coherence. There is no evidence that any physical factor exerts a directive inﬂuence sufficient of itself to determine any part of the whole speciﬁc cﬁect, although this may happen under extraordinary conditions, as when gravity impresses a bilateral symmetry upon the compulsorily upturned egg of the Frog, and so determines the median plane of the embryo.

Intimately bound up though these external conditions are with the proper conduct of the whole series of events whereby the organism comes gradually to resemble the parents that gave it birth, they can only operate in conjunction with internal factors which must be sought for not only in the initial structure and constitution of the germ-cells, but in the mutual interactions of the developing parts. CHAPTER IV

INTERNAL FACTORS

1. THE INITIAL STRUCTURE OF THE GERM AS A CAUSE OF DIFFERENTIATION

§1. THE MODERN Fomu or THE PItEI<‘0RMA'[‘IONlS'1‘ DOCTRINE.

As has been already pointed out, it was in the hands of Roux that the principle of germinal localization first advanced by His assumed the rank and importance of a theory of development. On this ‘ Mosaik-Theorie’ of self-differentiation the precise relation observable between the several parts of the embryo and certain deﬁnite regions of the undeveloped germ is not merely customary or normal, but necessary and causal. The germ-cell is endowed with a preformed structure corresponding to the structure of the organism which is to arise from it; each part of this structure is predetermined for the formation of some particular member of the embryo, and out of it no other member can, under ordinary circumstances, be made. The causes of its development, regarded as a speciﬁc activity of the organism, as leading to the production of a form which is like that of the parents which gave it birth, lie wholly within this prc—cxisting structure, and of each part within each part. Although the influence of the environment and, in later stages, of the parts on one another is not entirely excluded, the factors on which the differentiation of the whole and of each part depends are essentially internal, and all that happens is that by a continued process of cell-division the parts are separated from one another and the structure thus made palpable and manifest. Cell-division in the embryo is therefore qualitatively unlike.

The supposed proof, by Pﬂiiger’s experiment with forcibly inverted Frogs’ eggs, of the complete ‘isotropy’ or equivalence of all parts of the cytoplasm, compelled Roux to locate the IV. 1 INITIAL STRUCTURE OF THE GERM 159

selfdifferentiating substance, the Idioplasson, in the nucleus, a. view adopted and elaborated by Weismann, while the facts of regeneration. necessitated the assumption of a reserve Idioplasson endowed with the potentialities of the whole, which is not qualitatively divided during development, but handed on intact to some or all of the tissues of the body.

The hypothesis then assumes the existence in the nucleus of the fertilized ovum of as many separate units as there are separately inheritable characters in the embryo, arranged there according to a plan which conforms to the structural arrangement of the parts which they represent, the separation of these units by continued ‘unlike’ nuclear division, and their distribution to the cytoplasm, where they determine the formation of the structures to which they are beforehand assigned.

For Roux the theory was no mere speculation, but the result of observation and experiment. The natural occurrence of half or partial monsters, the correspondence of defects in the embryo to injuries to the egg, and the coincidence of the first furrow with the sagittal plane in a large percentage (80 Z) of, unfortunately, only a small number of eggs, suggested the selfdiiferentiation of predetermined parts, while the last-mentioned observation led directly to the experiment in which one of the ﬁrst two blastomeres of the Frog’s egg was killed and a halfembryo produced from the survivor.

By means of a hot needle Roux succeeded in at least partially destroying one of the ﬁrst two blastomeres. The other continued to segment, and passed through the ordinary stages; ﬁrst the Hemimorula, with animal and vegetative cells and a segmentation cavity (though the last was sometimes absent), then by continued cell-division the Hemiblastula, followed by the Hemigastrula and Hemiembryo. In all cases the living tissues formed an exact half——either a right half or a left halfof a normal embryo, and ended abruptly at the (sagittal) plane of separation of the living and the dead. The segmentation cavity, however, was observed to extend in some cases across this plane, while in others it was confined wholly within the living cells. ‘

In the Hemigastrula the greatest extent of the archenteron—160 INTERNAL FAC’1‘O1tS IV. 1

produced by the overgrowth of the blastoporic lip—-was parallel to the plane of separation: the yolk-cells were pushed into the segmentation cavity. The Hemiembryo had half a medullary

9

FIG. 76.—A and B. Meridional sections through Herniblustulae of the Frog: kn, remains of nucleus; In’, large reticular nuclei; 1:, nuclei In the yolk; F,b1astocoe1; 4*, vacuoles. C. Hemigastrulalatemlis. Oblique lon itudinal section. I). Hemiembryo sinister, transverse section; .93, me ian plane ; the right half of the egg is already completely cellulated, post-generation of the germ-layers has begun; ch, notochord, of whole size; mw, medullary fold; u, gut; j, two yolk-cells that have remained embryonic. (From Korschelt and Heider, after Roux.)

tube, mesoderm on one side, half a gut cavity—open towards the dead blastomere——but a whole notochord (attributed by Roux to

premature regeneration). IV. I INITIAL STRUCTURE OF THE GERM 161

Anterior half-embryos—occurring when the second furrow by an ‘ anachronism ’ appears first-—were also observed, but posterior never. Quarter and three-quarter embryos were obtained by killing three or one of the first four cells, and Hemiblastulae superiores by killing the four yolk-cells in the eig-l1t~celled stage.

The self-differentiation of the ldioplasson is thus demonstrated. Subsequently, however, the reserve Idioplasson comes into play, and the missing half is formed by a peculiar process, to which

FIG. 77.——A and B. Normal Frog embryos with medullary folds (m), open (A) and closed (B). C. Hemiembryo dexter with almost complete post-generation of the eotoderm; u,yo1k-plug. D. The same, older, but with less post-generation. E. Hemiemhryo anterior (‘i’) with beginning post-generation. (From Korschelt and Heider, after Roux.)

Roux has given the name of ‘post-generation ’ to distinguish it from regeneration, in which lost parts are formed out of already (liiferentiated tissue.

The injured blastomere first degenerates, is then reorganized, and ﬁnally post-generated.

In the ﬁrst of these phases the cytoplasm and yolk are seen to be vacuolated, while the injured but not completely killed nucleus has given rise to small and large nuclei of normal structure

Jmxxntson M 162 INTERNAL FACTORS IV. I

surrounded by masses of pigment and in addition to irregular chromatic masses —some pale, some deeply staining. Separating it from the uninjured half is athin layer of yolkless protoplasm.

Reorganization is accomplished by any one of three methods. The ﬁrst is nucleation followed by cellulation, that is to say, masses of cytoplasm arise round normal nuclei, some of which are derivatives of the injured nucleus, while others have migrated across from the living blastomere. The second method involves an abundant immigration of whole cells from the living embryo which resuscitate or feed on the yolk and nuclei of the injured half. In the third method overgrowths take place at various points of the tissues of the living half-embryo, the cells dipping into and feeding on the yolk as they pass across.

In post-generation, which only occurs after the ﬁrst method of reorganization, each layer of the uninjured reforms the corresponding layer of the injured, the cells of the former exerting a directive stimulus upon the reorganized tissues of the latter, which thus pass through a process of ‘ dependent ’ differentiation. The ectoderm grows over from in front backwards and from below upwards, so that a structure superﬁcially resembling a. yolk-plug is formed at the hinder end; as it does so it becomes diﬁerentiated into the usual two layers. The medullary tube is formed in like manner. The mesoderm is formed from the ventral side and later divided into vertebral and lateral plates. The notochord was a whole from the beginning. The missing half of the gut is formed directly from the half-gut of the living embryo, not by any process of blastoporic invagination.

The conclusions drawn by Roux from this experiment have already been stated. Development is conceived of as a process governed essentially by a factor which is entirely internal, the preformed structure of the nucleus of the germ. Into the validity of this factor we have now to inquire by examining the evidence oifered by the very numerous and similar experiments, performed on the eggs of animals of all kinds, which the ‘ Mosaik-Theorie’ has evoked. We shall take these experiments in order, beginning with the form employed by Roux himself, the common Frog 1.

‘ For literature see following section. IV. I INITIAL STRUCTURE OF THE GERM 163

§ 2. AMPHIBIA.

It will be remembered that the Frog’s egg possesses, when freshly laid, a symmetry which is radial about the axis, the line joining the centres (animal and vegetative poles) of the pigmented and unpigmented portions of the egg; the symmetry is marked internally by the arrangement of yolk and protoplasm, for the latter lies most abundantly in the smaller unpigmentcd, the former mainly in the larger pigmented portion. Since the yolk

FIG. 78.~—Formation of the grey crescent in the Frog's egg (R. temporaria). A, B from the side; 0, D from the vegetative pole. In A, C

there is no crescent, in B, 1) a part of the border of the pigmented area has become grey.

is heavier than the protoplasm the white pole is turned downwards, with the axis vertical, the egg being free to rotate inside its jelly membrane. Shortly after fertilization, however, the radial is replaced by a bilateral symmetry (in Rana ﬂwca and I2. temporm-z'a), a grey crescent being formed on one side of the egg along the border of the pigmented area by the retreat of pigment into the interior (Fig. 78). The grey crescent eventually becomes white and added to the white area. In Rama caculenta M 2 164 INTERNAL FACTORS IV. 1

it is stated that the egg‘-axis becomes inclined to the vertical. This would seem to be an error, due to a confusion between the original white area and the white area enlarged by the addition

FIG. 79.—Diagrams of the closure of the blastopore in the egg of the common Frog (R. tempo:-aria). In A~—D the egg is viewed from the vegetative pole, in E, F from below. The dorsal lip is at the top of the ﬁgures. In D the ventral lip has just been formed and the blastopore is circular. In E the rotation of the whole egg has begun, and in F is complete.

of the grey crescent (compare Fig. 44 with Fig. 78). The egg is now bilaterally; symmetrical about the plane which includes the egg-axis and the middle (‘point of the crescent. IV.1 INITIAL STRUCTURE OF THE GERM 165

Since the crescent is formed on the side opposite to the entry of the spermatozoon, the latter is considered by Roux to determine the symmetry-plane of the egg. The ﬁrst furrow is also stated to generally lie in this plane, and the dorsal lip of the blastopore, which marks, of course, the sagittal plane of the embryo, appears in the region of the grey crescent. The dorsal lip grows through 75°, starting 25° below the equator and passing beyond the vegetative pole ,- with the rotation of the whole egg and therefore also of its axis in the direction opposite to that through which the dorsal lip has travelled, the anterior end of the embryo, whose dorsal side is now uppermost, comes to lie a little above and behind the animal pole, while its posterior end is marked by the now fast-closing blastopore (Fig. 79).

According to Roux the ﬁrst furrow and the sagittal plane coincide and the ﬁrst two blastomeres are right and left, unless, by an anaehronism, the second furrow, which separates anterior from posterior (this should be dorsal from ventral), occurs ﬁrst. The third furrow, therefore, separates dorsal from ventral (correctly speaking, anterior from posterior). In the small number of eggs examined by Roux the percentage of coincidences was found to be high (80 %) and the deviations were attributed to experimental error.

Oscar Hertwig has, however, stated—-on the strength of a small number of observations on eggs compressed between horizontal glass plates—that the angle between the two planes may have any value; while Schulze and Kopsch think it probable that they coincide in the majority of cases. The recent examination by the present author of a large number of cases has shown that the angle in question may indeed have any value from 0° to 90°, but that there is a decided tendency to small values, that is to coincidence, as may be seen from the annexed frequency polygon (Fig. 80), although at the same time there is no correlation, and therefore no causal connexion, between the two planes. Between the plane of symmetry and the sagittal plane, however, the correlation is considerable, and the tendency of the two to coincide much greater (Fig. 81), a result in conformity with the statements of other observers (Roux, Schulze, Morgan). Within certain limits, therefore, the right and left 166 INTERNAL FACTORS IV. 1

halves, the dorsal and ventral sides (since the dorsal lip always

appears in the region of the grey crescent) and the anterior and posterior ends (since the anterior end is always a little above the

-90-oo -10 -so -50 -40 -50 -20 -no

o no '20 '30 Ho 050 +60 910 +00 *9!)

FIG. 80.—-Frequency polygon of the angle between the ﬁrst furrow of the Frog's egg and the sagittal plane of the embryo. n = 889, M = 2-12°

1-914, 0- = 40-39°:-646.

animal pole) are predetermined in the undivided egg. But the manner in which the material of the egg is cut up by the first two meridional divisions makes no diiference to the result. In IV. 1 INI'1‘IAL STRUCTURE or THE GERM 167

these two divisions the qualitatively distinct parts of the cyto. plasm may be separated from one another in any assignable manner, the nuclear material being distributed to these in any assignable order. The symmetry of segmentation has no relation

H0

I00

70

60

Frequency.

50 ‘IO

30

ID

-90 -80 -70 -no 50 40 '30 '20 -10 o 0-Io v20 950 +40 950 can #70 0-60 9-90

FIG. 81.—F1-equcncy polygon of the angle between the plane of symmetry of the Frog's egg and the sagittal plane of the embryo. n = 509, M = 2-23:-889, rr = 29-75:-629. either to the symmetry of the embryo or to the symmetry of the undivided egg, for the first furrow tends to lie either in or at right angles to the plane of symmetry and is not correlated with

it to any great extent‘. The third furrow, however, being‘ always ‘ See, however, Appendix A. 168 INTERNAL FACTORS IV. I

at right angles to the axis, invariably separates anterior from posterior.

An observation of Morgan’s has some bearing on this question ; even when the first furrow divides the egg unequally a normal embryo results.

Even when the ﬁrst two blastomeres are equal the cells produced by their subsequent division may migrate across the plane of separation (Kopsch). In Nectmws there appears to be no relation between the first furrow and the sagittal plane (Eycleshymer).

The hypothesis of qualitative nuclear division has been tested

by O. Hertwig in another way.

FIG. 82.—'l‘he first four phases of segmentation of the F1-og’s egg. Normally (the three upper ﬁgures) and under pressure between horizontal plates (the three lower ﬁgures) I, II, 111, 1 V, the ﬁrst, second, third, and fourth divisions. 1-16, the cells produced by successive (livisions, numbered as follows : The egg divides into

1”5 3 7 2 6 4J8

1 95'T4:?'T77 52'1“06164?28”'ﬁs

The animal cells in the normal egg and the corresponding cells in the compressed egg are stippled. (After 0. Hertwig’s account.)

The eggs are aﬂixed to glass plates by their jelly, fertilized, and allowed to rest until the axis has become vertical. Pressure is then applied by a second plate, in the direction of the axis, the plates being horizontal, or at right angles to the axis, the plates being vertical or oblique. In such compressed eggs the direction of elongation of the nuclear spindles is distorted, and IV. 1 INITIAL STRUCTURE OF THE GERM 169

consequently the ‘qualitatively unlike’ nuclei compelled to assume an abnormal arrangement. Between horizontal plates the ﬁrst two furrows are, as normally, meridional and vertical (Fig. 82) ; the third, however, is parallel to the first (instead of latitudinal), and the fourth latitudinal (instead of meridional).

Between vertical plates the first is meridional and at right angles to the plates, the second latitudinal (horizontal) and near the animal pole, instead of meridional, the third furrows are parallel to the first, and the fourth meridional and at right angles to the first in the four upper blastomeres.

Now if nuclear division is a qualitative process, if, further, the nuclei of the compressed egg are divided in the same way in successive mitoses as are the nuclei of the normal egg (that is to say if the ﬁrst two nuclei are right and left, each of these then divided into dorsal and ventral, and each of these again into an anterior and a posterior), then as a result of compression their distribution and arrangement must be altered (as the ﬁgures show), parts which should be anterior will be lateral or ventral, and vice versa, and a monster will be formed. Such eggs give rise to normal embryos. Nuclear division is therefore not qualitative. From this conclusion there is only one escape; it may be argued that the orientation of the nuclei remains unaltered when the direction of the spindles is changed by the pressure, and that therefore the order of sequence of the cell-divisions may be varied ad libitum, may be as ‘ anachronistic ’ as is pleased, without affecting in the least the mode of distribution of the, qualitatively unlike, parts, a contention, surely, which would only be urged for the sake of supporting a thesis.

0. Hertwig has also repeated Roux’s experiment, killing, or at least injuring, one of the ﬁrst two blastomeres by a needle or by means of electricity. The uninjured half segments to form a mass of cells lying on the top of the dead blastomere, as a blastoderm lies on the yolk of a meroblastic egg, and separated from it by a segmentation cavity, although the latter may be wholly within the living cells. This is Roux’s Ilemiblastula, but in Hertwig’s case the dead cell lies below, a point, as we shall see, of considerable importance. Later on a blastopore is formed, but always, according to Hertwig, within the bounds of 170 INTERNAL FACTORS IV. 1

the living portion, not at the edge; the blastopore is usually symmetrical to the plane of separation of the two blastomeres. Beneath the lip of the blastopore an archenteron is formed, and notochord and mesoderm are differentiated. The closure of the blastopore is, however, prevented by the resistance oﬁered by the dead yolk-mass, which lies ventrally and posteriorly, just as it is retarded by the yolk in a large-yolked Fish egg; in fact, were this dead yolk removed, the living portion, Hertwig maintains, would develop normally. A nearly normal embryo may, in fact, be formed, but more usually there are considerable abnormalities. The mass of dead yolk, though partially enclosed by the growing edge of the ectoderm, is sufficiently great to impede the ultimate closure of the blastopore, and sometimes of the medullary folds as well; the latter frequently diverge round the yolk-plug of dead and living tissue, the chorda being split as well; they may be symmetrical, or one side may be much less developed than the other, owing, it is asserted, to an asymmetry in the resistance offered by the dead cell, and in the more extreme cases this inequality may be so pronounced as to give rise to a condition which does not differ in any way from Roux’s Hemiembryo lateralis ; there is one medullary fold, a notochord, mesoderm on one side, and a gut cavity in the yolk-mass ; to the bare side of the yolk-cells is attached the dead blastomere, only partially covered by extensions of the cctoderm of the living half. Such cases are, however, very rare; in the majority more, sometimes much more, than a half is produced from the living blastomere, for Hertwig denies that the missing parts are post-generated, as Roux maintains, though he admits the overgrowth and immigration of cells, as well as the persistence of living nuclei where the injury has been only partial ,- all these contribute to the formation of the embryo, which would be complete were its development not hindered by the presence of the inert mass of yolk.

The diiferences of interpretation put by Roux and Hertwig on the same phenomena appear to be radical. A reconciliation is, however, possible, for Morgan has observed that the whole or half development of the injured cell depends upon the position it takes up. If the original position—with the black pole IV.J INITIAL STRUCTURE OF THE GERM 171

uppermost-—is retained, then a half-embryo is formed, as is indeed the case, according to Hertwig’s own evidence, when the egg is prevented from turning over by compression.

In such half-embryos Morgan ﬁnds no traces of Roux’s postgeneration, although a whole embryo may be eventually formed by regenerative processes, referred by the author to the retarded development of the living parts of the injured half. It should be noticed, however, that Roux’s statements are conﬁrmed by Endres and Walter.

Should, however, the white pole be uppermost, then a whole embryo of half-size results. In Hertwig’s experiments, of course, the original position was usually not retained.

FIG: 83.-.~—Doub1e embryo of Rana fusca, from an egg compressed in the direction of the axis and inverted in the two-cell stage. (After Schultze, from Korschelt and Hcider.)

A B

FIG. 84.—Double monsters of Rana fusca, obtained by the same method. (After Schultzc, from Korschelt and Heidcr.)

This conclusion is still further strengthened by Schultze’s observations on the development of eggs inverted in the twocelled stage and kept so. Each blastomere develops independently and a double monster is produced (Figs. 83, 84). The result is not due to the pressure, for controls similarly compressed developed normally.

The details of development of these eggs have been carefully worked out by Wetzel (Figs. 85, 86). In each inverted blasto172 INTERNAL FACTORS IV. I

mere the yolk sinks next to the plane of separation, while the protoplasm and pigment rises to the outer side. The two cells are now related as two whole eggs united by their vegetative poles, their axes in one and the same (horizontal) straight line; each has, in fact, acquired a totally new polarity of its own. When segmentation has been completed a groove appears, in the plane of separation, and gradually extends round the whole circumference ; the ends of the groove are forked, but the branches of each fork unite as the groove grows round. The groove is, in fact, a blastoporic lip common to the two, the branches the individual lips, the material between them and ﬁnally covered over by them a common yolk-plug, the space between the two

FIG. 86.——Section through a. double blastula of the Frog ' (Rana fusca). Ia, blastocoels. FIG. 8-3.—-Double embryo ob- (After Wetzel, from Korschelt

tained by the same method: h, and Heider.) heads; 1», metlullary grooves;

c, line of union of the latter.

(After Wetzel, from Korschelt

and Heider.)

a common archenteron extending into an archenteric space in each individual. Subsequently medullary folds and notochord are developed in each below the groove, that is on the vegetative or postero-dorsal side of each, but anteriorly each grows out free of the other, and in this region medulla, notochord, and gut are single. The result is‘, therefore, two embryos placed back to back, and united by a common yolk-plug.

Experiments in which the four animal or the four vegetative cells of the eight-celled stage are killed are not very conclusive, as the develonment of the survivors does not go very far. IV. I INITIAL STRUCTURE OF THE GERM 173

Samassa has, however, shown that a short archenteron with dorsal lip, together with traces of notochord and mesoderm, may be formed from the four animal cells alone, the dead yolk-cells making a ﬂoor to the archenteric cavity and protruding as a large yolk-plug; and Morgan has obtained a similar result with the four vegetative cells‘ alone.

From all these investigations it seems reasonable to infer that each of the ﬁrst two blastomeres of the Frog’s egg may under certain circumstances acquire the polarity of a whole, and be capable of giving rise to an embryo whose complete development is only prevented by the impediment offered by the presence of the other, whether living or dead. Were it possible to completely separate the two blastomeres, we may surmise that each would become a perfect embryo: a surmise which is raised to a certainty by our knowledge of what happens in the newt. For in this form it is possible to separate the two cells by means of a noose of ﬁne hair tied round a __,,.. the egg in the plane of the first ' i‘ ’ ' _.,furrow, as Herlitzka has shown, },‘1G_ g7__Egg of the newt

and in case gach half seg-- (Triton cristatus), tW0 (E0111pletely normal embryos, obtained

ments as a whole and develops by tying a thread W) round the

into a whole larva of rather more egg the f'li1I‘r0iv§'é ky,ijc1Iy - - - _ mem l'2].:n(3. er erl Z a, .l'Ol'l1 than ha’It'S1ze (F135 87)‘ H01" Korschelt and Heider.)

litzka has further investigated the dimensions of the organs in these embryos ,- those of the me dulla and notochord he ﬁnds are the same as in embryos developed from a whole egg, those of myotomes and gut a little less. The size of the nuclei and cells in the medulla and myotomes is the same as in the whole (3%) embryo 1 ; the number of nuclei in the medulla is the same, in the myotomes one-half. He concludes that some organs need a certain minimum number of cells for their differentiation, while the cells must attain to certain dimensions.

1 It will be convenient henceforward to designate the whole egg, or the embryo or larva developed from it, by the symbol }, each of the first two blastomeres and the embryo or larva, whether complete or Incomplete, formed from it by —§, and so on; thus a % embryo means one which arises from three out of the ﬁrst four blastomeres. 174 INTERNAL morons IV. 1

The discovery of Herlitzka has been followed up by the researches of Spemann, who has employed the same method of constriction, but with variations of degree, direction, and time. These diﬁerences have given the most interesting results.

The ﬁrst furrow, according to this author, is usually at right angles to the sagittal plane and separates the material for the dorsal and ventral halves of the embryo ; only rarely do sagittal plane and first furrow coincide. Both cases were, however, experimentally investigated.

FIG. 88.—Three stages in the roduction of a double monster by strong median constriction of the §Iewt’s egg. (After Spemann, 1903.) a. Beginning of gastrulation; there is a separate lip in each half. b. I. and 1'. Med., edullary folds of left and right embryos; *, point where the medullary grooves separate; Bl, blastopore. c. The doubleheaded larva.

When the ﬁrst furrow is in a horizontal plane, a slight constriction in the two-celled stage separates the dorsal lip of the blastopore in one portion from the ventral lip in the other. Medullary folds are developed in the first half only, the second forms a sort of yolk-sac appendage which is later absorbed by the single normal embryo. With tighter, but still incomplete, constriction the dorsal half alone becomes an embryo; it contains either the whole or only the dorsal portion of the blastopore. The ventral half contains either no portion of the blastopore or IV. I INITIAL STRUCTURE OF THE GERM 175

only the ventral lip; it develops mesoderm but undergoes no further dilferentiation, and eventually drops oﬁ. Should the constriction be delayed until after the dorsal lip has appeared both halves may form an embryo, but the ventral embryo is usually imperfect. These dilferences are attributed by Spemann to differences in the time or place of constriction.

When a. transverse constriction is made after the appearance of the medullary plate the anterior half develops as a whole, with complete nervous system and optic vesicles; the posterior forms a medullary plate, but no folds, and dies. After the appearance of the medullary folds, however, the anterior and posterior halves produced by transverse constriction develop as halves, although a case is described where each had a pair of auditory vesicles.

When the sagittal plane coincides with the ﬁrst furrow, constriction in the two-celled stage gives rise to double-headed monsters; if the constriction is slight the most anterior organs only are involved (duplicitas anterior), the cerebral hemispheres, epiphysis, hypophysis, and paraphysis being doubled; there may be two complete pairs of eyes, or the median eyes may be more or less fused (Diprosopus triophthalmus). Tighter constriction brings about a reduplication of the chorda, auditory vesicles, and fore limbs (Dicephalus tetrabrachius). Similar effects are obtained by constriction in the blastopore stage, but not later; halves separated in the median plane when the medullary folds have arisen die as halves.

We shall now brieﬂy consider another experiment by which the independence of one another of the parts is demonstrated. Schaper removed the brain, eyes, and, probably, the auditory vesicles from newly hatched tadpoles of Rana esculenta. The wound healed up. The mouth moved to the anterior end and the suckers up the sides of the tadpoles, which lived for nearly a. week, in the course of which they grew 2mm. They then showed signs of weakness and were preserved. It was found that the mouth had opened, that labial cartilages, pterygopalatine bar, jaw-muscles, gill-bars, gills, heart and bloodvessels, trigeminus and vagus ganglia, oesophagus, pronephros and glomus, and dorsal muscles had all been diﬁferentiated. 176 INTERNAL FACTORS IV. I

There was, however, no operculum. The anterior end was occupied by a mass of mesenchyme. There was no sign of a regeneration of any of the lost organs except the anterior end of the notochord. The nerve ganglia were normal, but the spinal cord underwent degeneration. In spite of this the creatures could execute spontaneous and reﬂex movements.

What this experiment shows is, of course, that the organs of the trunk are not dependent for their development upon the presence of the brain; fresh researches would be necessary to determine how far in each case the parts are self-diﬁerentiating. In the second place the brain and eyes, when removed at this stage, cannot be remade by the tissues, but remain behind. So far, therefore, the body is at this stage an inequipotential system, as Driesch would call it. We know, however, that at an earlier stage the parts of the body are equipotential. We have, in fact, only another instance of that loss of totipotentiality, of that increase of independence and self-diiferentiation which takes place as development proceeds. Here, however, we are anticipating a conclusion which can only be completely stated after a discussion of the experiments performed on eggs of other types.

Ll'l‘ERA'l‘URE

H. ENDRES and H. E. WALTER. Anstichversuche an Eiern von Rana fusca, I'°' und lI*°' '1‘ei1, Arch. Eat. Mech. ii, 1896.

A. C. EYCLESHYMER. Bilateral symmetry in the egg of Nectum.9, Anat. Anz. xxv, 1904.

A. HERLITZKA. Contributo allo studio della capacitia. evolutiva dei due primi blastomeri nell’ uovo di tritone (Triton crislatus), Arch. Eur. Mech. ii, 1896.

A. HERLITZKA. Sullo sviluppo di embrioni completi da blastomeri isolati di nova di tritone, Arch. Ent. Mech. iv, 1897.

O. HERTWIG. Ueber den Werth der ersten Furchungszellen fiir die Organbildung des Embryo, Arch. mikr. Anat. xlii, 1893.

F. KOPSCH. Ueber das Verh-liltniss der embryonalen Axen zu den drei ersten Furchungsebenen beim Froseh, Iutemaf. Monatsc-hr. Anal. 14. Phys. xvii, 1900.

T. H. MORGAN. Half-embryos and who1e—embryos from one of the first two blastomeres of the Frog’s egg, Anat. Auz. x, 1895.

T. H. MORGAN and E. TORELLE. The relation between normal and abnormal development (iv) as determined by Roux’s experiment of injuring the first-formed blastomeres of the Frog's egg, Arch. Ent. Mech. xviii, 1904. ' IV. I INITIAL STRUCTURE OF THE GERM 177

T. H. MORGAN. The relation between normal and abnormal development of the embryo of the Frog, (v) as determined by the removal of the upper blastomeres of the Frog's egg, Arch. Ent. Mech. xix, 1905.

W. RoUx. Ueber die Zeit der Bestimmung der Hauptrichtungen des Froechembryo, Leipzig, 1883; also Ges. Abh. 16.

W. RoUx. Ueber die Bestimmung der Hauptriohtungen (lee Froschembryo im Ei und ﬁber die erste Theilung des Froscheies, Bresl. iirtz. Zeitschn, 1885; also Ges. Abh. 20.

W. ROUX. Uebcr die kiinstliche Hervorbringung halber Embryonen, Virchoufs A1'ch., 1888; also Ges. Abh. 22.

W. Roux. Ueber das entwicklungsmechanische Vermtigen jeder der beiden ersten Furchungszellen des Eies, Verh. Anal. Ges., 1892; also Ges. Abh. 26.

W. RoUx. Ueber Mosaikarbeit und neuere Entwicklungshypothesen, Anat. Hqfte, I" Abt., ii, 1892-3; also Ges. Abh. 27.

W. Roux. Uebcr die Speciﬁcation der Furchungszellen und iiber die bei der Postgeneration und Regeneration anzunehmenden Vorgiinge, Biol. Centralbl. xiii, 1893; also Ges. Abh. 28.

W. ROUX. Ueber die ersten Theilungen des Froscheies und ihre Beziehungen zu der Organbildung des Embryo, Anat. Anz. viii, 1893; also Gas. Abh. 29.

W. ROUX. Die Methoden zur Hervorbringung halber Frosch-Embryonen, uml zum Nachweis der Beziehung der ersten Furchungeebenen des Froscheies zur Medianebene des Embryo, Anat. Anz. ix, 1894; also Ges. Abh. 31.

W. Rovx. Ueber die Ursachen der Bestimmung der Hauptrichtungen des Embryo i1n Froschei, Anat. Anz. xxiii, 1903.

P. SAMASSA. Studien ﬁber den Einﬂuss des Dotters auf die Ga.strula.tion und die Bildung der primiiren Keimbliitter der Wirbelthiere : II. Amphibien, Arch. Ent. Mech. ii, 1896.

A. SCIIAPER. Experimentelle Studien an Amphibienlarven, Arch. Em‘. Mech. vi, 1898.

O. SCHULTZE. Die kiinstliche Erzeugung von Doppelbildungen bei F1'oschla.rven mit Hilfe abnormer Gravitntionswirkung, Arch. Ent. lllech. i, 1895.

0. SCHULTZE. Ueber das erste Auftreten der bilateralen Symmetric im Verlauf der Entwicklung, Arch. mikr. Anat. lv, 1900.

H. SPEMANN. Entwickelungsphysiologische Studien am Triton-Ei, I, II, III, Arch. Ent. Mech. xii, xv, xvi, 1901, 1903.

G. WE'rzEI.. Ueber die Bedeutung der cirkuléiren Furche in der Entwicklung der Schultze'schen Doppelbildungen von Roma fusca, Arch. mikr. Anat. xlvi, 1895. 178 INTERNAL FACTORS IV. 1

§3. P1scEs.

Morgan has shown that in the fishes Ctenola/)ru.9, Serramw, and Fmzdulus there is no deﬁnite relation between the first (or second) furrow and the median plane of the embryo.

In Fumiulus when one of the first two blastomeres is removed the other gives rise to a perfect embryo of more than half-size. In segmentation the first furrow is in the plane of what would be the second furrow of the whole egg, the second in that of the third, and the third more or less in that of the fourth ,- but as the successive furrows are at right angles to one another in both cases it is permissible to assert that the segmentation of the half blastomere is total.

Some of the protoplasm of the half that has been removed ﬂows in underneath the other, is nucleated by it and added to its mesoderm. The size of the nuclei is the same in % and -} embryos.

Morgan also found it possible to remove two-thirds of the yolk, but not more, without interfering with the normal development of the whole egg.

Injury to the germ—ring on one side of the dorsal lip resulted in a deﬁciency in the mesoderm of that side posteriorly ; anteriorly both sides were equally well formed.

So Kastschenko has shown for Elasmobranchs and Kopseh for Teleostei and Elasmobranehs that injury to the lip of the blastopore will only interfere with the normal bilaterality of the embryo when effected quite close to the middle line.

A series of experiments similar to the last has been carried out by Sumner on the eggs of various ﬁsh (principally 1*'zmrlulus, also Ewocoetzza, Salvelinus, and Batrac/ms).

By means of needles inserted into various parts of the blastederm the exact mode in which the material of the latter is used in the formation of the embryo is ﬁrst determined. It is shown that the material for the embryo is brought into position not by a conerescence of the lateral lips of the blastopore (sides of the germ—ring), but rather by an axial concentration of the cells originally situated in that ring at the posterior margin or dorsal lip, the cells so concentrated being continually pushed forwards in the middle line until the anterior end of the embryo comes to lie near the original centre of the blastoderm. IV. 1 INITIAL STRUCTURE OF THE GERM 179

Injuries to various parts of the blastoderm and its margin (the germ-ring) prove that it possesses a certain degree of ‘isotropy’.

Thus the entire embryonic region of the posterior margin was destroyed by electrocautery. The adjacent edges closed up and completed the germ-ring ; at the posterior point of this a new embryonic shield was formed, in which ectoderm, endoderm, and notochord became differentiated. Or, again, a lateral piece of the germ-ring immediately adjacent to the embryo was removed without preventing the formation of a normal, bilaterally symmetrical embryo, although, as Morgan fou11d, the mesoderm might be deﬁcient posteriorly on the injured side.

LITERATURE

N. KASTSCHENKO. Zur Entwickelungsgeschichte des Selachierembryos,

Anat. Anz. iii, 1888. F. KoPscH. Experimentelle Untersuchungen iiber die Keimhaut der

Salmoniden, Vcrh. Aunt. Gesell., 1896. F. KOPSCH. Experimentelle Untersuchungen am Primitivstreifen

des Hiihnchens und an Sc;/Ilium-Embryonen, Verh. Auat. GeseIl., 1898.

T. H. MORGAN. Experimental Studies on Teleost eggs, Anat. Anz. viii, 1893.

T. H. MORGAN. The formation of the ﬁsh-embryo, Journ. Morph.

x 1895. ’F. B. SUMNER. A study of early ﬁsh-development, experimental and

morphological, Arch. Em‘. Mech. xvii, 1904.

§ 4. Amrmoxus.

In this form the blastomeres may be separated by shaking. Their development has been observed by Wilson, whose account

‘ 8 '’GD

0

FIG. 89. A. Normal } gastrula. of Amphioams. B. Gastrula from blas tomere. C Gastrula from a i‘ blastomere. J). Gastrula of normal size from an egg-fragment. (After Wilson, from Korschelt an Heider.) has since been conﬁrmed by Morgan. The isolated blastomeres soon become rounded; the ﬁrst division is always transverse to the long axis.

N 2 180 INTERNAL FACTORS IV. 1

A i‘ blastomere segments like a whole ovum, except that the second division may be unequal, and gives rise to a normal blastula, gastmla, and embryo of half the normal size; % blastemeres behave in the same way (Fig. 89). Incomplete separation of the cells after the first or second division leads to double embryos in the ﬁrst case, double, triple, or quadruple embryos in the second. The double embryos may make any angle with one another and continue to live for some time (Fig. 90).

A % blastomere segments like a whole egg (the second division is, however, always unequal) and gives rise to a normal blastula and gastrula, more rarely to an embryo of one-quarter size.

FIG. 90. Double nrastrulae of Amphioams, from incompletely separated

blastomeres. 1;‘), u’, separate blastopores; u, common blastopore.

(After Wilson, from Korschelt and Heider.)

A % blastomere will segment normally, or nearly so. A blastula is rarely formed, usually an open curved plate of cells, which are ciliated and swim about but do not gastrulate.

Lastly 115 blastomeres divide but produce only an irregular heap of cells.

It is obvious that the potentiality of a blastomere to form a whole embryo diminishes with its germinal value, the ratio it bears to the whole ovum; but it is not so clear whether this diminished capacity is due to the lack of some necessary, speciﬁc, organ-forming substance or merely to the small size and lack of IV. I INITIAL STRUCTURE OF THE GERM 181

undifferentiated material. Morgan has attempted to answer this question by counting the number of cells in the various organs of whole and partial larvae. It appears from his estimate that the number of cells in the archenteron, in the notochord and in the nerve-cord of -1-, 5, and 71,- larvae is constant, and that though the size of the whole body is less, the dimensions of the notochord and nerve-cord are about the same in all three. It would seem, therefore, that there is a minimum size and a minimum number of cells necessary for the formation of these organs. Whether the failure of the % blastomere to develop into a larva is, however, due to more lack of material or the absence of some speciﬁc substance needful for the development of some particular organ is still undecided ; for the 1% and :1; cells contain substance from both the animal and vegetative portions of the egg, while the §- are composed of one or the other substance exclusively, the furrows of the third phase being equatorial.

LITERATURE

T. H. MORGAN. The number of cells in larvae from isolated blastomeres of Anzphioxzts, Arch. Ent. Mech. iii, 1896.

E. B. WILSON. Amp7n'0.z'us and the Mosaic theory of development, Joum. Morph. viii, 1893.

§ 5. COELENTERATA.

Many years ago Metschnikoif observed a perfectly normal separation of the blastomeres at a certain period in the development of certain medusae (Oceania). If when the cells reunite in a later stage the order of their rearrangement is not constant, the egg-substance must be in some measure isotropic.‘

More recently the equipotentiality of the blastomeres of these forms has been experimentally demonstrated by Zoja (Fig. 91).

The cells were separated by means of a needle. -3 and :1blastomeres (in the case of Lz'rz'ope, Gem/onia, Mitrocoma, Clytia, and Laozlice) and % and 1-15 blastomeres in the case of the two last mentioned will give rise by normal segmentation to blas ‘ It should not be forgotten that in 1869 Haeckel had cut the blastulae of Crystallodes (a Siphonophor) in pieces and obtained from the larger fra ments normal larvae. The development of small pieces was retar ed and abnormal (Zur En!wickelimysgeschichte der Siplconophovm, Utrecht, 18%). 182 INTERNAL FACTORS IV. 1

tulae with closed blastocoel and these to normal Planula larvae. In Olytia a hydroid was eventually reared and in Liriope a medusa with four primary tentacles from 73 and 3%; cells. It is to be noticed that, since the third division in these eggs is equatorial, the ,1, and 1% blastomeres must be either animal or vegetative.

Zoja also ﬁnds that in Laodice and Clytia the number of cells in partial larvae is proportional to the germinal value. In Li;-iope the number of cells in the endoderm is equal to that in the whole larva.

35%

FIG. 91. Partial development

in Coelenterata. (After Zoja, 1896.)

a-cl. Laodice cruciata: a,§ blastomeres: 12, § blastomeres ; c, blastula from blastemere; d, larva during formation of endoderm. e—l. Liriope mucrouara. c— 2'. Development of blastomere : e, division of blaste more into f; -§-; g, -,"5,;; h, end ofendoderm formation in embryo; 1', medusa. reared from blastomere. j—l. Development 01} blastomere: j, 3; k, ,4“; Z, embryo with endodenn.

7

LITERATURE

E. METSCHNIKOFF. Embryologische Studien au Medusen, Wien, 1886. R. ZOJA. Sullo sviluppo dei blastomeri isolati dalle uova di alcune meduse, Arch. Ent. Mach. i, ii, 1895-6. IV. 1 INITIAL STRUCTURE OF THE GERM 183

§ 6. ECHINOD]*Jlh\IA'1‘A.

Easy to obtain in large numbers and eminently suitable for experiment, the eggs of the sea-urchins and starﬁshes have provided the analytical embryologists with abundance of material; and it is in this group that the possibility of rearing a perfect larva from a single blastomere was first demonstrated by the classical researches of Driesch.

The experiments which have been carried out, mainly by Driesch, but also subsequently by others, are very numerous and fall into two principal classes.

In the first the development of parts—wheth_er isolated blastemeres or groups of blastomeres, fragments of unsegmented eggs fertilized or unfertilized, or pieces of blastulae and ga.strulae—has been observed, the rate of their development recorded, and the relation of the number of cells and dimensions of the partial larvae to their germinal value determined. By the second series of investigations the effect of alteration of the type of nuclear or cell-division and consequent displacement of the blastomeres upon the subsequent development has been discovered.

Before discussing these experiments, however, a word must be said as to the normal behaviour of the whole egg.

The structure, segmentation and formation of the primary germinal layers have been very completely studied by Boveri in the sea-urchin, St1'o2z_qyZocenh'o6u8 livirlus (Fig. 92). The axis of the ovarian egg is marked by the exeentric position of the nucleus ; at the point on the surface nearest which it lies the polar bodies are formed, and this point is the animal pole. At this point there is a fine canal (micropyle) in the jelly which surrounds the egg. After the extrusion of the polar bodies a brown pigment which had previously been uniformly distributed through the cytoplasm becomes aggregated in a dense but quite superﬁcial subequatorial zone, the upper somewhat ill-deﬁned border of which may, however, extend into the animal hemisphere. The pigment-free region left about the vegetative pole occupies from 7515 to -115 the volume of the whole ovum. The spermatozoon may, but need not, approach the egg by the mieropyle. The fertilization spindle lies in a plane parallel to the equator of the egg (the FIG. 92.——Norma.l development of the sea.-urchin, Sh-ongyloceulrotus l1'm'dus. (After Boveri, 1901.)

The animal pole is u permost in all cases, and in the ﬁrst two ﬁgures the jelly with the cana.i)(micropyle) is shown.

a, primary oocyte ; the pigment is uniformly eripheral.

b, ovum after extrusion of polar bodies: tiie pigment now forms a. subequatoi-ial band. The nucleus is ex-axial.

c, (1, first division (meridional).

e, 8 cells, the pigment almost wholly in the vegetative bla.ston1o1-vs.

j, formation of mesomeres (animal cells) by meridional division: the vegetative cells have divided into macromeres and micromeres.

g, blastula. h, mesenchyme blastula.

1', j, k, invagination of the pigmented cells to form the urchenteron of the gastrula. In j the primary mesenchyme is separated into two groups, in each of which, in A-, a. spicule has been secreted. In I: the secondary, pigmented mesenchyme is being budded off from the inner end of the archenteron. IV. 1 INITIAL STRUCTURE OF THE GERM 185

‘ karyokinetic plane’) and elongates at right angles to the spermpath, which thus lies in the plane of the ﬁrst, a meridional, furrow (a similar observation had been previously made by Wilson and Mathews on Towopuezzstzav).

The second furrow is likewise meridional and at right angles to the first, the third equatorial or a little nearer to one or other pole; the animal cells get only a very little of the pigment. In the next phase the animal cells divide meridionally and equally to form the eight mesomeres, while the vegetative cells divide latitudinally and very unequally into four large pigmented macromeres next the equator, and four small quite unpigmented micromeres grouped round the vegetative pole.

In the blastula stage all the cells are of the same size, vacuolated and ciliated, and the polarity of the egg is only determinable by the position of the pigment zone. In the next stage the clear vegetative cells derived (presumably) from the micro

FIG. 93.—Ec7n'nus: segmentation under pressure.

a, preparation for third division (radial); b, preparation for fourth division (tangential); b’, after fourth division; c, another form of the 8-cell stage (third division parallel to first); (I, the same after removal of the pressure. (After Drieseh, 1893.)

mercs wander in to form the primary mesenchyme~from which the first triradiate spicules are developed-—and this is followed by the invagination of the pigmented cells to form the arehenteron. Secondary pigmented mesenchyme cells are budded off from the inner end of the latter. The character and sequence of the divisions in segmentation are the same in other Echinid eggs, but the polarity is not marked by pigment, nor indeed recognizable till the micromeres have been formed, and again when the mesenchyme cells wander into the blastocoel.

By means of press11re——under a coverglass-—Driesch succeeded (E0/12'-mw) in altering the direction of the spindles and consequently of the cell-divisions (Fig. 93). The first and second furrows were at 186 INTERNAL FACTORS IV. I

right angles to one another and to the coverglass, but whether meridional or 11ot does not appear. The third furrows were again at right angles to the coverglass and at 45° to the ﬁrst two; a ﬂat plate of eight cells was thus formed. The next division resulted in the formation of sixteen equal cells still all lying in the same plane ; the formation of micromeres is thus suppressed. Such eggs gave rise to normal larvae.

In the case just quoted the egg-membrane was intact; but similar results were obtained when it was broken. The second furrow was sometimes at right angles to, sometimes parallel to, the ﬁrst, the third at right angles to both in the latter case, and the fourth

FIG. 94.—The effect of heat upon the segmentation of the Echinoid egg. a, b, c, d, four successive stages in the segmentation of the same egg ofEchinus; e, f, two successive stages in the division of the same egg of Sphaerechinus. (After Drieseh, 1893.)

parallel to the third ,- but if the pressure was released the blastemeres became rounded olf and the sixteen-celled stage consisted of two plates of eight cells each; one or two micromeres were sometimes formed. The development of these eggs is also quite normal. These results have been confirmed by Ziegler.

In this, as in the similar experiment of Hertwig on the Frog’s egg, the normal distribution and arrangement of the nuclei is interfered with without prejudice to the normality of subsequent development. Nuclear division cannot therefore, Driesch contends, be a qualitative process. Boveri has also shown in similar fashion that the order of segmentation may be disarrang-ed and . IV. 1 INITIAL STRUCTURE OF THE GERM 187

the formation of the micromeres suppressed in Stron_qylocem5rotzw and a normal larva still result. The relation, however, of the egg-axis, as determined by the pigment-ring, to the symmetry of the larva remains the same as in the undisturbed egg. Increased temperature, violent shaking, dilute sea-water, and calcium-free sea-water are all means by which Driesch has suc ceeded in disarranging the blastomeres of E’c/tinus in various ways (Fig. 94). Thus when the temperature was raised from 19° C. (the

FIG. 95.——Va.ria.tions in the segmentatioxi of Echimts mic;-otuberculntus produced by dilution of the sea-water. a, tetrahedral four-cell stage; I), eight cells, three premature micromeres; c, eight cells, two precocious microineres; (I, the same egg after the next division. the precocious niicromeres have divided unequally, two normal micromeres have been formed. (After Driesch, 1895.)

normal) to 31° C. the first two (not subsequent) blastomeres separated, though they sometimes reunited. The next division was normal, b11t with the next phase one or two blastomeres divided in a direction perpendicular to that of the others, or the direction of division was different in each. In the following stage the micromeres were wholly or partly suppressed. So also in dilute sea-vmter (20% fresh water) the third division was unequal 188 INTERNAL FACTORS IV. I

in two or more blastomeres, and in the following an excessive number of small cells was formed (Fig. 95). In aless dilute mixture (16% fresh water) the blastulac divided to form each two Plutei. Again by shaking the eggs in the eight-celled stage the blastomeres were disarranged and made to lie almost in one plane ; micromeres were, however, formed as usual from the vegetative cells, but not, of course, in the normal position (Fig. 96). In spite of these very considerable alterations in the size and positions of their constituent cells all these ova nevertheless gave rise to perfectly normal larvae.

By the use of calcium-free sea.-water either the mesomeres or the macromeres and micromeres were separated into two groups. In the latter case the larva had two guts, in the former a normal larva. resulted, so that, as Driesch points out, the ectoderm at the

FIG. 96.~—-Disarrangement of the blastomeres of Echimcs by shaking, a, eight cells; b, sixteen cells; notice that the micromeres are not close together. (After Driesch, 1896.)

antiblastoporic (animal) pole must in this case have been formed from the macromeres.

The conclusion to be drawn from these experiments seems obvious; not only the nuclei, but the parts of the cytoplasm too are equivalent; within at any rate fairly wide limits their normal arrangement may be disturbed without affecting in the least the normality of subsequent development; or, as Driesch phrased it, the destiny of a nucleus or blastomere is not determined by its original situation in the preformed structure of the egg, rather it is a. function of its position in the whole embryo to which that egg gives rise. Its character is decided not by its origin, but by its ﬁnal position.

Whether there really is a limit to the rearrangement is a question which must be reserved for future discussion. We are 4 IV. I INITIAL STRUCTURE OF THE GERM 189 A /‘

I

FIG. 97.—A and 0, formation of ex-ovates in the egg of Arbacia by dilution of the sea.-water; K, nucleus; m, egg-membrane. B and D, blastulae formed from A and C ; B becomes constricted into two blastulae, each of which gives rise to a. Pluteus; Dproduces a. single Pluteus. (After Loeb,from Korachelt and Heider.) 190 INTERNAL FACTClRS IV. I

also indebted to Driesch for a long series of experiments on the behaviour of isolated blastomeres and egg-fragments. In the forms which we have hitherto considered—the Vertebrates and Coelentera.———an isolated cell segments as a whole and gives rise to a total larva. In these Echinoderms the isolated blastomere also gives rise to a total larva, but its segmentation is partial; only after segmentation has been completed does the open blastula. close up and resume the polarity of a whole.

Various methods have been employed for separating the blastomercs. Loeb showed that in dilute sea-water (50 the egg

FIG. 98.-—Development of the isolated «} blastomere of Echinus micromberculatms-. 'l‘wo1n1eromeres,twonmcromeres, four mesomeresz a, §; b 4*

c, Hemiblastula: on the right 1s the remaining blastomere, dead. Driesch, 1892.)

swells, bursts its membrane and protrudes an ex-ovate which may be large and develop independently (Fig. 97). Driesch has used heat, pressure, violent shaking with fragments of coverglasses, and the calcium-free sea-water introduced by Herbst. The isolated 71; blastomere in Ea/ziuus becomes rounded and segments as a half, as though the other blastomere were still present. It forms four mesomeres, two macromeres, and two micromeres, IV. 1 INI'l‘IAL STRUCTURE OF THE GERM 191

and by division of these a hemispherical curved p ate, a halfblastula (Fig. 98). When, as may happen, the other astomere dies without being separated, it is embraced by the open side of the survivor. Only after the 32-celled stage is passed do the edges meet and close. A blastula of half-size is the result, which becomes later a gastrula an(l'a Pluteus of perfectly normal form but of half the normal size.

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FIG. 99.——Segmentation of an isolated } blastomere of Eolmms; :1, -1-, (2), Ag, c,h—145i o9I§e)m1cro1nere, one macromere, two mesomeres, d, 391;. (After l‘leSC , 8 .

7% blastomeres behaved in the same way, segmenting as parts, developingultimately into whole normal Plutei. Similarly -,1 blastomeres segmented partially (Fig. 99), with, in some cases, irregularities : two micromeres, for instance, were formed in the second division. The % blastulae will gastrulate, but progress no further.

A B

FIG. 100.—-A, Gastrula with meaenchymc cells and trimdiate spicules reared from a. 2; animal cell of Sphaerechinus. B, The same from a. 5- cell of Echinus: the gut is tripartite. (After Driesch, 1900.)

In the next stage—-% blastomeres—a diiference becomes noticeable between the behaviour of the animal and vegetative, cells. Both kinds.of cells will gastrulate, secrete triradiate spicules and 192 IN’I‘ERNAL FACTORS IV. 1

develop a normally tripartite gut (Fig. 100). But this only occurs in a small proportion of cases, smaller for the animal than for the vegetative cells. Further, the former frequently give rise to blastulae provided with a row of long cilia (Fig. 101), while the latter are delicate, and many die.

Boveri has pointed out that the ability of animal cells to gastrulate depends, in Stron_r/ylocemfrotus, on whether or not they contain a portion of the pigment-zone, and this on the position of the third furrow. Garbowski, however, denies that the pigment is itself an organ-forming substance. For in the ﬁrst place there is a variety of S51-ongyloceaztrotus livitlus, in which the pigment remains diffuse and never becomes concentrated to form a band at all ; and secondly, even though, when present, it is usually in the subequatorial position described by Boveri, this is not always so; it may be oblique to the egg-axis, or wholly in the

Era. l01.—Long-eiliatedblastula from animal» 01‘ Wh°11.Y in the a _} animal cell of Echimcs. (After vegetative hemisphere, D"es°h’1900‘) It would appear, therefore, either that it is merely associated with some other substance which has up to the present remained invisible, or that in the cases described by Garbowski animal cells, if they were pigmented, would gastrulate as readily as ordinary vegetative blastomeres, in which case the manner in which the pre-determined material is cut up in segmentation would be a matter of indilference.

The differences in the behaviour of the T15 cells are still more marked. The mieromeres will only divide to form a heap of about ten cells ; the mesomeres give rise to either long-eiliated blastulae or imperfect gastrulae, with or without skeleton and 1V.1 INITIAL STRUCTURE OF THE GERM 193

mesenchyme, and with an undivided archenteron (Fig. 102) ; the macromeres, on the other hand, become gastrulae provided with mesenchyme and spicules (Fig. 103).

Lastly, even 51-; blastomeres will occasionally gastrulate, pro~ babl y only if derived from a macromere. This gastrulation was

Fm. 102.—Echinus. Larvae reared from mesomeres (animal cells) of the 16-celled stage. a and b, mesenchyme gastrulae, a has spicules; r, gastrula without mesenehyme; d, a long-ciliated blastula. (After

Driesch, 1900.)

not, however, observed directly, but the germinal value of the smallest gastrulae found was calculated by a. method which will be described below. 517; cells will not reach the gastrula stage.

9 O (I. b 0

FIG. 103,- Two gastrulae of Erhinus reared from 3;, macromeres; both have mesenchyme cells, one has triradiate spicules. (After Driesch, 1900.)

The diﬁerences between the developmental capacities of animal and vegetative cells may be studied in another way. The four micromeres were removed by Driesch in the 16-celled stage; the remaining twelve meso— and macromeres produced a normal Pluteus (an experiment also performed by Zoja on Stro7zy_I/Zocenlrotvue). The eight (or four in the previous stage) animal cells alone, the eight vegetative cells alone also formed, in some cases, a perfect larva. Segmentation was in all these cases partial.

JENKINSON 0 19_4« INTERNAL FACTORS IV. I

From the fact that the four animal cells together may form a Pluteus while each individually fails to pass beyond the gastrula stage, Driesch has argued that this failure depends solely on lack of sufficient material, not on the want of any speciﬁc gut-forming substance, and has supported his contention by other evidence. By placing the eggs of lL'c/ﬁrms in dilute sea-water the third division was made unequal ,- the cells were then isolated in ealeium— free water. The large cells thus formed must contain either more of Boveri’s pigment-ring (or rather of the substance of the vegetative hemisphere) or else more of the animal hemisphere, probably the former, since a larger percentage gastrulate than is usual with animal blastomeres. According to Boveri, however, all ought to gastrulate if all contain the speciﬁc substance for the archenteron. Further, these large cells are able to form mesenchyme even when the gut is lacking although they possess the middle region of the egg in any case and may lack the micromere area.

Driesch also points out that a 7} blastomere and a § vegetative blastomere l1ave both the same amount of Boveri’s pigment-ring and mesenchyme area, the latter, however, only half as big a gut and half as many mesenchyme cells as the former, and lastly that »,‘;— and :—l% animal blastomeres will not only'gast1-ulate but form mesenchyme as well.

The limitation of the potentialities of ectoderm and cndoderm in later stages has been investigated also by Driesch—-by observing the development of pieces or fragments of the blastulac and gastrulae. These pieces are obtained by cutting or shaking. Loeb has employed dilute sea-water to make the blastula swell, protrude through its membrane and become constricted into two.

In Ea/dmes, S/1/merec/u'mw, and /18/erius, any piece of a blastula when first cut out is crumpled, but soon becomes rounded, and swims about and eventually gastrulates.

Monsters exhibiting a certain degree of duplicity have been produced by Driesch by shaking the egg of Arte/'ia..e in the two-celled stage (this produces apparently a partial separation of the blastomeres) and by placing the blastulac of 152'//imzs in diluted sea-water. In the latter case the gut is single, the skeleton double; in the former the gut is doubled, though the two may subsequently fuse, or even trebled, or may be IV. I INITIAL STRUCTURE 01*‘ THE GERM 195

merely forked. The two guts may be similarly oriented or turned in opposite directions, and it is interesting to observe that Driesch, who believes the ﬁrst furrow to coincide with the plane separating the two guts, accounts for the latter case by supposing that one blastomere had been rotated upon the other, so that their vegetative ends—the locus of the gut-forming substance-—faeed in opposite directions.

Neither the ectoderm of the early gastrula alone nor the endoderm is capable of giving rise to a larva, though the former can develope a stomodaeum, a statement conﬁrmed by Morgan.

The vegetative half of a gastrula of AS);/zaerec/ziizzm will give rise to a normal small Pluteus whether the cetoderm only, or together with it the tip of the archenteron, has been removed (Fig. 104) ; the animal half will not. VVhen the spicules, one or

Wﬁiel/‘

FIG. 104. —»The potentialities of the cells of embryonic organs; (1, I). The vegetative and aniinal portions of a gastrula of S1)/I(IeI'€(‘lIiIl1Is grauulm-is, cut equatori-ally in two; 0. Plut-eus reared from a fragment of a gastrula; (1. Normal tripartite pluteus gut. (After Dricsoh, 1896.)

J

both, are removed the skeleton is one-sided and the Pluteus eonsequently one-armed.

In As/elrias the arehenteron can form a new terminal vesicle and mesenehyme cells afresh when these have been removed (Fig. 105); but should the eoelom sacs be cut off ata later stage, the (secondary) archenteron can form no more, though it divides into the usual three portions. T

Parallel to the behaviour of isolated blastomeres is the segmentation‘ and development of egg-fragments, obtained by shaking, cutting, or (Loeb) dilute sea—water. The fragments may be taken from already fertilized eggs, or from unfertilized: in the latter case each, whether nucleate or enueleate, must be subsequently fecundated.

In segmentation there are a great many ditfereiices, which appear to depend on the nature of the fragment, that is to say, 196 on the part of the egg from which it

INTERNAL FACTORS

IV. I In

has been removed.

the case of the colourless egg of ]$’¢-/zz'm1.9 it is only possible to

FIG. 105.~-The potentialities of the cells of embryonic organs; a. N0l'1]1i1-l Pluteus of S'ph(wre(.'hiIms; b. Pluteus reared from a fragment of a gustrula; 0, 9. Normal Bipinnaria of A8tP1‘iflS _qlaciali.s-; «I, f. Bipinn-aria from the vegetative half of a gastrula; g. Larva. of Asterias with typical tripartite gut, but no coelom, from the vegetative half of a gastrula. removed after development of the eoeloni sacs. (After Dricscli, 1896.)

In another case wha.t appears to

guess at the nature of the piece by observing the mode of segmentation.

The ﬁrst and second divisions may both be equal, as also the third and fourth ; in this case (1)1-ieseli supposes) a large fragment is involved. Again, the ﬁrst, second, and third may be normal, but the fourth and ﬁfth equal: no micromeres, therefore, are formed, and the fragment is probably derived from the animal hemisphere. Or while the first and second are equal, one of the four cells will divide unequally; in the next division a large or small number of micromeres is formed, according, presumably, as a larger or smaller portion of the mieromere region has been included in the fragment (Fig. 106i.

be a. meridional half

divides equally twice, and then two of the four equally (mesomeres), two unequally (macro- and micromeres) (Fig. 107), while a supposed vegetative half is segmented to form four large and

four small cells.

(

Thus the type of segmentation is determined by the initial IV. I INITIAL S'l‘RUC'l‘URl*] OF THE GER,1\[ 197

structure, including the physical constitution, of the ovum alone, a conclusion in which Boveri concurs.

All these fragments will give rise to normal larvae, provided they are not too small. The least egg-fragment that will gastritlate has the same germinal value as the least blastomcre, namely,

(1

FIG. 106.-~ Segmentation of an egg-lragxiient of E¢-Iu'nu.s- supposed to contain the whole of the mieromere area of the egg, and some of the animal hemisphere. u. Eight cells, three equal pairs and one unequal pair; 1;. sixteen cells, four of the eight having formed mieromercs, the other four divided equally. (After Driesch, 1896.)

31-2- (calculated from the volume of the smallest gastrnlae found). Morgan, however, estimates the smallest egg-fiaginent capable of giving rise to a normal larva at from 315 to -616, the least blastenicre that will gastrnlate only

G

FIG. 107. — Segmentation of an e;,rg-fmgnient (Ez-Irinats) supposed to be an exact lneritlional half". (I. Eight cells, four l)e1ng im-s0Incre.~', two n'1-.Lcro1neres and two microineres; b. sixteen cells. (After Dneseh, 1896J

In partial blastulae, gastrulae, or larvae the number of cells is proportional to the germinal value ; and this is true not merely of the whole embryo or larva, but of each of its organs, the 198 INTERNAL FACTORS IV. 1

eetoderm, the mescnchyme, the archenteron. This assertion Of Driesch’s is corroborated by Morgan for the whole larvae, but not for the organs ; according to him the number ofeells invaginated to form the archenteron in a partial tends to approach that found in

a whole larva, about fifty. l)riesch has also determined the relation to their germinal value of the dimensions of partial blastulae, gastrulae, and Plutei and their organs (Figs. 108 FIG. 108.— Outlines of -}, 22, 1, 112). As the table (Table XX)

« l bl‘th fS7 ""7' . . . ., '(‘X;.teilf Dr?:‘S;‘11"°19"00_)” W‘ “ ‘””'S shows, the suifaees ale vex)

nearly as the germinal values, the ratios of the radii of the partial a11d total larvae consequently greater, of the volumes less than these values.

Flu. 1O9.——Ea1'ly gastrulae of Ec-hinu.~c: a. }, b. g, c. 1, (Z. 3,. (After Driesch, 1900.) IV. 1 INITIAL STRUCTURE OF THE GERM 199

TABLE XX

Relation of dimensions of partial blastulae to their germinal value. Germinal values.

1 1

1 . 1. 2+ Rzulii . . . 2 1-2 I .75 Surfaces . 4 2-2-5 1 -5625 Volumes . . 8 3375 1 -4

FIG. 1lU.—l’lutei of I9'rlu'nM.s-: u. b. .1_,, 4'. 1. The skeleton is shown. (After Drie.~xch, 1900.)

From the proportionality of the number of cells to the germinal value it follows that the cells of the whole and of the partial blastulae are of the same size.

If 1:’, S, and V are the radius, surl"aee, and volume of the whole

. . . 1 '

egg, and rm 3", nu the correspon(l1ng- magnitudes Ill a ; larva, S 1:? I’

then 8n = ——,whenee 1'” = _, and therefore 0” = - ~—_.—. By " 1; ~/ 713

this formula. the germinal value of a larva of unknown origin may be calculated. Finally, although segmentation takes place at the normal rate 200 INTERNAL FACTORS IV. I

in the isolated blastomeres, their subsequent development is progressively retarded as the germinal value decreases (Table XXI).

TABLE XXI

Showing the rate of development in partial larvae. Germinal value.

5 Day ! } : 7’; 7‘: L 5 A 1 ' Gastrula IEarly Gastrulal Still earlier Blastula f Gastrula ' 2 Pluteus Early Pluteus Gastrula with Blastula and 4 Skeleton Gastrula 3 Pluteus Pluteus Early Pluteus Gastrula 4 Pluteus Pluteus Pluteus Gastrnln

a C

FIG. 111,-SpIum'¢=chi/Hts Plutci, showing the gut. u -}, b. .1_., c. }. (After Driesch, 1900.) It is evident from these experiments that, in the Echinoderms, while the isolated parts of the egg‘, whether blastomeres or egg fragments, are only able to segment as they would have done had they remained in connexion with the whole, they are neverIV. I INITIAL STRUCTURE 014‘ THE GERM 201

theless capable of total development in a very marked degree. This capacity is not, however, unlimited, but diminishes as development proceeds, and it appears highly probable, and is indeed often admitted by Driesch, that the chief cause of this restriction is the lack of some specific substance and not the deﬁciency of mere material, though this may be a subsidiary factor. The progressive loss of potentialities is exhibited also by the embryonic organs as they are formed later on.

(L

FIG. 112. --Echinu.s~, the tripartite gut of the complete l'luleus: (1. }, b. c. in (After Drieseh, 1900.)

Before concluding this section we must notice an experiment which is the converse of those considered above, namely, the production of one embryo from two eggs.

An unsuccessful attempt was made by Morgan to obtain this result. This author only succeeded in showing that the eggs of AS’/1//ac/‘cc/rim/.9, when (leprived of their membranes by being shaken, tend to fuse together, and that from such pairs double monsters may be produced with two guts and two skeletons. One skeleton may, however, very large] y predominate over the other, which indeed remains rudimentary, and the guts may coalesce. 202 INTERNAL FACTORS IV. 1

Driesch, however, who has employed the same method as Morgan and has corroborated his results, has been able to produce, from the fusion of two ova, a larva single in all its parts—mesenehyme, skeleton, and gut. The organs are in correct proportion but larger than the normal (Fig. 113). The number of mesenehyme cells is about twice the normal.

From this experiment it may of course be concluded that the egg substance possesses a degree of isotropy, but not that that isotropy is absolute. Nothing, it should be remarked, is known of the way in \vhich the eggs which fuse to form one embryo are oriented upon one another. Garbowski, however, has stated

A ﬂ

Fla. 1113. A. Norlnal gastrnla of S_pIz(m'r-('h1'nMs._ B. Single gastrula formed by the fusion of two blastulae. (After Drieseh, from Korsehelt and Heider.)

that fragments of different eggs of ]3'c//iizzts (in segmentation stages) may be grafted on one another, and that the product of their union will give rise to’ a normal embryo, whatever the relative positions of the fragments.

The fusion of blastulae to form giant Planulae has been noticed by Metsehnikoff in the medusa, Jli/roco///a mmae, and in 0])//r_//at/'oc/Ia Korsehelt has observed a similar fusion of (llstinet eggs in the body cavity of the parent when the latter is hermaphrodite. Lastly, Sala and Zur Strassen have observed the fusion in twos, threes, or more of the eggs of ./1.vc¢m'.s-, a result experimentally produced by the first author by exposure to a low temperature, while Zur Strassen has found that such double eggs, provided they had united before fertilization and then been fertilized by asingle sperm, will produce perfectly normal embryos. IV. I INI'I‘IAL STRUCTURE OF THE GERM 203

LITERATURE

T. BOVERI. Die l’o1m-itilt von Ovocytc, Ei und Larve des SIron_r/,z/Iocentrofus Iiritlus, Zool. Jahrb. (Anat), xiv, 1901.

T. BOVERI. Ueber die PoIa.ritii.t des Seeigelcies, Vwh. d. ph_:/s.-nwd. Gas. cu Wiir:Inu'_q, xxxiv, 1902.

H. DRIESCH. Entwicklungsmechzmische Studien : I, II, Zcilschr. uv's.~:. Zool. Iiii, 1892.

II. DRIESCH. Entwicklungsn1ec1mniscl1e Studieu: II|—VI, Zei!s¢'7u‘., 1z'i.s-s. Zoo]. Iv, 1893.

H. Dlulascll. Entwicklungsmechanische Studien: VII-X, Mir/. Zaol. Slut. Ncapel, xi, 1895.

H. Dnusscn. Zur Analysis der Potenzen embryona.lcr Orgamzcllen, Arch. Eul. Merl). ii, 1896.

H. DRIESCII. Betmchtuxigen ﬁber die 01'gunisution des Eies und ihrc Genese, An-h. ]v.'m. Mo:-7:. iv, 1897.

H. DRIESCH. Primiire und sckundi'u'e Regulationen in den‘ Entwicklung dcr Echinodermen, Arrh. Eur. Mech. iv, 1897.

H. DRIESCH. Die Versclnnelzung der lndivid112L]it€‘Lt bei Ec11inidenkeimen. Arch. EM. Merh. x, 1900.

H. DRIESCII. Die isolirten Blastolneren des Echinidenkcimes, Arch. EM. Mech. x, 1900.

II. DRIESCH. Neue Ergihizungen zur Entwicklungs]physiologic dos Eel)inidenkeimes, Arch. Em. Illech. xiv. 1902.

H. DRIESCII. DreiApho1'ismen 7.111‘ Entwicklungsphysiologie jiingstcr Stndiell, Arch. 12'/1!. Jllcch. xvii, 1904.

II. DRIESCH. Altes und Neues zur Entwicklnngsplnysiologic dcs

jungun Astcridenkeiines, Ar:-h. Enl. Mc-ch. xx, 1906.

M. '1‘. GARBOWSKI. Ueber B1-ustomerentru.nspIa.ntationen bei Sceigeln, Bull. Iufmwal. do l'Arml. ties Sci. dc (,'r(u'orie, 1904.

M. T. (irARB()\‘\'Sl\' I. Ueber die l’oIzu'itiit des Scoigcleies, Hull. Internal. ((0 IA('mI. (105 Sci. dc ('rm'uvi¢', 1905.

M. '1‘. GARBOWSKI. Ueber Bla.sto1nerent1'ansplamtationen bci Sceigeln, Bull. Internal. Ac. Sv. Crucoric, 1905.

1*}. KORSCHELT. Uebcr Kerntllcilung, I‘}ircifung und Bci'1'ucI1i.ung bci 01)/H'_IjU(I'0(']I(I1)Il(’I'i/(ls, Zeitschr. 'mf.s's. Zoo]. lx, 1895.‘

J. LOEB. Ueber cine eiufaclne Methode, zwei odci-1110111‘ zusu.mn1engewaclisene Fmbryoncn nus oinem Ei hervorzubringen, P_/lii_qer’s Arch. Iv, 1894.

J. LOEB. Beitriige zur Entwicklungsmechanik der nus einem Ei entstchenden Doppelbildungen, Arch. Euf. Mech. i, 1895.

E. ME’l‘SCIINIKOFl~‘. Embryologisclie Studien an Medusen, Wien, 1886.

T. H. MORGAN. EXp€l‘I1HC1IId.I. studies on Echinodcrm eggs. Anat. Ans. ix, 1894.

T. H. MOREAN. Studies of the ‘ pa.rbial ' larvae of Spham'ech1'Ims, Arch. Ent. Mech. ii, 1896. 204 INTERNAL FACTORS IV. I

T. H. MORGAN. A study of a. variation in cleavage, Arch. Em. Mach.

ii, 1896. '1‘. H. M()RGAN. The formation of one embryo from two blastulae,

Arch. Euf. Mcch. ii, 1896. L. SALA. Experimentello Untersuchungen iiber die Reifung und Befruchtung der Eier bei Asc(u'i.~: mcgalocqrhalu, Arch. mi/cr. Anal.

xliv, 1895. 0. L. ZUR STRASSEN. Ueber die Riesenbildung bei Ascm-is-Eiern,

Jrch. Ent. Mech. vii, 1898. H. l. ZIEGLER. Ueber l*‘urc-hung unter Pressung, Verh. Anal. (:'escll.,

1894. R. ZOJA. Sullo sviluppo dei blastomeri isolati dalle uova. di alcunc

medusc (e di altri organisini), Arch. Eur. Mach. ii, 1896.

§ 7. Nimnirrinm.

In the behaviour of its isolated blastomeres the Nemertine egg resembles the Echinoderm very closely; the cells segment as parts but ultimately give rise to wholes. The limit of this capacity is, however, sooner reached, for there is a sharper distinction between the potentialities of animal and vegetative blastomeres.

Experiments have been made on (Jcrcb/'a/zzlzw («alone by VVilson, on Ccrcbraiulus nzaI'_I/‘iizaﬁus by Zeleny ; Zeleuy and Yatsu have further investigated the development of fragments removed from various regions of the egg.

The axis of the egg is marked by the excentricity of the nucleus and the point of extrusion of the polar bodies. 1\Iatura.tion precedes fertilization. The first two furrows are meridional and divide the egg into four equal cells- (A, 1!, C’, and 1)). Subsequent cleavage is, however, on the spiral plan seen in Turbollarian, Annelidan, and holoblastic Molluscan eggs.

In the Nemertine the egg-axis becomes the axis of the Pilidium larva, the animal being the aboral, the vegetative the oral, pole.

In both the § and the % blastomeres cleavage is partial and an open blastula is formed, or sometimes a flat plate of cells merely, sometimes a closed sphere. The 1}; later becomes a larva with an apical organ, mescnehyme and archenteron, but no lappets; the % larva has a solid archenteron but neither apical organ.nor lappets. Development is slightly retarded 3 % blastomeres give rise to a IV.1 INITIAL STRUCTURE OF THE GERM 205

larva. with mesenchyme and apical organ, but the archenteron is solid. After the next phase—eight cells—-the upper quartette

FIG. 114.———Ce;-cbrafulus. Larvae from upper quartettes of the 8-cell stage. A. Larva from whole quartette. B, C. The same, but one cell was injured. J), E, F. Slightly older, f'rom complete quartettc. (After Zeleny, 1904.)

(1 11-1 rl) produces a larva with an apical organ and meseuchyme but devoid of an archenteron ; the larva developed from the lower

quartette (,-l—~—Z)) has an archenteron but no apical organ, while a

/1 FIG. 115.~—('erebralulus. Larvae from lower (vegetative) quartet-tes of 8-cell stage. Note absence of apical organ, presence of large, hollow or solid, archenteron. (After Zeleny, 1904.) meridional half, comprising two macromcres and two micromeres, develops both organs, though it is usually asymmetrical. In the 16-celled stage the isolated apical cells (1 a 1-1 (I 1) will

give rise to a. ciliated blastula with mesenchyme and an apical 206 INTERNAL FACTOllS IV. I

organ, b11t without an archcnteron, while from the remaining twelve is derived a ciliated embryo with a large solid arehenteron, but destitute of an apical organ and lappets.

Wilson asserts that the size of cells in partial is the same as in total larvae ; their number appears to be proportional to their germinal value.

From fragments of blastulae normal, or nearly normal, dwarf Pilidia may arise, but the degree of development again depends on the origin of the fragment. VVhile the lower third becomes

FIG. ll6.——Cerebratulus. Larvae from portions of the 8-cell stage. A, II. Larva from lateral 4-cell group. Note presence of both apical organ and archenteron. C. Similar, but younger. I). Larva from upper quartette plus two cells of lower quartette. Note that there are three apical organs, a large bl-astocoel, a small archenterou, and two inxluggiiiations of eetoclerm at the sides of the archent-eron. (After Zeleny.

04.)

a eiliated embryo without an apical organ, a piece of the upper two-thirds developed (in one case) into a larva with a gut, with two apical organs, but no lappets (Zeleny). Wilson states that in Cerebrazfulus lacteus the archenteron is small in animal fragments, the apieal organ frequently, though not apparently always, absent in pieces of the vegetative hemisphere.

Animal fragments of the blastula have a greater total potentiality, a greater regulative capacity, than the animahblastom eres IV. '1 INITIAL STRUCTURE OF THE GERM 207

of the eight-celled stage. This is attributed by Zeleny to the greater time and opportunity afforded them of regaining the polarity of the whole.

The behaviour of egg—fragments depends not merely on the part of the egg from which they are taken but also on the time at which they are removed. '

Pieces taken from the ovum, no matter in what way, before the disappearance of the germinal vesicle segment as wholes and give rise to perfect Pilidia. Only in a small percentage (15 %) of cases is their development defective, the lappcts being absent, or the gut abnormal, or the apical org-an absent or multiplied (Yatsu). The proportion of perfect Pilidia obtained from fragments removed a little later, at the time of formation of the first polar spindle, is,

V however, less, only 52 0 It is during the conjugation of the H pronuclei that the deﬁnite localization — 4 H of speciﬁc substances seems to begin, V '

for now the fate of the fragment de . . . . 0 pends upon the direction in Wl)l(‘l1 the cut is made. Eggs from which the animal

. . FIG. 117.-— Cerebmtulus. portion——about one-third of the whole Diammn of egg with one

——has been taken away become perfect POW‘ b0d)'. and basal PW - - ' t b .( I: ‘I d.‘ ' Pilidia ; when, however, by removal of t§)neS1(":)(3f-c’],(i:,.ii;(;3;i:,1 Jiffy,

the vegetative third, or by an oblique vertical (V), aiul oblique cut, the fragiiient does not contain (1%4_°)“ts' (After Zelcny’ the whole of the lower two-thirds, the

lappets are defective and the apical organ sometimes absent as well. If, therefore, there is present at this stage a special material for this organ it cannot as yet be located in its ultimate position.

The cleavage of these fragments is always partial (Zeleny). After the first furrow the material for the apical organ has still not reached the animal pole : the animal region may be removed from both blastomcres without inhibiting its formation. bcrfeet Pilidia may also be produced from eggs in the two-celled stage when pa.rt of one blastoinere has been obliquely cut away.

Should, however, the blastomeres be cut apart before their complete

V 208 INTERNAL FACTORS IV. 1

separation, each develops into a Pilidium with one lappet but no apical organ; it seems, therefore, that the material for this structure is at this stage placed in the bridge of plasma connecting the two cells, and is fatally injured by the operation.

Taken together these experiments make it, to say the least, highly probable that there are in the Nemertine egg deﬁnite substances connected necessarily, that is causally, with the formation of certain organs.

These speciﬁc organ-forming substances may be said to be

preformed, but it is equally clear that they are not prelocalized, but only reach their ultimate destination in the course of development.

LI'l‘ERATURl‘}

E. B. WILSON. Experiments on cleavage and localization in the

Nemertine-egg, Arch. Ent. Mech. xvi, 1903. N. YATSU. Experiments on the development of egg-fragments in

Cerebratulus, Biol. Bull. vi, 1904. C. ZELENY. Experiments on the localization of developmental factors

in the Nemertine egg, Joan-n. E.l'p. Zool. i, 1904.

§ 8. CTE.\'OPIIOR.\.

VVith the Ctenophora we come to a group of animals in which the development of a total larva from a single blastomere is no longer possible, though the missing parts may eventually be regenerated.

The egg consists of a central yoll<—mass surrounded by a superﬁcial layer of granular protoplasm. After fertilization this protoplasm passes wholly to one side—the animal hemisphere-—and the egg divides meridionally into equal parts. The protoplasm then spreads itself over the outer surface of each cell, but returns to the animal side prior to the second cleavage, which is again meridional and at right angles to the ﬁrst. Once more the protoplasm is distributed over the whole outer surface of each hlastomere, only to be again concentrated before the third division, an unequal one, but nearly meridional. There are new four large cells lying together in a square and four smaller cells lying above them in two groups of two each, one at each end of-the plane of IV. I INITIAL STRUCTURE OF THE GERM 209

the second furrow. This becomes the transverse, while the first furrow marks the sagittal plane of the future organism. Each of these cells contains both granular protoplasm and yolk, but at the succeeding division nearly all of the former material passes into the eight small cells, which are nipped off and lie in an oval ring on the vegetative side of the egg 1 (Fig. 118). These small cells or micromeres will give rise to the ectoderm, the large cells or macromcres to the endoderm and mesoderm, the mesoderm

\

FIG. 11H. -Normal segmentation ofthe Ulenophorc egg. :4 u, Stomach or sagittal plane ; I» I), funnel or transverse plane. C, E, from mieromere pole; D, F, from the side. (After Ziegler, from Korschelt and I-leider.)

being separated off later on at the vegetative pole, which becomes

the oral pole of the embryo. In normal development, therefore,

the embryonic axes are marked out at quite an early stage. Chun was the first to isolate the % and § blastomeres of

‘ Ziegler's description is that in this (the fourth) division the furrow first appears near the animal pole, but yolk then streams from the larger vegetative into the smaller animal portion, until the latter becomes the macromere, the former the inicromere. Division is then completed and the micronieres are at the vegetative pole. This explains the divergences in the accounts of earlier observers (Agassiz, Kowalewsky, Chun, F01, and Metschnikoff). 210 INTERNAL FACTORS 1v. 1

Euc/(aria by shaking. These gave rise to half larvae, provided with four costae, four meridional canals communicating with two subsagittal and two subtransverse canals in the ordinary way, one subgastric canal, one tentacle, but a whole funnel and a whole stomach, the latter formed by an oblique invagination ; the side turned towards the missing blastomere was covered over by ectoderm.

Later on the missing half was regenerated, first the subgastrie vessel, then the meridional canals, over these the costae, and

ﬁnally a tentacle.

FIG. 119.~Development of isolated bl-astomeres in Ctenophora. a, /2. Segmentation of 1% blastomere of Beﬁio ovatu: a, two large, two small cells; b, each gives off a mieromere; c Larva from a .1, blastomere with four costae; (I, c. Segmentation of 1 blastomere ; f. The resulting larva with two costae; g. Larva. with six costae from ~2~ blastomercs. (After Driesch and Morg.m, 1896.) h. Isolated »1"‘,; blastomeres of liwﬁe omlu;

j. The resulting larva with four costae, one sense--oi'gai1, and one

stomodaeum ; 2'. Isolated -1'3, blastomeres; A-. The resulting larva with three costae, one sense-organ, and one stomodacum. (After Fischcl, 1898.)

Such half larvae are found in the tow-net after a storm and may become (l}olz'7za) sexually mature.

These experiments have been repeated and confirmed on Barbi‘ by Driesch and Morgan. The authors add that the segmentation of these isolated blastomeres is partial. The 71; larva has two costae only and a large and a small canal, the former passing to the costae, the latter to the opposite side and representing apparently IV. 1 INITIAL S'l‘RUC'l‘URE OF THE GERM 211

the other half of the funnel (Fig. 119 mg). Fischel has carried the analysis of the potentialities of the blastomeres a step further (Fig. 119 /z—zt). Though cleavage is partial, as described by Drieseh and Morgan, there are slight irreg~ulari.ties in the position of the blastomeres. A ,1; blastomere produces one eosta only, -1-55 blastomeres (four macro- and four mieromeres) four eostae, {-3(five macro- and five mieromeres) ﬁve, -19%, -1%, and 14;, when separated in the same way, respectively three, six, and two eostae. Similar results were obtained by meridional division of the egg in later stages. Fisehel conﬁrms the statements of the other

_FiG. 120.——u. Beriie oz-am: egg in the 16-cell stage. The mieromeres disarmugecl by pressure ; I». The resulting larva with two sense-organs, and four eostae radiating from each : one stoinodacuni. (After Fischel, 1898'.) 1'. Normal segmentation of the egg of Beriie from \Vl1lCll a small portion of the vegetative hemisphere has been removed; d. Larva produced from the same, with eight eostae, four endoderinal canals, and one (central) stomodaeum. (After Driesch and Morgan, 1896.) investigators as to the structure of the half larvae. In % larvae, however, not three, but four, canals are formed, and the stomodaeum is vertical. The sense-organ is not found in embryos developed from small fragments. Removal of the Inacromeres left the number of eostae unaffected, but displacement of the mieromeres by pressure led to the formation of an abnormal larva with two sense-organs and eight eostae arranged in two groups of four each round each sense-organ (Fig. 120 a, 6), or, if the mieromeres were separated into two lets, a group of four round one organ, a group of three round the other, and some scattered combs. With greater pressure three sense-organs were produced and the

P 2 212 INTERNAL FACTORS IV. 1

costae were very irregular. In these eggs there was, however, only one stomodaeum, and only the normal number of canals.

It appears, therefore, that the individual eostae are deﬁnitely related to the meridional divisions of the segmented egg, and further that the material which is concerned in their formation becomes located in the micromeres of the 16-celled stage.

Driesch and Morgan have also shown that if the vegetative hemisphere be taken away from the undivided egg, the animal portion, though it segments normally, is able to produce neither costae nor stomodaeum; it has, however, cndodermal canals. Loss of a small portion of the vegetative hemisphere, however, does not interfere with subsequent normal development (Fig. 120 0, II).

On the other hand an egg which has been deprived of a lateral portion divides, by three unequal divisions, into eight cells, all of which form mieromeres. Nevertheless, the larva has only four, five, or six costae, and some of these rudimentary, only three or four canals, and the stomodaeum, as in half larvae, is oblique.

As Driesch points out, the whole of the nucleus is here, and the embryo is still defective. Diﬁerentiation thus depends on the cytoplasm, but is quite independent of segmentation, which may be perfectly normal without determining the formation of a normal embryo. l)rieseh further argues that the observed defects are due to lack of suflicient mater-ial.merely, not to lack of any preformed specific substance. \Vhether such substances are already present in the unsegmented as they undoubtedly appear to be in the segmented egg, it is perhaps impossible, on the evidence, to decide, but it may be observed that the development of perfect larvae from those eggs from which a portion of the vegetative hemisphere has been detached does not ncccssaril y support the View advanced by Drieseh. '

LI'J.‘ERA'l‘URE

C. CHUN. Die Dissogonie, eine neue Form der geschlechtliehen Zeugung: 7. Zur Entwicklungsuieclianik der Ctenophoren, I"('.s'l.s'c7u'. L<'m'7.'m'!, 1892.

II. Dmascu and T. H. MORGAN. Zur Analysis der ersten l*lntwicklungsstadien des Ctenophoreneies, Arch. Eur. Illech. ii, 1896.

H. FISCHEL. Experiinentelle Untersuchungen am Ctenophorenci, 1, .‘.lI'fh. Enl. Zlfech. vi, 1898; II, 111, IV, ibid. vii, 1398.

II. E. ZIEGLER. Experimentelle Studien ﬁber die Zelltheilung: Die Furehungszellcn von Bertie ovata, Arch. Ent. Mech. vii, 189:4» IV. I INITIAL STRUCTURE OF THE GERM 213

§ 9. Crmnrorom AND MoLLUscA.

In the Chaetopoda and Mollusca (except the Cephalopods) it is possible to trace back almost every organ of the body to some particular cell, or group of cells, of the segmenting ovum, to describe, in fact, its lineage in terms of individual cells. Speciﬁc materials for the formation of the vaiious parts seem to be sundered from one another by the process of cleavage, which thus presents the characters of a ‘Mosaic’ work in an exceptional degree. 'l‘o what extent such a preformed structure (lees in reality exist experiment alone can decide, and the answer given by experiment, so far, at least, as the nucleus is concerned, is in the negative.

By means of pressure Wilson succeeded in preventing in Nerei.s~ the normal formation of the first quartette of micromeres ; instead, the egg divided into a flat plate of eight equal cells. The pressure was then removed. All eight cells formed micromeres, and later on eight instead of the usual four macromeres were found in the Troehophore la1'va. Four of these necessarily contained nuclei which would normally be placed in cells of the prototroch, and yet the larva was normal. In this case, then, the causes of differentiation cannot lie in the nucleus; they must be sought for, if anywhere, in the cytoplasm.

In the eggs of these forms segmentation is holoblastic and of the spiral type, with the cleavages alternately dexiotropic and laeotropic, and it is easy not only to determine the lineage of each organ, but to compare the oi-igin and the destiny of individual cells in a large series of forms. The first, second, and third quartettes of micromeres are usually destined for ectoderm formation, the second quartette cell in the D quadrant, 2 «I, being the first somatoblast from which the ectodermal structures of the trunk a1'e derived. The eiliated ring or prototroch is formed, in most cases, from certain derivatives of the ﬁrst quartettc, namely 1 a 2———I (l 5.’, the primary trochoblasts, each of which divides into a group of four; but it may be reinforced by secondary trochoblasts from the same or the second quartette. The second and third quartettes may provide larval nzesoderm or mesenchyme. The trunk mesoderm is usually derived from 4 (I. The remaining 214 INTERNAL mcwons IV.1

cells of this quartette (4 a—4 c) become with the residual macromeres (A—D) the endoderm (or mesendoderm).

A peculiar structure seen in some forms (Ilyamwsa, Dentalinm,

.41

FIG. 121.-—No1-nml cleavage of I(_I](m((.98(L A, B. Formation of polarlobe or yolk-lobe (til); (Y. ‘Trefoi1‘ stage; I), E, F. The polar-lobe passes into CD; 0. The second polar-lobe protruded in H; it passes into D. (After Cmmpton, from Korschelt and Heidcr.)

0/laclopteru.v, M_y:o.v/oirm) is the yolk-lobe, or polar lobe, as it should properly be called. This is a. mass of cytoplasm p1-oi-ruded at the IV. I INITIAL STRUCTURE OF THE GERM 215

vegetative pole after fertilization. It becomes attached entirely to one of the ﬁrst two blastomeres, namely to CD, into which it is withdrawn; prior to the next division it is again protruded, attached to D only, and then once more withdrawn : at the time of the next division it is protruded again, for the last time (Figs. 121, 122). '

Crampton has isolated the blastomeres of I/‘//amzwz and watched their development (Figs. 123-6). —;-, §-, % (A, B, and C), 1}, §

FIG. 122:-Normal cleavage of II_:/mmssa. (After Crampton, 1896.) a, 8—celled stage from above; I), 1C-celled stage from above; r, 24-celled

stage, from above; :1, optical section, sliglltly oblique, of a later

embryo, from below. Division of mesoblast pole-cell (rI‘).

(a macromere and a micromere), %, -,1; (a micromere), I53, 11} and 193 all continue to segment as though the missing blastomeres were present, except for certain small irregularities in the positions of the cells ; the A, B, and C blastorneres, separately or together, bud off the usual four generations of mieromeres, but D only the ﬁrst three ; the division of the mic-romeres continues in nearly normal fashion, The 3,. mic-romerye divides twice only. All die, however, 216 INTERNAL FACTORS IV. 1

without giving rise to larvae. A 13- macromere, on the other hand,

or a. -1-13 or later blastomere, will not segment at all. By lowering the temperature the cleavage may be made to

resemble that of whole ova.

FIG. ]23.—-Il_:/anassa: c-lenvzige of smaller blastoniere (AB). a, Undivided; b, embryo, fr_on1 sule ; (.', »j, embryo, from above; «I, -,'-‘:5 emln-yo, gram ahdove _the clot; lindficattesil the for1ne1t'1centre of the egg; 9% .1_,'3‘«e1}J1.bryo;)

rom si e; , grow 1 o ec 0 erm over ie nmcromeres; 3/, orma ion 0 the nncromeres of the foulth generation, from below; It, ciliated partial

embryo of 46 hours. (After Cranipton.)

The yolk-lobe inﬂuences the character of segmentation, for if it be removed CD divides like AB. More important ‘than this is IV. I INITIAL STRUCTURE OF TIIE GERM 217

the relation of the yolk-lobe to the mesoderm. When the former is cut olf the latter is not formed at all. There are four generations of micromeres and four equal macromeres, and the larvae, though eiliated and able to swim, never reach the veli_qer stage (Fig. 127).

1/

1/ I

FIG. 1'24.~Il;/«nus-.5-«I: ¢~1eavan.ge of CD. (4, Formation of yolk-lobe (yl) ; b, ‘trel'oil'; v, embryo from above; (1, early 1‘ e1ubr_yo, from above:

~'econ(lary yolk-lobe (yl) ; a, resting 3, embryo from above; f, ,‘E_. embryo

from above; (1, passage to 11,- stage, from above; h, embryo, from side ; i, division of n1esobla.st pole-cell (d‘), from below. (AftcrUra.1npt0n.)

The potentialities of the blastomeres are thus strictly limited both in segmentation and difl’e1‘entiation. This result is amply confirmed by Wilson's researches on I/eutulimzz and Patel/a.. 218 IN'l‘l*}RN.\L FACTORS IV. 1

In the latter genus the development of the isolated blastomeres, especially of those resulting from later divisions, has been minutely followed. Each separate first quartette cell (1 a—1 (1) forms (Fig. 128) a closed ectoblastic structure with an apical org-an at

FIG. l25.—IIgmm.~sa: cleavage of ;} blzmstomere (Al. (I, 3, froI_n side: b, ,“:_;, from side; 0, passage to 143, ﬁist form, from side; (I, rest1ng_-‘g. second form, from side; 0, pttssngo to fl, from side ; f, ;_,7,,«, from s1( e. (After Crauupton.)

“ii,” ‘I

Fm. 126.—I/_:/auassa: aw, cleavage of Q; blnstomercs; (1, ,9, stage, from one 1n1c_romere; 12, ~,*;; stage, from two micromeres; r, —,“,, stage, from three nucromeres. d—-f, cleavage of bliLSt01l1(‘1‘CS; cl, -J,-3 einbryo, from below: P, origin of fourth qunrtefte. from below; f, embryo of 48 hours. (After Urzuupton ) IV. I INITIAL STRUCTURE OF THE GERM 219

/‘B co

FIG. 12"’7.—~]I_1/a12assa: cleavage of the egg after removal of the yolklobe. a, :5; b, 1.-;_ r, first quartette formed; (1, second quxutette formed, ﬁrst qu2Lrtgtt§fl1v1de(l ; a—¢l, from above; e. from below: fourth quzu-tette formed; _/, cxllutod embryo, from side. (After Cr-.unpton.)

(I // 0

FIG. 128.-- Patcllu: development of isolated ;« micromeres. ((, _?; micromere; I), c, first division; «I, 3"-‘,3 stage, with two trochoblasts (stippled), one rosette-cell, and one p1-imary cross-cell; e, larva}. of 24 hours, from the side, showing trocholnlasls below, apical cells above. (Aft:-1' \Vi1Su11.) I 220 INTERNAL FACTORS IV. I

one end, and four cells (derived from a primary trochoblast) provided with long cilia. at the other. When the primary trochoblasts (1 a 2-1 (Z 2) are isolated each divides into four ciliated cells, the cilia arranged as normally in a single row, but no further (Fig. 129 a—e). If the daughter-cells of these troehoblasts (1 a 2. 1 -1 (Z 2. 1 and 1 a 2. 2--1 (Z 2. 2) are separated they divide once only (Fig. 129/’), while their daughter-cells do not divide at all (Fig. 129 g, /1). The sister-cells to these (1 «L 1—1 rl 1) behave in the same way, forming a closed larva with an apical organ at

'2

(*7 3

0 gr

5; I; 0 ll

FIG. 129. —-Pulellu. Development of isolated primary and secondary troehoblasts. a, Primary troehoblast; b, 0, first division; «I, second division ; 0, after 24 hours ; f; pair of primary t1'ocl1oblasts, the products of division of either 1'“ or 12"’, after isolation; ,1/, h, single prilnary trochoblasts, either 12”, 1*", 1”‘, or 1”"; i, a pair of secondary t1'oehoblasts, the products of 1"; j, Ir, single secondary trochoblasts. (After Wilson.)

one end and secondary trochoblasts at the other, while the isolated apical cells and secondary trochoblasts divide as often as they would in the whole embryo and put out their cilia (Fig. 129 i-,{', Fig. 130 u—e). Finally, the whole first quartette becomes an ectoblastie larva with an apical organ at one end and a prototroch at the other, but with no archenteron (Fig. 130/; 9). The cells of the second quartette (2 a——2 (I) likewise form each a hollow ectoblastic

vesicle containing larval mesenehymc, ciliated secondary trochoblasts, and a few (prcaual) ciliated cells (Fig. 131). IV. I INITIAL STRUCTURE OF THE GERM 221

The cells of the third and fourth quartettes do not develop at all.

A -E1, maeromere (1 A-1 D), however, segments partially, to form a micromere of the second quartette, from which secondary

e f 9

Fm. 130.-—I’uIella. Development of isolated cells of ﬁrst qualrtet-to and of the whole ql1iL1'i:0tl’;C. (1, lsolzited 1‘; I», its first division; ¢', its second division ; (I, after 12 hours ; 0, after 22 hours, showing apical cells zmd secoinl-My troelioblasts; _f, first division of isolated first quumtette; g, second division. (After Wilson.)

(I

FIG. 131. — l’uleIlu. Isolated cells of second qu-.u'tette. um, Cl0‘¢l.V1LgP,; «I. after 24 hours, showing sec.-ond:u'y trooheblusts and ‘.’ prezuml cells. (After Wilson.) trochoblasts are later developed, and one of the third. Subsequently an are-henteron is invaginated (Fig. 132). So the 116- macromere (2 A-—-2 D) forms a micromere (3 a-~31/) and gives rise to a. larva, with lentomesoblast (D) or entohlast only (A— C). 222 INTERNAL FACTORS IV. I

Isolated 71; (A—D) or is (AB or CD) larvae have an apical organ and a prototroeh complete or semicircular; they may remain open or close up ,- sometimes they gastrulate.

It is evident that each cell has exactly the same potentialities for division and diﬁerentiatiou when isolated as when in connexion with its fellows. Only in the shifting of the cells and closing up of the whole cell-mass is there anything which approaches to total development.

$3‘

.;

5

8 u

FIG. 1I)‘2.-—1’utella. Development of isolated basal (inacromere). a, First; 11, second division; 1-, d, -56- and 64-cell stage. In 4; the position of the 2-group is normal, not in (I ; 0, after 24 hours, showing two secondary troehoblasts ()roducts of 2“) and two feebly ciliated cells (preanalcel1s‘.’). (After ilson.)

The dependence of this speciﬁcation of the blastomercs upon deﬁnite substances preformed——tl1ough not prclocalized—in the unsegmented ovum is demonstrated by Wilson’s experiments on Deizlztliuril. The newly laid egg of Ijeuializmz contains a brickred yolk—mass surrounded by a thin hyaline eetoplasm ,- the latter is thickened a little to form an animal polar area and very considerably to form another large area. at the vegetative pole. The nucleus is in the centre (Fig. 133). VVhen the sperm enters the

nucleus breaks down, and its substance becomes conﬂuent with both IV. I INITIAL STRUCTURE OF THE GERM 223

areas. This continuity is soon lost and the egg becomes divided into three regions, a clear layer spread over the animal hemisphere, below this the yolk-mass with the fertilization spindle, and a. large,

_I/ /I 1

FIG. 133.-V 1)mlulimu. Normal cle-.wn;,re. The red pigmented part is represented by stippling. a, Freshly laid egg, polar view ; b, the szunc 20 minutes latter, after shedding the menibrame; c, the smile from the side; (I, one hour after fertilization, showing polar bodies; 0, first division and protrusion of first polar lobe; f. ‘trefoil’ stage; (1, the polar lobe has passed into CD; h, p1‘0t1‘u~siOn of second. polztr lobe (Z2) ; 2', second cleavage. (After Wilson.)

nearly yolk-free area at the vegetative pole, which is 11ow protruded as the polar lobe. This becomes attached, as described above for Ilqazzztssra, to the D blastomere. The first quartette of 224 INTERNAL FACTORS IV. I

micromeres is formed from the clear animal area; after this division the polar lobe of D moves up to meet the upper white area, and so it comes about that a large part of this lobe passes into the second micromere, 211, which is the ﬁrst somatoblast. In the other quadrants the second mieromeres are formed from the upper white area alone. All the cells of the third quartette and (it is not known certainly whether this is the mesoblast) 4 (I in the fourth are quite white. The larva is spindle-shaped with

FIG. 134.—l)enIalium. Cleavage after removal of the first polar lobe ; a, first division: the isolated polar lobe is seen below; b, 4—cell stage, from animal pole; 1', 8-cell stage, from animal pole; d, forum,tion of second quartette, from vegetative pole; 0, same stage as last, open type; f, same stage, from animal pole. (After Wilson.)

a broad, triple prototroch. Tlierc is an apical organ of long cilia and a tuft of sensory hairs at the opposite end (Fig. 135 (6).

When the polar lobe is removed in the ‘ trefoil ’ (2-celled) stage, segmentation continues, but no polar lobe is formed in any stage, the D cells are not larger than those in the other quadrants, the embryo has no lower white area, the segmentation cavity may be open below, and the larvae are deﬁcient and die (Figs. 134-, 135 6). Post-troclial region and apical organ are both absent and IV. 1 INITIAL STRUCTURE OF THE GERM 225

these missing parts are never regenerated ; the pretrochal region is, however, increased and the three rows of cilia in the prototroch are usually developed.

The polar lobe is thus clearly connected with the formation of the trunk and the preoral sense-organ.

The segmentation of theiisolated blastomeres is partial, as in the types already examined, but their subsequent behaviour depends on the presence or absence of the polar lobe. AB larvae or A, B, or C larvae closely resemble the deﬁcient trochophore just described ; CD or D larvae are normal, though the structures derived from the polar lobe are out of proportion to the rest

FIG. 135.——Dentalium. a, Normal trochophore of 24 hours; b, larva of 24 hours after removal of the ﬁrst polar lobc. (After Wilson. 1904-)

(Fig. 136 a-d, 1}). The failure of the egg to develop normally is not due to insufficiency of material, for the volumes of CD and D are less than that of a whole but lobeless egg.

The micromeres 1 a, 16, 1 c, when isolated, become actively swimming, ectoblastic embryos provided with a prototroch, but with no apical organ, gut, or post-trochal region; the other micromere, lzl, however, has an apical organ, though it is Still Without the hinder region and incapable of gastrulation (Fig. 136 e,f It appears, therefore, that the material for the apical organ is now placed in this cell, and this is proved by experiment ;

for if the polar lobe is cut away during the second cleavage, or

‘I Jzxxuson Q, 226 INTERNAL morons IV. 1

from the isolated CD blastomere, then the larva has an apical organ, but no post-trochal region. Hence between the first and second divisions the stuff which determines the formation of this organ must migrate out of the polar lobe into the animal hemisphere of the D cell.

In the unsegmented egg the speciﬁc material for both apical organ and post-trochal region is situated in the vegetative

FIG. 136.—DentaIimn. Larvae from isolated blastomeres. «, b, Twin larvae from the isolated CD (a) and AB (7)) halves of the same egg, 24 hours; 0, d, twin larvae from the isolated D (c) and C (d) quadrants of the same egg, 24 hours; :2, f, twin larvae from '.the isolated posterior micromere 1 d (e) and the 1 c micromere (f) of the same egg, 24 hours; g, }larva from one of the A, B, or C quadrants, 72 hours. (After Wilson.)

hemisphere. The animal half of the egg, whether removed after fertilization, or before and then fertilized, segments like an egg from which the polar lobe has been removed (Fig. 137 6), and develops neither apical organ 11or post-trochal region (rarely an apical organ if the fragment is large). The vegetative hemisphere, when out off before the entry ofithe sperm and afterwards IV. 1 INITIAL STRUCTURE OF THE GERM 227

fertilized, segments as a whole with the polar lobe in correct proportion and gives rise to a dwarf larva with all its parts complete (Fig. 137 a, c). Many, however, die.

A meridional half-egg is able to develop normally, or nearly so.

The behaviour of the enucleate vegetative fragment of the fertilized egg is interesting. Though deprived of nucleus and centrosome it forms its polar lobe synchronously with the divisions of the animal portion ; three times is the lobe protruded and three times withdrawn: the fourth time it is not taken back; this is the moment when the first somatoblast is given off. In this enucleatc fragment the polar lobe is not in proportion but of the same size as in the whole egg.

Even the isolated polar lobe, or pieces of it, exhibits periodic movements, amoeboid ,or of elongation, or protruding a smaller lobe.

(L

FIG. 137.—Dentalium. Development of egg-fragments, the plane of section being indicated in the small ﬁgures. a, b, Twins, after horizontal or oblique section near animal pole; c, trochophore developed from a. fragment resembling that shown in a. (After Wilson.)

The existence in the egg of preformed substances speciﬁed for the production of particular organs is thus incontrovertibly established, and as long as an egg-fragment or blastomere contains all these it will give rise to an embryo which is, in form, perfect, though lack of enough material (or the shock of the experiment) may bring its life to an untimely end.

At the same time these substances, though preformed, are not necessarily preloealized; their original is not the same as their ultimate situation, and in their redistribution we have to deal with a process which is, as Wilson remarks, truly epigenetic.

Before bringing this section to a conclusion reference must be made to another case in which it seems probable that deﬁnite

Q2 228 INTERNAL FACTORS IV. 1

organ-forming substances exist, though experimental proof is at present lacking.

In the immature egg of the aberrant parasitic Polychaet .413/zoatoma there is, according to Driesch, at the vegetative pole a mass of greenish cytoplasm, the surface of which is convex towards the reddish brown mass of the rest of_ the egg. When the polar bodies are formed the red substance retreats to the animal pole, leaving a clear equatorial zone.

Before segmentation a polar lobe is formed——as in Ilyauassa

FIG. 138.——Deve1opment of the egg of Jllyzosioma. The green substance on the vegetative side is shaded ﬁnely ; this passes into the polar lobe (e), and into the D quadrant (1'). The red substance of the animal hemisphere is coarsely dotted. (After Driesch.)

Fm. 139.—Development of the egg of My/zostoma. Formation of the cells 1 d (d‘ in c) and 2 d or X (d’ in e). The latter cell takes a portion of the substance of the polar lobe. (After Driesch.)

and Deutalz'um.—containing the green and part of the clear substance: it passes first to CD and then to D. When the latter divides it is again protruded and withdrawn. The cell 1 (3, like 1 a, 1 6, 1 c, contains the reddish protoplasm. The second micromere in the D quadrant—2 (Z——-takes part of the green substance, and Driesch suggests that the rest of the latter would eventually pass into the second somatoblast (4 ti = M) (Figs. ],38, 139). IV. I INITIAL STRUCTURE OF THE GERM 229

LITERATURE

H. E. CRAMPTON. Experimental studies on Gasteropod development, Arch. Eut. Mech. iii, 1896.

H. DRIESCH. Betrachtungen ﬁber die Organisation dcs Eies und ihre Genese, Arch. Ent. Mech. iv, 1897.

E. B. WILSON. Cleavage and mosaic work. Appendix to Crampton’s paper on Ilyanassa, Arch. Eut. Mech. iii, 1896.

E. B. WILSON. Experimental studies on germinal localization. I. The germ regions in the egg of Dental imn, Jomw. Ezrp. Zool. i, 1904.

E. B. WILSON. Experimental studies on germinal localization. II. Experiments on the cleavage mosaic in Palcllu and Dcnlulium, Journ. Exp. Zool. i, 1904.

§ 10. ASOIDIA.

As has already been shown, there is in the Vertebrata and zlmp/ziowue no constant relation between the first furrow and the sagittal plane of the embryo, although the symmetry of the embryo does stand in a very deﬁnite relation to that of the unsegmented ovum. Further, experiment has proved that the isolated blastomeres, up to a certain point at any rate, a.re capable of giving rise to normal total larvae. It is, perhaps, all the more remarkable that in another group of the Chordata, the Ascidians, the symmetry of the embryo is not only predetermined in the symmetry of the egg but also by the planes of segmentation, and that the potentialities of isolated blastomeres remain strictly what they would have been in the whole ovum.

The earliest experiments are due to Chabry (Figs. 140, 141). According to this author the first furrow in the egg of /1.9ci(h'ella asper.9a is meridional and coincides with the sagittal plane; the second, likewise meridional, coincides with the transverse plane; the third, equatorial, separates oral from aboral regions, and is therefore horizontal.

Isolated g blastomeres or 7"} right or left blastomeres give rise to imperfect larvae with only one ﬁxing papilla, only one atrial invagination and one pigment-spot, the eye; the last is absent in the 3,: left larva. % anterior larvae have one ﬁxing papilla, one sense-spot, a,gut, a cerebral vesicle and a posterior notochord ; 230 INTERNAL FACTORS IV. 1

in %} posterior larvae the ehorda and brain are absent, though the three germ-layers are all present.

In the segmentation of the isolated blastomeres the direction of division is altered, in that a furrow (the fourth) which in the whole egg is meridional and at an angle of 45° to the first two becomes parallel to the plane of separation, and so resembles the third furrow of the whole egg.

A right or left % blastomere or %, anterior or posterior, right

FIG. 140. -—A. Egg of Ascidiella aspersa in the two-cell stage. _One blastomere has been killed (G). B. The survivor (D) divides to form, in (J, a small normal gastrula. (After Chahry, from Koischelt and Heuler.)

0

FIG. 14l.—Larva. of Ascidiella aspersa produced from 1 blastomeres. F, sucker; At, atrium; No, notochord; En, endoderm. (After Chabry, from Korschelt and Heider.)

or left, blastomeres consist after this division of two tiers of four cells each ; the cleavage of these cells resembles, therefore, that of a whole egg.

It should be noticed that the blastomeres cannot be actually separated; it is only possible to kill one, by a needle, and note the development of the survivor.

The more recent researches of Conklin on Cyu//zz'a have conﬁrmed and extended Chabry’s results.

The immature egg of Qynl/ziu, (Fig. 142) comprises a central grey yolk surrounded by a peripheral layer of yellow pigmented IV. 1 INITIAL STRUCTURE OF THE GERM 231

FIG. 142.——Norma1 development of the egg of C'_1/nthia pm-tim. Maturation and fertilization. (After Conklin.)

A. Unfertilized egg before the fading of the germinal vesicle (clear), showing central mass of grey yolk (lightly dotted), peripheral layer of yellow protoplasm (thickly dotted), test-cells and chorion.

B. After the entrance of the sperniatozoon the yellow protoplasm has streamed to the vegetative pole (v); in it, excentrically, is the sperm nucleus (spat). The clear protoplasm derived from the germinal vesicle partly forms a. layer over the yellow protoplasm, in art remains at the animal pole (a). The grey yolk occupies the anima part of the egg.

C. The yellow and clear protoplasmic substances have both streamed to what will be the posterior side (P). A, anterior, V, ventral (animal pole), D, dorsal (vegetative pole).

D. View of the same egg from behind. The two pronuclei are seen side by side in the clear area. B, right; L, left. 232 INTERNAL FACTORS IV. I

protoplasm ; near one pole (the animal), and so determining the egg-axis, is the large germinal vesicle. Prior to the extrusion of the polar bodies the latter breaks down and provides a. third substance, a clear cytoplasm. On the entrance of the spermatozoon near the vegetative pole a remarkable change takes place in the arrangement of these materials. The yellow protoplasm flows down to the vegetative pole and arranges itself symmetrically to the axis; above it is a zone of clear protoplasm derived from the germinal vesicle; these two areas together occupy about onethird of the egg. The remaining, the animal two-thirds, are occupied by the grey yolk with only a small quantity of clear cytoplasm at the animal pole in which, after the formation of the polar bodies, the female pronucleus is embedded.

The egg is still radially symmetrical, but by further shiftings of these substances it becomes bilateral.

The spermatozoon moves in the clear protoplasm to the equator on one side, the posterior side as it will be, and the yellow protoplasm follows suit; the first is now disposed in a broad equatorial band encircling half the circumference of the egg, the second forms a wide crescentic mass below it and reaches nearly to the vegetative pole. The female pronucleus, with its clear area, now joins the male.

The egg is now bilaterally symmetrical about a plane which includes the axis, and passes through the middle of the clear and yellow substances and between the two closely apposed pronuclei.

It seems doubtful whether the movement of the clear and yellow material to the posterior side is immediately due to the agency of the sperm, for the latter does not always take the shortest route to the equator; it may enter on what becomes ultimately the anterior side ; and further, when two sperms enter ﬂley both travel to the same spot, appearing to be passively carried along by the independent streaming of the egg cytoplasm.

The two pronuclei next move into the axis of the egg in the animal hemisphere, together with their clear protoplasm. The grey yolk is thus largely displaced and shifted into the vegetative region. The egg, therefore, now consists of. an animal IV. 1 INITIAL STRUCTURE OF THE GERM 233

hemisphere composed almost entirely of the clear substance, and a vegetative hemisphere, the anterior portion of which includes grey yolk and nothing else, the posterior portion the yellow protoplasm together with a little yolk near the vegetative pole. The cleavage furrows separate these substances from one another in a perfectly definite way (Fig. 143). The ﬁrst is

A '5' 1"”

FIG. l43.—Cleavage of the egg of Cynthia pm-tita. (After Conklin.)

A. 8 cells, from the left side.

3. 8 cells, from above. The yellow crescent (thickly dotted) is limited to the posterior dorsal cells: the animal cells are clear, the grey yolk (sparsely dotted) is found in the dorsal cells, both anterior and posterior.

C. 16 cells, from the dorsal side.

D. 74 cells, dorsal view, showing the division of the 4 neural platecells (n. ), and 4 chords cells (ch.). There are 10 endoderm cells (e), 6 muscle-cells (m.s.), 8 posterior (p.m.¢-h.) and 2 anterior (u.m.ch.) mesenchyme cells. 234 INTERNAL FACTORS IV. I

g /4

' Fm. 144.— nthia parh'tq. Succe 've stages in the development of the same righ alf embryo, the left stomere having been iniured in IV. 1 INITIAL STRUCTURE OF THE GERM 285

meridional and sagittal, the second meridional and transverse, the third equatorial and horizontal. Since the animal pole will be ventral, the vegetative dorsal, the right and left sides of the future embryo are simultaneously determined.

The four ventral animal cells consist almost entirely of clear protoplasm, the two anterior dorsal entirely of grey yolk, the two posterior dorsal almost exclusively of the yellow substance.

The next furrows are, roughly, meridional and at angles of 45° to the ﬁrst two; they emphasize the bilateral symmetry, for in the anterior half they meet the first furrow above, the second below, and conversely in the posterior half.

The direction of the divisions in the next phase may be most easily gathered from the ﬁgures. In the vegetative hemisphere four anterior ehorda-neural cells are separated oﬁ ; the six posterior cells include all the yellow substance and will give rise to the posterior mcsenchyme and the muscles of the tail; six endoderm yolk-cells remain, four anterior, two posterior. The next division sees the separation of the ehorda from the neural cells in front, of an outside ring of muscle-cells from an inside ring of mcsenchyme cells behind, and of an anterior mcsenchyme cell on each side from the anterior endoderm.

A few anterior animal cells become associated with the neural cells in the formation of the nervous system; the remainder of this hemisphere is ectodermal.

In gastrulation the endoderm cells first sink inside, followed by the ehorda in front and the anterior and posterior mesenchyme cells and the muscle-cells at the sides and behind. Overgrowth is greatest at the anterior dorsal lip, the blastopore thus becoming posterior.

In normal development, therefore, the axes of the embryo are

the 2-cell stage-. The yellow protoplasm is indicated by coarse stippling. (After Conklin.)

a, 8-cell stage, posterior view. V, ventral; D, dorsal. b, 16-cell stage, anterior view. ' Normally, A 5.1 and a 5.3 lie in front of A 5.2 and a 5.4. c, 16-cell stage, posterior view. In normal eggs the cells B 5.2 and b 5.4 lie behind B 5.1 and b 5.3. d, 30-cell stage, dorsal view. e, j; 34-cell stage; e, posterior, f, dorsal view. In normal e gs the cell B 6.4 lies laterally to B 6.3, and the cell B 6.1 lies between 6.3 and A 6.1. g, 46 cells: posterior view. h, 48 cells: dorsal view. The cells B 7.1 and B 7.2 should lie next the middle line. 236 INTERNAL FACTORS IV. I

predetermined by thearrangement of certain substances in the egg, and segmentation exhibits the character of a ‘ Mosaic ’ work, segregating these specific materials into the rudiments of deﬁnite organs. The causal connexion between these materials and the organs to which they are beforehand assigned is demonstrated by experiment.

FIG. 145.— Cynthia purtita. Half and three-quarter embryos. (After Conklin.) a, right half gastrula, dorsal view. The neural plate, chorda, and mcsoderm cells are present only on the right side and in their normal osition and numbers. b, left half of young tadpole, dorsal view. he notochord is normal except for size and number of cells: the muscle and mesenchyme cells are present only on one side: the neural plate (71.1). is abnormal in form but not in position. c, right half of young tadpo e, dorsal view: slightly younger than b. m’ch., mesenchyme, ma, muscle-cells. d, left anterior three-quarter embryo, dorsal view. The anterior half is entirely normal with anterior mesen chyme (m'ch.) on both sides. Posterior mcsenchyrne and muscle-cells upon the left side only. IV. I ‘INITIAL STRUCTURE OF THE GERM 237

‘The segmentation of a 4} or of 7*} right or left blastomeres is partial except for the alteration in the direction of the furrows of the fourth phase already described by Chabry. The blastomeres divide as though the other half of the egg had not been killed (Figs. 144,146, 147). Eventually a half gastrula is formed with exactly half the normal number of neural,chorda,endoderm, mesenchyme and muscle-cells ; the gastrula cavity is open on the side of the dead blastomere. Many are, however, abnormal in that the endoderm and muscle- and mesenchyme cells do not invaginate (exogastrulae) or show invaginations of the ectoderm (pseudogastrulae).

The half gastrula may develop into a half larva, with the notochord, muscles, and mesenchyme lying on one side of the medullary folds. The only signs of the regeneration of the missing half are the overgrowth of the ectoderm on the naked side, and the passing over of a few muscle-cells ; the full number of the latter is, however, never found on both sides (Fig. 145 a—c).

The % larva is especially interesting, particula.rly when a posterior blastomere has been killed, for here the anterior end is complete with anterior mesenchyme on both sides ; the posterior mesenchyme and muscles are, however, missing on either the right or left (Fig.145(l). The chorda and neural plate are stated to be smaller than the normal in these larvae, and there is no sense-vesicle.

An embryo developed from the two anterior blastomeres of the 4-celled stage alone possesses the full number of neural, chorda, anterior endoderm and anterior mesenchyme cells ; its hind end is covered by ectoderm but shows no trace of muscles or posterior endoderm or mesenchyme (Fig. 146 0, (Z).

The two posterior blastomeres produce an embryo devoid of neural plate and notochord, of anterior mesenchyme and endoderm. By overgrowth of the ectoderm a sort of solid gastrula is formed with a central mass of endoderm flanked by musclecells and posterior mesenchyme. No tail appears (Fig. 147).

21,-, %, and I15 blastomeres also segment and develop partially. An anterior % has half the proper number of neural, notoehordal, anterior endoderm and mesenchyme cells, but the neural plate does not become folded and the chorda cells protrude irregularly behind. A posterior quarter may gastrulate in the way described for the posterior half, but exogastrulae and pseudogastrulae occur. 238 INTERNAL FACTORS IV. I

Conklin has also divided the gastrula into anterior and posterior halves ; neither can regenerate the missing parts, though the cut surface of each becomes covered by ectoderm.

Driesch, however, states that in another form, P/zallzma manmzillata, anterior and posterior half gastrulae will, if

FIG. 146.—~CyntIu'a pm-tita. Anterior half and three-quarter embryos.

(After Conklin.) a, b, anterior ventral three-quarter embryo, the posterior dorsal cells (B 4.1, B 4.1) containing all the yellow pigment

having been killed in the 8-cell stage. a, ventral view; b, dorsal view.

c, d, anterior half embryo: a, dorsal view, showing the neural plate (stip led); d, deeper focus showing two rows of chorda cells (the nuclei are s aded), and ectoderm and endoderm.

separated before complete closure of the blastopore, each develop into a. larva lacking only the otolith and the eye. In a later stage of gastrulation this becomes impossible, although the IV. I INITIAL STRUCTURE OF THE GERM 239

posterior portion of a young Ascidian will regenerate a. new head with pharynx and eyes ; the anterior half dies.

Driesoh has also maintained that the isolated blastomeres (72,, 3., %) of the same species give rise ultimately to total gastrulae and these to larvae. The larvae are, however, unable to hatch out and are defective—with rudimentary eyes, otolith and suckers-or devoid of these organs altogether.

FIG. 147.—Cg/nthz'a parfita. Posterior half embryos. (After Conklin.) a, 32-cell stage, dorsal view. Cleavage is quite normal. b, 76-cell stage, dorsal view. Muscle-cells (B 8.8, B 8.7 and B 7.8) and posterior mcsenchymc cells (B 8.6, B 8.5, B 7.7, and B 6.3) and posterior endoderm cells (B 7.1, B 7.2) are seen. c, later stage, deep focus, showing ventral endoderm (0 end.) with a. mesenchyme (m'ch.) on each side. d, same stage as last, showing muscle-cells (ms.).

LITERATURE L. CHABRY. Contribution a 1’embryologie normale et tératologique des Ascidies simples, Journ. de l’Anat. et de la Phys. xxiii, 1887. E. G. CONKLIN. The organization and cell-lineage of the Ascidian egg, Joum. gicad. Nat. Sci. Philadelphia, xiii, 1905. 240 INTERNAL FACTORS IV. I

E. G. CONKLIN. Mosaic development in Ascidian eggs, Joum. Exp.

Zool. ii, 1905. _ H. Dnmscn. Von der Entwicklung einzelner Ascidienblastomeren,

Arch. Ent. Mach. i, 1895. H. DRIESCH. Ueber Aenderungen der Regulationsﬂihigkeiten im Verlauf der Entwicklung bei Ascidien, Arch. Ent. Mech. xvii, 1904.

§ 11. GENERAL CONSIDERATIONS AND CONCLUSIONS TO BE DRAWN mom ma Fonaaomc Exrammnnrs.

From the foregoing experiments certain general conclusions may now be drawn.

( 1) First, thedivision of the nucleus during segmentation at least is not the qualitative process imagined by Roux and Weismann. The pressure experiments of Hertwig, Driesch, and Wilson have clearly demonstrated that the nuclei of the segmenting ovum may be disarranged without prejudice to the subsequent normal development of the embryo; and this result is amply conﬁrmed by the normal behaviour of eggs (of sea-urchins) in which the mutual positions of the blastomeres, and therefore also of the nuclei, are altered, as well as by the very numerous cases in which it has been shown to be possible to rear a perfect embryo or larva from an isolated blastomere or blastomeres.

It is evident that during segmentation at least the nuclei are equipotential, and the hypothesis of self-diiferentiation in the form originally propounded by Roux can no longer be upheld. It has, in fact, been now abandoned by its author.

(2) But though in this direction its labours have ended negatively, modern experimental research is yet able to point to a positive achievement of the greatest value and signiﬁcance. For the same series of investigations has shown that the cytoplasm of the undeveloped ovum is not the homogeneous and isotropic substance which the experiments of Pﬂiiger led Roux to consider it, but heterogeneous, containing various speciﬁc organ-forming stuffs which are deﬁnitely and necessarily connected with the production of certain parts or organs of the developing animal. If the polar lobe of the mollusc Ilyamzssa be removed the larva has no mesoderm; if the polar lobe of Dentalium be taken away the larva has no trunk and no apical organ; the egg of a Ctenophor IV. I INITIAL STRUCTURE OF THE GERM 241

is capable of segmenting when deprived of its vegetative hemisphere, but the larva to which it gives rise has no combs and no sense-organ. The egg of the Ascidian Cynmia exhibits in its cytoplasm various substances, easily distinguishable by their colour ; in segmentation these are distributed to the various blastomeres in a perfectly deﬁnite way. Remove one or more of these substances and the embryo will lack the organ or organs normally produced from them. Again, a Nemertine egg from which, at a certain stage, the vegetative portion has been cut away, gives rise to a Pilidium without lappets and without a sense-organ.

In these cases, then, at least, the existence of speciﬁc organforming stuffs in the cytoplasm of the egg has been proved; the removal of the stuffs inevitably entails the absence of the corresponding organ later on. But though it may therefore be justly said that such stuﬁs are preformed, it is not invariably true that they are preloealized, present, that is to say, at z'u2'tz'o in what will be their eventual situation. For example, the stuff which determines the formation of an apical organ in the larva of Deutalium is originally in the polar lobe of the vegetative hemisphere : but between the ﬁrst and the second divisions it migrates into the animal portion of the egg ; and the same may be said of the formation of the sense-organ in Cerebratulus and the Ctenophora.

It seems highly probable, though it cannot be said to have been demonstrated, that such specific organ-forming substances are of universal occurrence.

The behaviour of the isolated blastomeres of the eggs of different forms, as well as of the blastomeres of the same form at different stages, is markedly different. The .1, or ,1; blastomere of an Ascidian egg can never produce more than g- or %; larva; the isolated blastomeres of Palella produce just so much, dividing just so many times, as they would have produced had they remained in connexion with their fellows; a radial group of blastomeres (one or more ,1, blastomeres, or two or four or more 113- blastomeres) of a Ctenophore egg gives rise to a larva. with one, two or more costae and meridional canals, as the case may be. On the other hand, the isolated 13 or 71; blastomeres of a Nemertine ovum will produce a. complete la.rva, while, after the next dii./°ision, the totipotentiality is lost; for an. isolated

muxmsox R 24-2 INTERNAL FACTORS IV. 1

animal cell gives rise to a larva without an archenteron, an isolated vegetative cell to one in which the apical organ is lacking. So in Amp/u'o.m4s % and § blastomeres develop into whole embryos, but g blastom eres, whether of animal or vegetative origin, will not gastrulate. In other cases, however, the potentialities do not become restricted so soon. Either the four animal or the four vegetative cells of a Frog’s egg will give rise to an embryo in which the blastoporic lip and archenteron are developed, while in Echinoderms the least blastomere that will gastrulate is 315. It is not, nevertheless, every cell of the 32—celIed stage that is capable of developing an archenteron, but only those, as Driesch has conceded, of vegetative origin.

But whatever differences there may be in the potentialities of the cells of different ova, or of the same ova at different stages or in different regions, it is clear that sooner or later the capacity of a part to become a whole i lost; and it seems only reasonable to conclude that this loss of totipotentiality is due to the loss of one or more necessary cytoplamic substances, to the lack of specific material, and not merely of material. In the egg of Ampﬁiowus, which is telolecithal, there is an obvious difference, in the amount of yolk and size of the granules, between the animal and vegetative hemispheres; a diﬁerence in the capacities of these two regions ha been experimentally demonstrated in the Nemertine egg; and, although it may often appear homogeneous, the ovum of the sea-urchin, in all probability, possesses that ‘stratification’ of potentialities, that is of organ—forming stuifs, at right angles to its axis, on which Bovcri has so strongly insisted, for Driesch has admitted that vegetative cells ‘gastrulate’ more readily than animal cells ; it would appear, then, that a gutforming substance is present, and a mesenchyme-forming substance too, but that the concentration of these substances steadily diminishes from the vegetative to the animal pole. Radially about the axis the distribution of these substances may be taken to be uniform, since meridional fractions of the egg, %, §, % or g or g blastomeres, are invariably totipotential.

The same explanation may be applied to other cases. Hence, speaking generally, the limitation of the potentialities of the parts will depend simply on the original distribution of the IV. I INITIAL STRUCTURE OF THE GERM 243

substances in the unsegmented egg. Should they be already segregated (wholly or in part) in distinct regions, as in Ascidians, Molluscs, and Ctenophors, then even the first blastomeres will to that extent be unable, when isolated, to give rise to more than they would have done in the uninjured egg; should theybe uniformly distributed about the axis, but unequally along the axis (Nemertines, Echinoderms, Amp/u,'oams), then a limitation of potentialities will appear when the third division separates the animal from the vegetative cells‘ ; should the arrangement be

FIG. 148.—Diagran1matic sections through the unsegmented egg of Lolig/o pealii. (From Korschelt and Heider, after Watase.) A is in the transverse, B in the sagittal plane of the future embryo. In B the anterior side (m.) is more convex than the posterior (h.). d, dorsal (animal pole); v., ventral (vegetative pole); 1., left; r., right. The protoplasm is black, the yolk shaded.

uniform about the egg-centre, then animal and vegetative blastomeres will be alike totipotential (Coelenterata).

At the same time it would appear to be true that the capacity of total development may be lost for mere lack of material, for Drieseh found that while all four (or all eight in the next stage) animal cells of the sea-urchin egg might give rise to a Pluteus larva, the isolated cells would not. The isolated cells may, how ‘ We may notice that O. Maas (Arch. Ent. Mech. xxii, 1906) has found that the isolated anterior ﬂagellated and posterior granular cells of the Amphiblastula larva of S3/can are not equipotential. The latter can ﬁx and give rise to a. young sponge, the former cannot. 244 INTERNAL FACTORS IV. I

ever, gastrnlate ; when once the gastrula stage has been reached, therefore, further development may be simply a matter of sufficiency of substance. So again the octants of the Ctenophore egg are alike, but each can produce one costa, and one only. There are then certain preformed organ-producing‘ substances, and the arrangement which they either possess before, or acquire

"‘l.'tnL" FIG. l49.—Three segmentation stages in the blastoderm of Sepia aﬁciualis; the segmentation is of the bilateral type. l, left; r, right; I—V, first to ﬁfth cleavages. The top sides of the ﬁgures are anterior. (After Vialleton, from Korschelt and Heider.)

during, segmentation determines the ﬁxed relation which can be observed in almost all cases between the structure of the egg and the axes of the embryo. Thus in Amphibia the head of the embryo is formed at or near the animal pole, and the plane of egg symmetry becomes the sagittal plane. In Mollusca (not

Cephalopoda) and Annelida the apical sense-organ is developed at the animal pole, while the D quadrant is posterior. A blasteTV. I I-NITIAL STRUCTURE OF THE GERM 245

pore is in very numerous eases formed at the vegetative pole, the bilateral symmetry of the eggs of Cephalopoda (Figs. 148, 14-9), Inseeta, some Crustacea, and Aseidia becomes that of the embryo; in Ascaris the ﬁrst four cells lie in one plane, which becomes the median plane, the germ-cell is posterior and the animal pole dorsal; and soon.

(3) The instances in which the segregation of the organforming substances into the blastomeres takes place at once are especially interesting, as they appear to fulﬁl exactly the requirements of the ‘ Mosaik-theorie ’, if that doctiine be transferred from the nucleus to the cytoplasm, for here, undoubtedly, segmentation is a qualitative process, a sundering of potentialities. It must be pointed out, however, that even in these cases the factors which determine segmentation and those which determine diﬂ’erentiation may be as distinct as they are elsewhere.

In the Frog there is a much greater correlation between the symmetry of the egg and the median plane of the embryo than between the former an(l the plane of the ﬁrst furrow.

A % or % blastomere of a Nemertine or a sea-urchin segments as a part, and yet it eventually develops into a whole larva: and the same is true of egg-fragments. All the isolated blastomeres of a Molluse segment as parts, yet in 1)ental2'um, that one which contains all the necessary stuﬂfs, CD or D, can give rise to a complete embryo, while its fellows cannot. The converse case is to be found in Ctenophors ; here the egg which has been deprived of its vegetative portion segments as a whole and yet produces an embryo devoid of eostae and sense-organs.

There are also certain forms (/Imp/u'o.'ru8 and V61 tebrates, Coelenterata) where the isolated cell segments as a whole, and this appears to depend on the rapidity with which the part can rearrange its material or resume the polarity of the whole (Wctzel). It is further to be noticed that the capacity of a part for total segmentation,like its power of total differentiation, may differ in the same egg at diﬁ’erent times and under diﬁerent conditions ; the fragment of an immature egg of Care//ratulua divides Iikea whole egg, an isolated blastomere like a part ; again, in Ilyanaaaa the cleavage of a part may be made total by lowering the temperature.

When, further, it is remembered how closely similar the 246 INTERNAL FACTORS IV. I

geometrical pattern of segmentation may be in the eggs of different animals, whether of the same or of diiferent groups (spiral segmentation of the eggs of Molluscs, Chaetopods and Turbellaria, similar segmentation of the eggs of, for example, some Echinoderms, Polyzoa, and Vertebrates), without necessarily involving a similarity in the fate of cells which are identical in origin, it will, I believe, be conceded that the factors which are responsible for cleavage and those which determine diﬁerentiation are distinct, though the two may, as in the socalled ‘Mosaik’ segmentations, coincide: and where, as in Ascidians, the Ctenophora, the Cephalopoda, this coincidence is complete, the symmetry of the egg, the symmetry of segmentation, and the symmetry of the embryo are one.

The former factors must be sought for in the arrangement of yolk and protoplasm, possibly in the relative strengths of the centrosomes, in surface-tensions, in the pressure exerted by eggmembranes and so forth ; the latter in deﬁnite cytoplasmic organforming stulfs.

(4) Like the parts of the egg the parts of the elementary organs of the embryo are at ﬁrst equipotential, but exhibit a gradual limitation of potentialities as their development proceeds. As Minot puts it, there is a ‘ genetic restriction ’ of capacities. Thus the arehenteron of Asterias can form a new terminal vesicle when this has been removed, but only before the outgrowth of the coelom sacs. The anterior half of the newt embryo will develop into a whole embryo when out off before the medullary folds have arisen, but not after. When one vitelline vein of a chick is destroyed its fellow will form a. whole, not a half heart; and Sumner’s experiments quoted above have shown that the margin of the Teleostean blastoderm is isotropic.

(5) There still remains to be noticed a point of great importance. It certainly has not been demonstrated, and it cannot be pretended that there are in the developed egg as many speciﬁc substances as there are separately inheritable qualities in the body. On the contrary, the evidence of experiment speaks against such a supposition; for, as Driesch has urged, were every separately inheritable quality to be represented in the germ by a distinct morphological unit, the equipotentiality of IV. I INITIAL STRUCTURE OF THE GERM 247'

the isolated parts would be inconceivable. Although the formation of the larval and elementary organs (germ~layers) may depend upon such stuﬂfs, yet within each such elementary organ the parts are at first and remain for a time equipotential.

Diiferentiation is progressive. The origin of the primary differentiations has been found; it remains for the experiments of the future to discover how fresh distinctions arise until the organization of the whole is complete. Suggestions as to the importance, in this respect, of certain other internal factors have been made, notably of the structure of the nucleus, and the reactions of the parts on one another. These suggestions, for they are hardly more than that at present, we shall have to discuss. First, however, we must brieﬂy consider the part played by the spermatozoon in determining the structure of the egg and the symmetry of the embryo.

§ 12.

It is of the greatest interest to observe that the deﬁnitive distribution of these speciﬁc substances in the unsegmented egg, and so the determination of the embryonic axes, may be brought about, partly at least, by the spermatozoon.

In the Frog (Ifamz fusca, It. temporaria, and In’. 1Jalu3lrz's, probably also 1?. esculenta) the egg soon after fertilization becomes bilaterally symmetrical. As first Roux and later Schulze have shown, this is due to the appearance on one side of the border of the pigmented area of a grey crescent, caused by the retreat of the pigment into the interior. The grey crescent subsequently becomes white and added to the white area. The side on which it appears is opposite to that on which the spermatozoon had entered, and later becomes the dorsal side of the embryo ; there is also a considerable correlation between the plane of symmetry‘thus bestowed upon the egg and the future sagittal plane, since the latter lies in a large majority of cases in or close to the middle point of the crescent.

This relation between the point of entry of the sperm and the egg- and embryonic symmetry has also been experimentally demonstrated by Roux, who, by applying the sperm to any arbitrarily selected meridian of the egg (by means of a fine cannulus, _a brush, or a silk thread), was able to show that 248 INTERNAL FACTORS IV. I

fertilization can take_ place from any meridian, and that the point of entry so selected becomes the ventral side later on; in other words, the fertilization meridian becomes the sagittal plane. This, as we have seen, is true, or approximately true. Roux, however, believed that the plane of the first furrow also coincided with the other two, in fact that it was the ﬁrst furrow which determined the sagittal plane, and he brought forward evidence to show that the first furrow, if it did not pass through the point of entry of the sperm, at least either included, or was parallel to, the inner end of the crooked path of the spermatozoon within the egg, the so-called ‘ copulation ’ path (Fig. 150). As we now know there is very little correlation between the first furrow and the sagittal plane. Nevertheless Roux’s work remains of

FIG. 150. —Roux’ diagrams to show the relation of the sperm-path (Pig/.) to the ﬁrst furrow in the Frog's egg. In A the furrow includes the sperm-path, in B it is parallel to it, in C it is parallel to the inner portion of the path (copulation path), in D it includes only the very last portion of the copulation path. (From Koischelt and Heider, after Roux.)

the greatest significance, for it seems extremely likely that, while it is the actual point of entrance of the sperm and the ﬁrst, radially directed, part (‘penetration path’) of its track within the egg, which determines the position of the grey crescent, and so of the sagittal plane, it is the second part which determines the direction of the first furrow, since the centresome will divide at right angles to this ‘copulation’ path, or line of junction of the two pronuclei; the axis of the fertilization spindle is of course given by the line of separation of the two centrosomes, its equator by the plane including the two pronuclei and the egg-axis, and this spindle-equator is the plane of the first furrow.‘

In the egg of the sea-urchin Tomopzzezw/es the deﬁnitive eggaxis would appear to be ﬁxed not by the original position of the egg ‘ See, however, Appendix A. IV. I I°NITIAL STRUCTURE OF THE GERM 249

nucleus at all, but by the position of the excentric segmentation nucleus. Both male and female pronuclei move through the egg for a longer or shorter distance till they meet, and the combined nucleus then takes up its deﬁnitive excentric position (Fig. 151). The plane of the first furrow is given by the egg-axis and the point of entrance, approximately, of the spermatozoon, which may thus be said to determine the symmetry of segmentation, and so of the embryo, since this plane becomes, it is said (Selenka,

FIG. 151.—D1ag1~ams from successive camera. drawings of the living eggs of Toaropneustes, to show the determination of symmetry during fertilization.

The original position of the eg nucleus is shown by the position of the dotted circle marked 9; its pa.t by the succession of such circles.

E, the entrance-point of the spermatozoon, and cone of entrance. The male pronucleus is rendered in black, its path marked by the line uniting the black dots.

M is the meeting-point of the pronuclei. The cleavage nucleus, c, is shaded; its successive positions are indicated. The axis of the fertilization spindle is shown by the line with a small circle at each end passing through the cleavage nucleus; the arrow passing through this is the deﬁnitive egg-axis, its point the animal pole. The cleavage nucleus is slightly excentric, and nearer the animal than the vegetative pole.

F is the meridian of the first furrow.

A and B are opposed in polarity, similar in other respects. 0 and D differ in polarity, and otherwise as well. (After Wilson and Mathews.) 250 INTERNAL FACTORS IV. 1

Garbowski, Driesch), the median plane of the Pluteus. In this case there is practically no distinction between ‘ penetration ’ and ‘copulation’ parts of the sperm-track, and, the centrosome dividing perpendicularly to this path, the point of entrance naturally lies in, or nearly in, the first furrow. The other alternative, as Wilson has pointed out, is to suppose that the deﬁnitive polarity of the egg existed before fertilization, but exerted no inﬂuence upon the egg-nucleus, although able subsequently to compel the fertilization nucleus to take up a position in its axis.‘ There are other Echinoderms, however (:1a5er2'a.9, Stroizyyloccietrotus), in which the original and the deﬁnitive egg-axes coincide.

Another very interesting case is Uynt/zia (Fig. 142). In the immature egg of this Ascidian the cytoplasm is concentrically arranged, since the yellow pigmented protoplasm forms a peripheral investment for the central grey yolk: the egg-axis is determined only by the position of the germinal vesicle. Upon the entrance of the spermatozoon the yellow substance together with the clear plasma of the germinal vesicle, which has in the meantime broken down, streams towards the vegetative pole and becomes radially arrayed about the axis. By a further streaming movement these two substances are carried up with the male pronucleus—— or pronuclei in cases of dispermy—to the equator, so marking the future posterior side of the now bilateral egg. These changes are evidently an eifect of fertilization: it cannot, however, be asserted that the point of entrance of the sperm determines the posterior end, since it may enter on the opposite side and get carried across.

There are a few other instances in which a11 alteration of the egg-structure has been noticed to follow on fertilization. In Ctenophora the peripheral layer of granular cytoplasm becomes aggregated at the animal pole (Agassiz) ; in Polyclads (Lang) and Cirripedes (Groom) the egg becomes telolecithal ; and so in P/rzyew

‘ In Diplogaster longicauda also it appears to be the point of union of the two pronuclei which determines the deﬁnitive egg-axis and orientation of the embryo. The polar bodies are always extruded and the female pronucleus formed at that end of the ellipsoid egg which is turned towards the ovary, while the sperm enters at the opposite end. The pronuclei may meet and units at either end; but it is at the end where their union takes place that the smaller of the first two blastemeres is found and, later, the binder end of the embryo (H. E. Ziegler, Zeitschr. wise. Zool. lx, 1890). IV. I INITIAL STRUCTURE OF THE GERM 251

(Kostanecki and Wierzejski): the ‘polar rings ’ appear in Allololzoja/tom, Clepsine, and Rhync/aelmis (K. Foot, Whitman, Vejdovsky), while in Teleostei the periblastic hyaline layer becomes concentrated as the blastodisc at the animal pole. The case of Dentalium has been already referred to (p. 223, Fig. 133).

Although the cases in which the role played by the spermatozoon in determining egg and embryonic structure are, as has been observed, not very numerous, yet it is fully to be expected that renewed investigation will show that some such rearrangement of the materials of the egg is in many, perhaps in all, instances one of the ﬁrst results of fertilization.

LITERATURE

A. AGASSIZ. Embryology of the Ctenophorae, Mcm. Amer. Acad. Arts and Sciences, N.S. x, 1873.

E. G. CONK LIN. The organization and cell-lineage of the Ascidian egg, Jam-n. Ac. Nat. Sci. Philadelphia, xiii, 1905.

K. Foo'r. Preliminary note on the maturation and fertilization of the egg of Allolobophora foetida, Journ. Morph. ix, 1894.

T. T. GROOM. On the early development of Cirripedia, Phil. Trans. Roy. Soc. elxxxv, B. 1894.

K. VON KOSTANECKI and A. WIERZEJSKI. Ueber das Verhalten der sogen. achromatischen Substanzen im befruchteten Ei (Physa fontinalis), Arch. milcr. Anat. xlvii, 1896.

A. LANG. Fauna. und Flora des Golfes von Neapel: XI. Die Polycladen, Leipzig, 1884.

W. Roux. Die Bestimmung der Medianebene des Froschembryo

(lurch die Copulationsrichtung des Eikernes und des Spermakernes, Arch. mikr. Anat. xxix, 1887; also, Ges. Abh. 21, Leipzig, 1885.

F. VEJD0vsK1'V. Entwickelungsgeschichtliche Untersuchungen, Prag, 1888-92.

C. 0. WIIITMAN. The embryology of Clepsine, Quart. Jouru. Micr. Sci. xviii, 1878.

E. B. WILSON and A. P. MATHEWS. Maturation, fertilization and polarity in the Echinoderm egg, Journ. Morph. x, 1895.

§ 13. WIIAT PART DOES THE NUGLEUS PLAY IN DIFFERENTIATION ?1 We have already criticized and rejected the hypothesis that the division of the nucleus during segmentation is a qualitative process. It may still be urged, however, that the nucleus is not

‘insigniﬁcant in dilferentiation. In support of this contention

the following arguments have been put forward. 1. It is urged that the nucleus is essential for the life of the cell,

1 See also Appendix B. 252 INTERNAL FACTORS IV. 1

and not only for its metabolism (Figs. 152, 153) but for the production of form (Fig. 154).

2. It is obvious that the germ-cells are the vehicles whereby the inheritable characters of the species are transmitted from one generation to the next; they are the material basis of inheritance. The germ-cells of the two sexes are, however, as unlike as possible in every character except their nuclei, in which, as the study of maturation has abundantly shown, they are exactly alike. In fertilization, the union of the two germ-cells, two distinct processes are involved. The first is the mutual stimulation whereby the lost _ power of cell-division is restored. This ',5 ' is a process independent of the nuclei (in Metazoa). The other is, however, the union of the nuclei (or their chromosomes); and since the offspring are held to inherit equally from the two parents, it has been - - supposed that with the union of the nuclei, the only parts of the cells that are

FIG. 152.——Egg-cell of

Dytiscus mwginalis in its follicle with two nurse cells. From the nurse cells nutritive material passes into the egg-cell, towards which the nucleus

exactly alike, the paternal and maternal contributions are intermingled, and that therefore it is in the chromatin of the nuclei that the vehicle of transmission of hereditary qualities must be looked for.

sends out pointed pseudopodia. (After Korschelt, from Korschelt and Heider.)

3. Thirdly, it has been pointed out (originally by Roux) that karyokinesis appears to be a mechanism expressly adapted to the simultaneous division of a large number of qualitatively different units. From the existence of karyokinesis it is argued that the nucleus is not homogeneous, and that in development the various units of which it is composed are directly concerned in the process of dilferentiation.

The doctrine of the individuality of the chromosomes has been brought forward in support of this belief, as well as the

constancy of their number. 4. The diminution of the chromosomes. Boveri has observed

in Aacmia, in the nuclei of purely somatic cells, apeguliar process IV. I INITIAL STRUCTURE OF THE GERM 253

to which he has given the name of the diminution of the chromatin. The middle parts of the chromosomes in somatic cells become divided transversely into small granules, the ends remain rodshaped. The granules of the middle parts are alone divided and

pass to the daughter nuclei, while the ends are cast out into the cytoplasm and there degenerate. The chromosomes of the germ-cells (or of the parent-cells of the germ-cells) do not undergo this change, but remain intact (Fig. 155).

It might be argued that in such a. case as this the determination of certain characters —-somatic-—is brought about by the expulsion of the chromatin into the cytoplasm.

5. A great deal of stress has been laid on the importance of the experiment, due to Boveri, in which the enucleate fragment of the egg of one species of sea-urchin gives rise, when fertilized with the sperm of another species, to a larva _ _ _ which exhibits the charams i..F’1»‘TI..i§’.3 3?‘.-:...‘3.‘£T"""q~i‘.’.f ‘.”...’l.‘i‘.§f.'.i'“iL'

of the male parent alone. placed at the point of origin. B, the

- same in Cmurbita pepo. C, root-hair 6' Boven has brought for of Cannabis .-atim: the nucleus is at

ward evidence to show that the growing point. (After Haberlandt, the abnormal development of from Korschelt and Ileider.) Echinoderm eggs which follows on polyspermy is in reality due to the irregular distribution of the chromosomes to the daughtercells. Hence it is argued that the chromosomes are qualitatively diﬂferent.

These are the reasons which are, or have been, brought forward in support of the belief that the nucleus is not insigniﬁcant in diﬁerentiation. We may now discuss them in order,

A B C 254 INTERNAL FACTORS IV. 1

premising only one thing, namely, that both maternal and paternal nuclei are certainly not necessary for the production of a new individual, however beneﬁcial it may be for the elements of two parents to be commingled in the body of one oﬁspring. In artiﬁcial parthenogenesis an egg is stimulated to development by a solution, and produces a normal larva; this creature possesses nuclei of maternal origin. The

_ e converse case is seen in the

W "K "- i ‘ phenomenon of ‘ merogony’.

M An enueleate egg-fragment, fertilized with sperm of the

!!lll|

same species, gives rise to a new organism, provided only with paternal nuclei.

The older view that the union of the pronuclei was the essential act in fertilization ean no longer be held, for, certainly, both nuclei are not necessary. The most, therefore, that can be said will be that a complete set of the chromosomes (or smaller

chromatin units) of the species FIG. 154.——0n the left the protozoan

Stentor, cut into three pieces, each con- ls necessary for d1ﬁerentla't10n'

tainingha piecle of thﬁ nucleus. On Whether this is so or not we the rig t, eac piece as regenerated - - , the missing parts and become a com— must now mqune'

lete Stentor. (After Gruber, from 1. It will of course be al°“°h°1t “ml Heme") lowed thatthe nucleus is necessary for the life of the cell and to a certain extent for the assumption of form. The numerous and familiar experiments on Protozoa have sufficiently established this point} A nucleate piece will live and regenerate lost parts, an enucleate piece will not (Fig. 154-). 2. It will also be admitted that the maturation processes in the germ-cells of the two sexes are extraordinarily alike and end in the reception by each germ-cell of one-half the normal ‘ See especially M. Verworn, Die physiologisehe Bedeutung des

Zellkerns, 12ﬂ1'1ger's Arch. li, 1892 ; in this paper an account of the work of Gruber and others will also be found. ‘ IV. 1 INITIAL STRUCTURE OF THE GERM 255

FIG. l55.——The process of chromatin diminution as seen in the somatic cells of Ascaris megalocephala. (After Boveri, 1899.?

1. Mitosis in the 2-celled stage. In the first somatic cel (S, or AB), the primary ectoderm, the chromatin undergoes diminution, not in the germ-cell (Pl). la chromosomes being ‘diminished’.

2. 4-cell stage, T-shape. In A and B the discarded masses of chromatini are seen. S, (EM St), second somatoblast or endomesostomodaeal ru iment.

3. 4-cell stage, lozenge-shape. In A and B the next mitosis is beginning, in P, and E M St the nuclei are in the resting stage. A is anterior, 2:] and g are dorsal, all four cells lie in one plane, the sagittal plane of t c em ryo.

4. In EM St the chromatin is being diminished. Division of P, into P3 and S3 (C), the secondary ectoderm. a, b, primary ectoderm of right, a, 8, primary ectoderm of left side.

5. The endoderm (E,, E,) has now been separated from mesoderm and stomodaeum. P3 has just divided into P4 and S‘§d)' tertiary ectoderm.

6. Diminution of chromatin in S, (D). The our endoderm cells (E) beginning to be invaginated: on each side two mesoderm cells (M) in which franular chromosomes may be seen, and two stomoclaeal cells (St). Ventra. view. 256 INTERNAL FACTORS IV. 1

number of chromosomes of precisely the same shape and size. In fertilization the two sets of chromosomes—each in half the normal number-are united. It is of course difficult to believe that this extraordinary resemblance of the nuclei, while all the other characters of the cells are unlike, is without signiﬁcance.

3. With regard to the third point a distinction must be made between two hypotheses. The ﬁrst is the individuality of the chromatin, the second the individuality of the chromosomes.

For the ﬁrst, as we shall see, independent evidence exists, and mitosis is certainly a mechanism admirably adapted to the simultaneous division (or separation of already divided halves) of a large number of qualitatively distinct bodies.

But the second hypothesis in no way follows from the first, for the grouping of the unlike units may obviously be different at each successive division without in the least impairing their individuality. There is, indeed, evidence for the persistent individuality of the chromosomes in only one or two cases. Boveri has noticed, in the segmenting egg of A.<:carz'.s-, a constancy in the position of the pockets of the nuclear membranes which lodge the ends of the chromosomes, and a similarity in the arrangement of the chromosomes in the nucleus in successive nuclear divisions. Sutton again has observed that in Bracﬁ_g/stola each chromosome forms a separate reticulum, situate in a separate pocket of the nucleus. There are other cases in which each chromosome forms its own vesicle (Echinoderm eggs). These instances are, however, few and far between. There are so many nuclei in which nothing can be observed of the chromosomes in the resting stage, often there is nothing but a fine granular mass, and a study of the early and end stages of mitosis seems to show that the chromosomes are precipitated from solution in the nuclear sap and redissolved when the division is over.

In such a solution the chromatin elements would retain their individuality, their qualitative diﬁerences, just as each one of a number of crystals retains its distinctive properties in a mixed solution and exhibits them when recrystallization occur. In such a sense, and such only, is it possible to speak in general of the individuality of the chromatin. '

4. With regard to the diminution of the chromosomes in IV. I INITIAL STRUCTURE OF THE GERM 257

Ascm-is, Boveri has pointed out that the extruded outer ends of the chromosomes are very irregularly distributed to the daughtercells, whereas they should surely, if they are determinants, be equally divided between the two. More than that, however, Boveri has found evidence which shows that not only does the diminished chromatin not determine the characters assumed by the cytoplasm, but that on the contrary it is the cytoplasm which decides which chromosomes shall be diminished, and which remain intact.

Boveri has observed certain cases of dispermy in Ascaria, which are followed by simultaneous division of the ovum into four, consequent, presumably, on a quadripolar mitosis which is due, in turn, to the presence of an extra pair of centrosomes. The total number of chromosomes present in such eggs is 3 n, where n is the reduced number, since each spermatozoon introduces n. This number becomes doubled by division, and the number is then 6n or 12, since 1» in Ascm-2'3 megalocep/zala v. bivalens = 2. These twelve chromosomes have to be distributed over the four cells into which the ovum divides: their distribution is irregular. The next division is described as being tangential in two on? the cells, at angles of 45° to the first divisions in the other two: it leads to the formation of two groups of four cells each, in each of which the cells are arranged in the T shape characteristic of the normal four-celled stage. The normal four-celled stage is reached by (1) an equatorial division, followed by a meridional division in the animal cell (/1 B), and a latitudinal division in the vegetative cell, which separates a cell S2, which like A and B is somatic, from a cell P, at the vegetative pole. The chromosomes in P2 are intact, in A, B, and 82 they are diminished (Fig. 155).

Boveri argues that if the diminution of the chromosomes were an intrinsic property of the chromatin, there should always be 6 (3 7;) chromosomes intact and the rest diminished, just as there are always 4« (2 n) intact in the normal egg, no matter how the chromosomes had become distributed over the ﬁrst four cells and their descendants. This, however, is not the case. Whole chromosomes are found only in those cells——one in each group of four—which correspond in position to the P2 cell of the

JENKINSON S 258 INTERNAL FACTORS . IV. 1

normal egg; the chromosomes in the other cells are all diminished, and no cell contains chromosomes of both kinds. Further, the number of whole chromosomes may be 5 in one cell and 3 in the other, or 4 and 3, or 3 and 3, or 4 and 2, or 3 and 2.

Boveri concludes, it seems with perfect justice, that it is the (vegetative) cytoplasm which has determined that the chromosomes it contains shall remain whole, while, contrariwise, the animal cytoplasm of the other cells brings about the process of diminution.

In Ascaria lumbricoizles there is a diminution of the chromosomes very similar to that seen in A. megalocep/mla, and, to judge from

FIG. 156.—Pluteus of_Sphaerechz'nus granularis from in front and from the side. (After Boveri, 1896.)

certain abnormalities similar to those just mentioned, brought about by the same means (Bonnevie). In Dytiacus there is something like a diminution of the chromatin in the nurse-cells, while the oocyte retains all the chromatic material intact (Giardina). The distinction seen here may, however, only be that observed in all ova between the nutritive and reproductive functions of the nucleus (trophochrornatin and idiochromatin).

5. Boveri’s hybridization experiment, on which rests the assertion that the nucleus alone conveys the inheritable characters of the species, was as follows.

The Plutei of Sp/laerec/ainua _qranuZarz's and of Ecﬁinus microtuberculatue were found (at Naples) to be markedly diﬁerent-. IV. I INITIAL STRUCTURE OF THE GERM 259

The larva of the former (Fig. 156,) is short and squat, the oral lappet not divided into lobes, and the skeleton provided with a fenestrated anal arm—produced by three long parallel bars united by numerous cross-bars--an apical branch to the oral arm, and, at the apex, a square ‘frame ’ formed by the union of twigs from the last mentioned and from the apical arms.

The larva of Ea/limzs, on the other hand, is long and lank, the oral lappet is deeply cleft, and in the skeleton the anal arm is not fenestrated ; there is no apical branch of the oral arm, and the ex

FIG. 157.-—Pluteus of Echinus mhromberculatus from in front and from the side. (After Boveri, 1896.)

tremity of the apical arm is thickened and club-shaped (Fig. 157). Boveri ﬁrst fertilized the ova of Spﬁaerecﬁimw with the sperm of Ea/zz'mw, and so produced a hybrid larva whose characters were intermediate between those of the two parents in the following respects—in form, being shorter and broader than that of Ea/ainus, longer and narrower than that of Spﬁaerec/zinus, in having the oral lappet slightly divided, and in the skeleton, the extremity of the apical arm being swollen and branched, the anal arms being double but not fenestrated, and the oral arms being (sometimes) provided Vvith an apical branch (Fig. 158). s 2 260 INTERNAL FACTORS IV. I

A mass culture of egg-fragments of Sp/Eaerec/2imw—containing presumably whole eggs, and nucleate and enucleate fragmentswas then made, and this was fertilized with the sperm of Ea/tinua. Amongst the larvae developed in such a culture Boveri found a small number (twelve) of dwarf individuals which possessed the paternal (Ec/zimw) characters alone (Fig. 159 6), and he suggested that these had come from enucleate fragments of eggs. He was further strengthened in this opinion by the fact that the nuclei were small, which he attributed to their containing only one-half the normal number of chromosomes.

This conclusion has been adversely criticized by Seeliger. Seeliger, working at Trieste and subsequently at Naples, has

FIG. 158.—Pluteus of the_ cross Sphaerechinus granularis 9 x Echinus microtuberculatus c7’, from in front and from the side. (After Boveri,

1896) pointed out that the diiferences between the Plutei of these two sea-urchins are not as great as Boveri asserted them to be—-the extremity of the apical arm in I]c/ainus is, for instance, not always club-shaped—and that therefore it cannot be so conﬁdently asserted that the hybrid is intermediate. Secondly, he asserts that in an ordinary hybrid culture (whole eggs of Sp/zaerecﬁz'nus fertilized by sperm of Ea/timw) there are to be found together with variable larvae of more or less an intermediate type, individuals with purely paternal characters, though the pure Sp/merecﬁz'nu.s type never occurs, and that the nuclei are variable in size. This contention has been upheld by Morgan, and, after a good deal of controversy, has ﬁnally been admitted by Boveri himself. No conclusion can therefore be drawn from the original experiment. IV. I INITIAL STRUCTURE OF THE GERM 261

An explanation of the discrepancies in the results of the diﬁerent observers has been offered by Vernon. Vernon has found that Stranyylocentrotus livirlus (which has a Pluteus almost exactly like that of Ea/ainzw) is at a minimum of sexual maturity in the summer months, but that from that time onwards the power of the male to transmit its characters to the hybrid larva, when crossed with Sp/aaerec/mms 9 , increases, and that it is possible to obtain a culture consisting entirely of larvae of the paternal type, in respect, that is to say, of the skeleton. Vernon was inclined to look upon this difference in maturity as a seasonal variation, but Doneaster has since attributed it to the effect of temperature alone.

The transmission of the characters of the skeleton may therefore be the function of the nucleus, though, as just pointed out, there is no stringent proof of this. There are other larval characters, however, which, as Driesch has justly urged, must depend on the cytoplasm of the ovum, the colour, for example, if the egg is pigmented, and the size of the larva, so far as this depends on the size of the egg. From a series of hybridization experiments between Fm. 159"”, Fully

E0/ainus, :5):/uwrec/zi7ms, Stroizgz/loce7zt1'otu3, fonned platens of E-ch,-_

and /lrbacia, Driesch indeed concludes gut; m1'c(21'otubefr1culatusf.‘ that the rate of cell-division up to the tile °1i,1,:l,§, E’},i,-,,,:,“;:;e formation of the primary mesenchyme, r<leared(?from fan enu - _ - e eatc )c<rg— raoment the vacuolation of the ectodeim cells (in of Sphamf,=,h,.,m;* ,.er_

S};/merec/zines), the number of the primary tilized with the sperm mesenchyme cells, are all characters which ﬁve (After depend on the ovum, and the ovum alone, i i

and therefore on its cytoplasm. With regard to the skeleton Driesch admits that this, and hence, indirectly, the shape of the larva, may be inﬂuenced by the sperm, at least when the egg of Sp/taercc/¢z'nu8 is fertilized either by Ea/Linus or Strongyl0centrotus. .Fischel has made similar experiments and arrived

at a like conclusion. 262 INTERNAL FACTORS IV. I

Boveri has traversed some of these statements of Driesch’s. The number of primary mesenchyme cells is, he maintains, a mean between the paternal and maternal number, and the form of the larva may be intermediate before the skeleton is developed. Between these contradictory opinions it is not easy to decide, but it may be mentioned that the number of mesenchyme cells is peculiarly liable to variation, as Driesch has himself found.

Before concluding this section we should mention some most interesting experiments on heterogeneous cross-fertilization.

Loeb was the first to show that by the addition of a small quantity of calcium chloride and sodium hydrate to sea-water, it was possible to fertilize the eggs of Stron_9gylocent1-otu.s- [)mp1tmine with the sperm of Arte)-ias ockracea. The matter has since been more completely investigated by Godlewski, who has succeeded in rearing Plutei from the eggs of sea-urchins fertilized with the spermatozoa of the Crinoid Amfedon. The necessary condition of success is the prior treatment of both eggs and sperm with an abnormally alkaline sea-water (about 1'75 c.c.

T73 NaOH per 100 c.c. of sea-water).

The sea-urchins employed were Sp/merec/«mus, 8trou_qylocentrotua, and Ecﬁimzs. The eggs of the first reached the gastrula stage, of the two last the Pluteus. Fertilization was found to be perfectly normal, with production of a vitelline membrane, rotation of the sperm-head, and so on. Cleavage was of the Echinoid type (maternal) with formation of micromeres at the fourth division; no such micromeres occur in Antetlon, the fourth division being meridional in both hemispheres. Primary mesenchyme was formed (this again is absent in Anterlou), but the number of cells was slightly in excess of the normal: and after gastrulation the skeleton of the Pluteus was developed (except in 15'];/(ae7'ec/tinzts); the larva of Antedon of course has no skeleton.

The Antedon chromosomes persisted and the nuclei of the hybrids were intermediate in size between those of the parent forms.

When an enucleate fragment of the egg of Ecﬁimw was fertilized by Antezlon all the processes mentioned above were normal, except that segmentation was irregular, and that development ceased after gastrulation. The death-rate of these larvae was high. IV. I INITIAL STRUCTURE OF THE GERM 263

6. It remains for us now to consider the conclusions which Boveri has drawn from the behaviour of eggs in which owing to pathological mitosis the distribution of the chromatic elements was irregular.

The investigations of O. and R. Hertwig had previously shown that if the eggs of sea-urchins were weakened by treatment with poisons (strychnine, quinine, and others), then the vitelline membrane which normally prevents the entrance of more than one spermatozoon was not formed, that consequently several

FIG. 160.——Diagram of one case of irregular chromosome distribution in adoubly fertilized egg; an b. , c,, and d, ; a,, be, c,, and d2,and as, b,, c3; and d, are the three complete, speciﬁc sets of unlike chromosomes. (After Boveri, 1904.) '

spermatozoa entered, with the resulting formation of multipolar mitoses and irregular cleavage.

Boveri has succeeded in inducing dispermy in the eggs of sea-urchins without the use of the poisonous reagent, by simply adding an excess of sperm to the eggs. The number of abnormal larvae found in such a culture is greater than that observed when the quantity of spermatozoa used is less, and in proportion to the number of dispermic fertilizations.

In dispermic eggs there is an extra pair of centrosomes, and these two usually form with the other two a quadripolar ﬁgure with four spindles occupying the four sides of the square(Fig. 160). In the equators of these four spindles are placed the chromosomes, 264 INTERNAL FACTORS IV. 1

the number of which is 39: x 2, where n is the reduced number (in E'c/limzs 9), since there are two male and one female pronuclei and all the chromosomes have divided. Cell-division occurs across each spindle and the whole breaks up simultaneously into four. To these four cells the 37: x 2 chromosomes are distributed, and, as Boveri points out, the chance of each cell obtaining a complete set of the n chromosomes of the species (supposing these to be diﬂferent from one another) is

C (1

FIG. 16l.——La.rvae from doubly fertilized eggs; (I, of SphaerechimI.s_4/r_anu?qris; b—d, of Echimts micrombcrt-ulatus; a, r, and (Z are from tripartite, b from a quadripartite egg. (Aft-er Boveri, 1904.) small. Such eggs, as a matter of fact, practically always give rise to abnormal larvae. Out of 1,170 examined not a single one was normal.

The chance, however, that one cell of the four will receive a complete set of chromosomes is far greater, and when the =5 blastomeres were isolated it was found that 25 % gastrulated, though none gave rise to Plutei. Moreover, all four developed differently. IV. I INITIAL STRUCTURE OF THE GERM 265

Again, by shaking the eggs it is possible to prevent the division of one centre. A tripolar mitosis is the result and a simultaneous division into three. Out of 695 such eggs 58 became normal Plutei. This is also in accordance with the theoretical expectation, for the probability of each of three receiving a complete set of chromosomes when there are 3 u x 2 to be distributed is much greater than when these chromosomes have to be distributed amongst four cells.

The abnormal larvae are of various types, some showing no trace of the Pluteus organization, some a small trace, such as a skeleton on one side. Others have no gut, or some of the pigment cells are missing (Fig. 161).

The distortion of development cannot be attributed to the

deficiency in the number of chromosomes. Each of the cells 31; x 2 3 n

4 = 72- chromosomes, for the larvae reared from them may have all their nuclei of the same size (containing the same number of chromosomes), and it is known (artiﬁcial parthenogtnesis and merogony) that a larva will develop quite well with only 11. Nor is it needful that all the parts should have the same number of chromosomes, for normal larvae may be experimentally produced, with 71 chromosomes in the nuclei of one region of the body and 2 n in the rest (see below). At the same time dispermy does not necessarily involve abnormality : in such eggs the two pairs of centres may remain apart and no quadripolar ﬁgure be produced: these develop into ordinary Plutei (Fig. 162 B). Conversely, quadripolar mitoses may be found in monosperm eggs, but these produce pathological blastulae. No explanation of the abnormality, therefore, is possible, so Boveri concludes, except that it is due to the irregularity in the distribution of the chromosomes. For normal development it is essential that each cell should receive not a deﬁnite number but a deﬁnite combination of chromosomes, a complete set of the chromosomes of the species. It is the lack of this complete combination in one or more cells that is the direct cause of the derangement of development.

Hence the chromosomes are not alike, but qualitatively diﬁerent. _

Their division is, however, never (except in reduction) a

of a quadripartite ovum may have 266 INTERNAL FACTORS IV. I

qualitative but always a quantitative process. The combination is handed on in its entirety to every cell of the body. The chromatin has in this sense an individuality, for however much the chromosomes may disappear while the nucleus is at rest, the set of qualitatively distinct units always re-emerges (though not necessarily in the same order) to be quantitatively divided between the two daughter—cells. We reach, therefore, a conception of the chromatin akin to Roux’s idea of the ‘ Reserve-idioplasson ’, the germ-plasm which is passed on entire to all the tissues of the body.

And now Boveri proceeds to develop a theory already suggested by Driesch of the part played by the chromatin in differentiation. The cytoplasm is heterogeneous, anisotropic; and these diﬁerenees in the cytoplasm are sufficient to account for the earliest diiferentiations, the pattern of segmentation, the determination of the embryonic axes and possibly some few others. But then the chromatic elements come into play—not the chromosomes, which are too large and too few to be regarded as determinants—but far smaller subdivisions of these. Under the inﬂuence of the cytoplasm, diiferent in the various cells, some of the units become patent and active, while others remain latent, and by the reaction of the former upon the cytoplasm new diﬁerentiations arise which will in their turn call forth into activity new nuclear elements.

Whatever value may be attached to these speculations, there can be no doubt of the importance of the experiments on which they are founded. Provisionally we have good ground for believing in the dissimilarity of the chromatic elements and of their necessity for differentiation. It only remains to be seen whether further investigations on other forms will bear out the interpre— tation which Boveri has placed upon his observations.

There is one more point worthy of notice. An ordinary sexually produced individual possesses in its nuclei 2n chromosomes, that is, two complete sets. In maturation one division of the chromosomes is transverse, and Boveri suggests that in this transverse division homologous units are separated from one another, or, to use the expression made familiar by current Mendelian theory, members of pairs of allelomorphs. c

Before bringing this section to a close a brief reference must IV. I INITIAL STRUCTURE OF THE GERM 267

be made to the relation established by Boveri, as a result of the examination of sea-urchin larvae (Ecﬁimw), between the number of chromosomes on the one hand and the size of the nucleus and the size of the cell on the other.

The number of chromosomes in a larva, or part of a larva, may be varied in the following ways.

1. In merogony the number is half the normal number, i.e.

'71. (The larva is termed by Boveri hemikaryotic arrhenokaryotie.)

2. In artiﬁcially parthenogenetic larvae it is also 12. (Hemikaryotic thelykaryotie.)

3. In ‘ monaster’ eggs the number is twice the normal, i. e. 4 oz. (Diplokai-yotic.)

This condition is brought about by shaking the eggs, and so preventing the division of the centrosome. The pronuelei unite and enter a resting stage. From this the chromosomes emerge in the full number and split, but since the centrosome remains single, there is no cell-division, and the chromosomes once more form a resting nucleus, but now in twice the normal number.

Later the centrosome divides and the egg develops normally.

4. If the eggs are kept for twenty-four hours in sea-water and then fertilized with sperm which has been weakened in dilute potash the male pronucleus lags behind its centre. The latter divides about the female pronucleus, and thus in the two-celled stage one blavstomere has a male and a female nucleus, and therefore 21; chromosomes (amphikaryotie), while the other has only a female (thelykaryotic). In this stage, or subsequently, the male unites with the female pronucleus, and so one-half, or onequarter, possibly only one—eighth, of the larva has nuclei with 2 72, the remainder nuclei with n chromosom_es.

5. It sometimes happens in dispermy that the two pairs of centrosomes remain apart without forming a quadripolar ﬁgure (Fig. 162). Such eggs, as we have seen, develop normally. Sooner or later the cells become divided into two sets, one with nuclei containing 2 71 chromosomes (amphikaryotie), derived from the female and one of the male pronuelei, the other having n chromosomes from the remaining male (arrhenokaryotic).

A superficial examination shows that the nuclei of hemikaryotic embryos and larvae are smaller but more numerous than in 268 INTERNAL FACTORS IV. I

normal (amphikaryotic) embryos or larvae of the same stage. In diplokaryotie larvae, on the other hand, they are larger and fewer; while in larvae in which one portion is amphikaryotic, the other hemikaryotic, the two regions are distinctly marked oﬂ by the diﬁerence in the size and number of their nuclei (Fig. 163).

The exact relation between the number of chromosomes originally entering into the composition of the nucleus and the dimensions of the nucleus and the cell has been ascertained by Boveri to be as follows :—

I. The surface-area of the nuclei is directly proportional to the number of chromosomes. The chromatin, it is worth noticing, is often placed beneath the nuclear membrane.

FIG. l62.—Dispermic eggs of Strongylocentrotus Iividus. A, The four centres have united to form a tetraster. B, The two pairs of centres have remained apart to form two independent spindles, one of which contains the female and one of the male nuclei, the other the remaining male nucleus. (After Boveri, 1905.)

II. The number of nuclei is inversely proportional to the number of chromosomes ,- but III. Cell-volume is inversely proportional to the number of

cells in larvae of the same stage developed from whole eggs _

of the same size; hence

IV. Cell-volume varies directly with the number of chromosomes and with the surface area of the nucleus. This Boveri terms the ﬁxed relation between nucleus and cytoplasm.

V. It follows from I and IV that nuclear-volume increases more rapidly thon cell-volume.

VI. Conversely, the law holds good of egg-fragments of different sizes but containing the same number of chromosomes.

Since, in the larvae developed from such fragments, the IV. I INITIAL STRUCTURE OF THE GERM 269

thickness of the cell-layers is the same, the surface may be taken as an index of the volume and is proportional to the original volume of the fragments. Since the number of chromosomes is the same, the cell-volumes should be the same, and the number of cells ought to be proportional to the surface. This was, as a matter of fact, found to,be true in the few instances examined.

This result is merely a restatement of Driesch’s rule, that in larvae developed from isolated blastomeres the surface of the larvae and the number of cells are both proportional to the germinal value.

It appears, therefore, that the moment at which segmentation shall end may be determined by the moment at which this constant relation between nucleus and cytoplasm is reached.

{.'S‘°7°i‘o\

J” .6.

6

FIG. 163.—— a, Gastrula reared from an eggof Slrougylocenfrotus lim'du.s-, in which the sperm nucleus had passed entirely into one of the first two blastomeres. Optical section of a stained preparation. (After Boveri, 1905.)

b, Ectoderm in the neighbourhood of the blastopore of J. dispcrmic gastrula of .S'trong_r/locentrotus lividus. (After Boveri, 1905.)

Driesch has, however, recently reported certain exceptions to his rule, amongst the mesenchyme cells in particular. These may be more or less numerous than they should be, may differ in size and in the size of their nuclei. It is possible, as suggested, that the division of the centrosome has been suppressed (monaster) with consequent reduplication of the chromosomes. Such an explanation may account for the variability in the size of the nuclei observed by Seeliger in his hybrids.

In the Alga /5:piro_qym Gerassimow has observed a close relation between the size of the nucleus and that of the cell; here, however, it is the nuclear volume which is directly proportional to the cell-surface. 270 INTERNAL FACTORS IV. 1

LITERATURE

K. BONNEVIE. Ueber Chromatindiminution bei Nematoden, Jen. Zeitschr. xxxvi, 1902. J

K. BONNEVIE. Abnormitiiten in der Furchung von Ascaris lumbricoides, Jen. Zeitschr. xxxvii, 1903.

M. Bovmu. Ueber Mitosen bei einaeiﬁiger Chromosomenbildung, Jen. Zeitschr. xxxvii, 1903.

T. BOVERI. Ueber die Befruchtungs- und Entwicklungsﬁihigkeit kernloser Seeigeleier und ﬁber die Moglichkeit ihrer Bastardirung, Arch. Ent. Mech. ii, 1896.

T. Bovmu. Zur Physiologie der Kern- und Zelltheilung, S’.-B. ph_1/s.med. Gee. W1212bz¢2'g, 1896.

T. BOVERI. Die Entwicklung von Ascaris megalocephala, mit besonderer Riicksicht auf die Kernverhiiltnisse, Festschr. Kuppfer. J ena, 1899.

T. Bovmu. Ueber mehrpolige Mitosen als Mittel zur Analyse des Zellkerns, Verh. phys.-med. Ges. Wzlnzburg, xxxv, 1903. '

T. Bovnm. Ueber da.s Verhalten des Protopla.sma.s bei monocentrischen Mitosen, S’.-B. phys.-med. Ges. Wdrzburg, 1903.

T. Bovmu. Ueber den Einﬂuss der Samenzelle auf die La.rvencha.raktere der Echiniden, Arch. Ent. Mach. xvi, 1903.

T. BOVERI. Noch ein Wort iiber Seeigelbastarde, Arch. Ent. Mech. xvii, 1904.

T. BOVERI. Protoplasmadifferenzierung als ausléisender Faktor fiir Kernverschiedenheit, S.—B. phys.-med. Ges. Wﬁrzbu;-g, 1904.

T. BOVERI. Ergebniase ﬁber die Konstitution der chromatischen Substanz des Zellkerns, Jena, 1904.

T. Bovmu. Ueber die Abhﬁngigkeit der Kerngrosse und Zellenzahl der Seeigel-Larven von der Chromosomenzahl der Ausgangszellen, Jena, 1905.

L. DONCASTER. Experiments in hybridization with especial reference to the effect of conditions on dominance, Phil. Trans. Roy. Soc. cxcvi, B. 1904.

H. Dnmscn. Ueber rein-miitterliche Charaktere an Bastardlai-ven von Echiniden, Arch. Ent. Mech. vii, 1898.

H. Dnmscn. Ueber Seeigelbastarde, Arch. Eut. Mech. xvi, 1903.

H. DRIESCH. Zur Cytologie parthenogenetischer Larven von Strongylocentrotus, Amh. Ent. Mech. xix, 1905.

A. FISCHEL. Ueber Bastardirungsversuche bei Echinodermen, Arch. Ent. Mech. xxii, 1906.

J. J. GERASSIMOW. Ueber den Einﬂuss des Kerns auf das Wa.chsthum der Zelle, Bull. Soc. Imp. Nat. Moscou, 1901.

J. J. GERASSIMOW. Die Abhﬁngigkeit der Grosse der Zelle von der Menge ihrer Kernmasse, Zeitschr. allg. Phys. i, 1902.

A. GIARDINA. Origine dell'oocite e delle cellule nutrici nel Dytiscus, Internat. Monatachr. Anat. u. Phys. xviii, 1901. ' IV. I INTERACTIONS OF THE PARTS 271

E. GODLEWSKI. Untersuchungen ﬁber die Bastardierung der Echiniden- und Crinoidenfamilie, Arch. Ent. Mech. xx, 1906.

M. KRAHELSKA. Sur le développement mérogoniquc des ueufs de Psammechinus, Bull. Internai. Acarl. Sci. Cmcovie, 1905.

J. LOEB. Ueber die Befruchtung von Seeigeleiern durch Seestem samen, P_ﬂﬂger's Arch. xcix, 1903. J. LOEB. Ueber die R.eaction‘des Seewassers und die Rollo der Hydro xylionen bei der Befruchtung der Seeigeleier, Pﬂiiger-’s Arch. xcix, 1903. J. LOEB. Weitere Versuche ﬁber heterogene Hybridisation bei Echino dermen, Ifﬂﬁger-‘s Arch. civ, 1904. A. P. MATHEWS. The so-called cross-fertilization of Asterias by

Ar-bacia, Amer. Journ. Phys. vi, 1901-2. T. H. MORGAN. The fertilization of non-nucleated fragments of

Echinoderm eggs, Arch. Eut. Mech. ii, 1896.

W. ROUX. Ueber die Bedeutung der Kerntheilungsﬁguren, Leipzig, 1883, Ges. Abh. 17. Leipzig, 1895. O. SEELIGER. Giebt es gesehlechtlich erzeugte Organismen ohne miitterliche Eigenschaften ? Arch. Em. Mech. i, 1895.

O. SEELIGER. Bemerkungen ﬁber Baatardlarven der Seeigel, Arch.

Ent. Mech. iii, 1896.

E. TEICHMANN. Ueber Furchung befruchteter Seeigeleier ohne Beteiligung des Spermakerns, Jen. Zeitschr. xxxvii, 1903.

H. M. VERNON. The relations between the hybrid and parent forms

of Echinoid larvae, Phil. Trans. Roy. Soc. cxc, B. 1898. H. M. VERNON. Cross-fertilization among Echinoids, Arch. Eut.

Mech. ix, 1900.

2. ON THE ACTIONS OF THE PARTS OF THE DEVELOPING ORGANISM ON ONE ANOTHER

Up to the present we have discussed only the original structure of the fertilized ovum as an internal factor in diﬁerentiation. We have now to consider to what extent the development of each part may be due to inﬂuences exerted upon it by other portions of the embryo.

One of the ﬁrst suggestions of such an interaction was put forward by Oscar Hertwig. ‘We recognize’, he says in his Zez't- mul Streiyfrage, ‘apart from the nature of the egg-substance itself, the conditions of development, ﬁrst in those perpetually changing mutual relations in which the cells of an organism are placed to one another, and secondly in the influences exerted upon the organism by the external world ’ ; and again, ‘physiologically expressed, the dissimilar diiferentiation of cells consists in the reaction of organic substance to dissimilar stimuli,’ and 272 INTERNAL FACTORS IV. 2

he proceeds to illustrate his meaning by referring to the influence of light in determining whether the sexual organs shall be formed on the upper or under side of a fern prothallus, to the dependence of the fate of indiﬁerent buds of plants upon external conditions, to the formation of stems on the upper, roots on the lower side of a horizontal stolon of Amfennularia, as demonstrated by Loeb, and to certain alterations in the character of epithelia and other tissues which are brought about by pathological conditions.

For a more detailed elaboration of this very fruitful idea we are indebted to Herbst. Herbst classiﬁes organic reactions to stimuli as either directive or formative. The former are of two kinds, tactic when the response is some locomotion of a freer body, tropic when the movement exhibited in response is a movement of growth. The reactions may be further classified according to the nature of the stimulus, gravity, light, heat, electricity, chemical substances, currents of water, the contact of a solid surface. Thus we have positive and negative heliotropism, galvanotaxis, geotropism, galvanotropism, thigmotaxis, and so on, and many examples of all of these are cited. These are mostly familiar facts, but some, for their special interest, may be quoted here. Thus the hydroid Sertularellw in one-sided illumination produces stolons instead of hydranths, the tubes of the Polychaet Sergmla grow upwards towards the light (or against gravity), an inverted stock of Sertularia produces a polyp sympodium at the upper—originally lower——end, the stems of Aulemuluria are negatively, the stolons positively geotropic, spermatozoa swim to solid bodies, and stolons of Hydroids attach themselves to some foreign object.

Speaking generally, a speciﬁc reaction depends not merely on the nature of the stimulus, but on the structure of the reacting body itself. Further, the degree of sensitiveness to the stimulus is not constant. It varies at different periods of life, under the influence of external conditions such as temperature, and with the strength of an already existing stimulus (Weber’s Law).

Now many of the events of ontogeny, as we have seen, resolve themselves into movements of the parts of the organismmovements of free parts, locomotion, and growth movementsIV.2 INTERACTIONS OF THE PARTS 273

and the suggestion is made by Herbst that these movements are to be interpreted as so many responses to stimuli, which are then not merely directive but formative, the stimuli being either external or exerted by other parts of the body. Thus the spreading of the blastoderm over the yolk may be a case of oxygenotaxis, the movement of cells of the Frog’s blastula towards one another described by Rouxl as Cytotropismus may be due to a mutual chemotactie stimulus, the migration of cells to the surface of the Arthropod ovum is perhaps oxygenotactic, the vitellophags are ehemotactic, the application of mesoderm cells to the outgrowing axis cylinders of nerve ﬁbres, of connective tissue and muscle-cells to blood-vessels, of the choroidal cells to the optic cup (they are absent ventrally when the choroid ﬁssure fails to close), the reunion of separated blastomeres in Triclads and others, may all be tactic phenomena of a positive kind, while the dispersal of the elements of a complex may be negative. Similarly, the outgrowth and anastomoses of nerves, glands, ducts, the concrescence of layers may be tropisms of various sorts.

The suggested stimuli in some cases proceed from the external environment, in others they are exerted by one part on another. The former have already been discussed, but it may be pointed out again that the effect called forth depends both on the stimulus and on the reacting organism or organ. Neither in these cases nor in any others is the production of speciﬁc form due to the action of an external agency upon a homogeneous material. For even in those instances where the fate of some apparently indifferent tissue seems to be decided by the action of such an agency-—as in the production of stems from whichever side of an Anlemmlaria stock happens to be uppermost (Loeb)—it is evident that all that the external agent can do is to decide between a limited number of deﬁnite alternatives in respect of each of which the parts of the organism are equipotential. Responses to stimuli of the second kind possibly play an important part in ontogeny. Unfortunately, the experimental evidence for this is at present somewhat meagre.

In the ﬁsh Funrlulus the yolk-sac is deeply pigmented, the pigment-cells being especially closely aggregated round the

‘ See above, Ch. II. 1, p. 55.

unumrson T 274 INTERNAL FACTORS ' IV. 2

blood-vessels. When the creature is deprived of oxygen the pigment disappears, and it is supposed that under normal conditions the pigment cells are chemotactically attracted by the oxygen in the blood (Loeb).

Two other cases come from Echinoid larvae. Driesch has been able to displace, by shaking, the two groups of primary mesenchyme cells, those destined to secrete the skeletal spicules of the

FIG. 164.-Diagrams of Driesch's experiment into the responsiveness to stimuli of mesenchyme cells in Echinus. A, Normal arrangement of mesenchyme cells. B, The cells deranged by shaking, returning in C to their original position. (From Korschelt and Hcider, after Driesch.)

larva. The cells, displaced towards the animal pole, return to their original position after six hours, and subsequent development is normal 164). Driesch has attributed this movement of the cells to a stimulus exerted by the endoderm.

Again Herbst has found that when the larvae of sea—urchins are placed in solutions of lithium salts or in sea-water deprived of the sulph-ion, or devoid of the carbon—ion, the skeleton of the IV. 2 INTERACTIONS OF THE PARTS 275

Pluteus is either distorted, in the ﬁrst two cases, or absent, as in the last. The distortion takes the form of a multiplication of the triradiate spicules, and the arms are correspondingly multiplied. In water devoid of CO2 there are no spicules and no arms} Herbst has therefore urged that normally the outgrowth of the ciliated ring into the arms is due to a stimulus—thigmotropic, perhaps——exe1-ted by the tip of the spicule. The converse of this is seen later on when the arms diminish in length as the calcium carbonate of the Pluteus skeleton is made use of by the developing urchin.

The development of the lens of the vertebrate eye has also been asserted to be due to a contact_stimulus exerted by the optic cup upon the overlying ectoderm. Spemann was the ﬁrst to show that when the medullary plate of the Frog (Rana fusca) was injured in front of the optic vesicle, the optic cup (if developed at all) may or may not come in contact with the ectoderm, the latter being, at this point, uninjured. In the ﬁrst case a lens is formed, in the second not. Similar experiments have been carried out by Lewis on other species of Frog and on Amblystoma, with similar results. Lewis has also shown that a lens will be formed from any patch of ectoderm taken from some other part of the body and grafted over the optic cup. Schaper’s evidence in support of this conclusion is doubtful. This author removed the whole of the nervous system from a 4 mm. tadpole by a horizontal cut with the exception alone of the downwardly growing fore-brain and optic vesicles. The wound healed and development continued. The optic cup was solid, being filled with a mass of degenerating cells. A lens was formed opposite the upper edge of the optic cup. It did not, however, separate from the ectoderm, but diﬁerentiated in silu, with development of lens ﬁbres. Hence Schaper argues that the formation of the lens is a process of self-differentiation. The contact of the optic cup is, however, as he admits, in this case not excluded.

A more serious objection is raised by Miss King, who has found (in the Frog) that even when, owing to a prior injury, the optic cup is far away from the skin, or in some cases absent altogether,

‘ See above, Ch. III. 8, Figs. 73, 74. T 2 276 INTERNAL FACTORS IV. 2

the lens may still be formed. It may also be mentioned that Mencl has described a double-headed monster of Salmo salar in which one head has two well-formed lenses, though the other parts of the eye are absent. The lenses, however, lie quite close to the fore-brain, which may have exerted the stimulus, supposing a stimulus to be necessary.

It will be seen that the evidence is extremely contradictory, and forbids any deﬁnite conclusion.‘

It is curious that in tadpoles grown in certain solutions (sodium chloride, bromide, and nitrate) the optic cup is often at some distance from the ectoderm, and that in these cases the lens is absent, though it may happen that the upper edge of the optic cup is sharply bent down in front of the aperture, as though to form a lens from the edge of the iris, as occurs in the regeneration of the lens, after extirpation, in the Newt.”

Spemann and Lewis have also found that in the absence of contact between the optic cup and the ectoderm the cornea is not developed, that is to say, the overlying ectoderm does not ‘clear’ (lose its pigment), it does not thin out, and Descemet’s membrane is not formed. The more extensive series of experiments is due to Lewis, who turned back a flap of skin and wholly or partially removed the optic vesicle, the skin being then returned to its place. In the first case there was no eye, no lens, no cornea, though on the other side of the body all were present. In the second case the lens and cornea were both formed, provided that enough was left of the vesicle for the optic cup to come into contact with the ectoderm. As a control experiment the ﬂap of skin was merely removed and replaced; the development of the eye was normal. If at a later stage, when corneal changes had already begun, both optic cup and lens were taken out, the cornea was not completed. The pigment indeed disappeared, but the ectoderm did not diminish in thickness and Descemet’s membrane was not formed.

Removal of the lens alone did not interfere with the production

1 Stockard has recently shown that when the egg of Fzmdulus is placed in MgCl, the two optic cu s wholly or partly fuse while a single or double median lens is deve oped (Arch. Ent. Mech. xxiii, 1907). This would seem to support the view advanced by Lewis.

2 See above, Ch. III. 8, p. 134.

C IV.2 INTERACTIONS OF THE PARTS 277

of the cornea provided that the optic cup could touch the ectoderm, and if this same condition is fulﬁlled, the cornea will be differentiated in the ectoderm that grows in after the layer that originally lay over the eye had been destroyed, but if the cup and lens are taken away no new cornea is formed even after four weeks. The size of the cornea is in proportion to the area of contact. Finally, if, at a later stage still, the optic cup and lens are taken away, the already formed cornea degenerates. Should the results of these extremely interesting investigations be conﬁrmed, the development of some parts at least of the vertebrate eye would be processes of ‘ dependent diiferentiation ’ 1, dependent, that is, on causes outside themselves, on stimuli exerted by other parts, though not, of course, wholly dependent, since the structure of the reacting, aswell as that of the stimulating, organ is contributory to the quality of the eifect. How far such an action of the parts upon one another is of general occurrence as a factor in differentiation future research alone will show. It seems probable, however, that it will in any case be found to be limited to the ﬁrst moments in the formation of the organs 2, for we know from other evidence that as development proceeds the system which was ‘ equipotential ’, to use Driesch’s phrase, becomes ‘ inequipotential ’, that the parts lose their totipotence, that they gain independence and the power of self-differentiation, that the value of the correlations between them diminishes. This is what Minot has called the law of genetic restriction. But in early stages there are correlations between the‘ organs, the differentiation of one does depend upon that of another, and it is this mutual stimulation of the parts that ﬁgures so prominently in the theory of development worked out by Driesch. This theory we shall have to examine in the following chapter.

' 1 Although the expression ‘dependent differentiation‘ is due to Roux, and although Roux expressed the opinion that certain developmental processes—the beginning of parthenogenetic development, the determination of the position of the grey crescent and ﬁrst furrow by the entrance-point of the spermatozoon, the ‘post-generation’ of the dead blastomere by the living one in the Frog's egg, for instance—-were of the nature of responses to stimuli, still the role assigned by the ‘Mosa.ikTheorie’ to the inﬂuences of the parts upon one another in differentiation was subordinate, and restricted to the later stages of development.

2 Except, presumably, in cases of ‘composition’ where elements of different origin unite in the formation of a. single organ. 278 INTERNAL FACTORS IV. 2

LITERATURE

H. Dnmscn. Die taktische Reizbarkeit der Mesenchymzellen von Echinus microtuberculatus, Arch. Ent. Mech. iii, 1896.

C. HERBST. Ueber die Bedeutung der Reizphysiologie fiir die kausale Auffassung von Vorgfingen in der tierischen Ontogenese, Biol. Centralbl. xiv, xv, 1894, 1895.

C. HERBST. Experimentelle Untersuchungen iiber den Einﬂuss der

veriindert-en chemischen Zusammensetzung des umgebenden Mediums

ztuf die Entwickelung der Thiere: II. Mitt. Zool. Stat. Neapel, xi, 1895.

C. HERBST. Ueber die zur Entwickelung der Seeigellarven nothwendigen anorganischen Stoﬁ‘e, ihre Rolle und ihre Vertretbarkeit. I. Die zur Entwiclrelung nothwendigen anorganischen Stoffe, Arch. Ent. Mach. v, 1897. 11. Die Rolle der nothwendigen anorganischen Stoﬁ'e, ibid. xvii, 1904.

0. HERTWIG. Zeit- und Streitfragen der Biologie, Jena, 1894.

H. D. KING. Experimental studies on the eye of the Frog embryo, Arch. Ent. Meek. xix, 1905.

W. H. LEWIS. Experiinental studies on the development of the eye in Amphibia. I. On the origin of the lens, Amer. Journ. Anal. iii, 1904.

W. H. LEWIS. Experimental studies on the development of the eye in Amphibia. II. On the cornea, Journ. Exp. Zool. ii, 1905.

J. LOEB. Untersuchungen zur physiologischen Morphologie der Thiere: II. Wﬁrzburg, 1892.

J. LOEB. A contribution to the physiology of coloration in a.ni1na.ls, Journ. Mmph. viii, 1893.

E. MENCL. Ein Fall von beiderseitigen Augenlinsenbildung wiihrend der Abwesenheit von Augenblasen, Arch. Ent. Mech. xvi, 1903.

W. Roux. Ueber den Antheil von ‘Ausltssungcn ’ an der individuellen Entwicklung, Arch. Ent. Mech. iv, 1897.

A. SCHAPER. Ueber einige Fiille atypischer Linsenentwicklung unter abnormen Bedingungen, Auat. Am. xxiv, 1904.

H. SPEMANN. Ueber Linsenbildung bei defekter Augcnblase, Anat.

Anz. xxiii, 1903. CHAPTER V DRIESCH’S THEORIES OF DEVELOPMENT

GENERAL REFLECTIONS AND CONCLUSIONS

To the inquirer into the causes of development the central diﬁiculty must always be the problem of diﬁerentiation. Growth and division of the nucleus and the cell, processes which always accompany differentiation, are, as we have seen, side issues ; but the increase of structure, the production of form out of the relatively formless germ, and the gradual passing of this into a new individual which is like the parents that gave it birth, this is the marvel which has always excited the wonder of the observer, and demands all his wit to understand and to explain.

Experimental investigation, as far as experiment has at present gone, has shown, ﬁrst, that a certain complexity of the physical and chemical environment is a necessary condition of normal development ; that complexity may, it is true, vary within certain limits, but those limits can only be transgressed under pain of abnormality or death. In the second place it has been demonstrated that the initial structure of the germ, and the mutual interactions of its parts as they develop are both indispensable internal factors. It now only remains for us to discuss the value of those theories which are not only attempts to explain, to give the most general account of, the phenomena in causal terms, but also serve to provide a light to guide the investigations of the future. One such hypothesis we have already examined in some detail, Roux’s hypothesis of selfdiffercntiation. According to this belief not only is the development of each part determined by causes which reside in itself alone, but the parts—or rather their determinant representatives, which are all ea: /1._1/pot/Eesi necessarily present ab 2'mYz'o in the undeveloped germ—are located in the nucleus. The qualitative division of the nucleus sunders these units from one another, which then determine the characters assumed by the cytoplasm, and so the whole process happens.

As we have seen, the hypothesis, in this its original form, is 280 DRIESCH’S THEORIES OF DEVELOPMENT V

untenable. Quite apart from the fact that the complex architecture of the nucleus still demands an explanation—an explanation of the same kind, perhaps, which would at once involve us in an inﬁnite regress—the facts which experiment has brought out show conclusively that nuclear division is never a qualitative process. In the other direction the hypothesis errs in attributing a homogeneity, an isotropy to the cytoplasm, for the same experiments have proved the existence in the ovum of deﬁnite substances, necessarily concerned in the production of the primary organs of the embryo.

There is, however, no evidence to show that--as imagined in Weismann’s, and to a certain extent in Roux’s hypothesis—there

is a separate morphological unit for each separately inheritable‘

character of the species; such an idea would indeed seem to be precluded by the ease with which, in some cases at least, the germ may be divided into parts, each of which is endowed with the potentialities of the whole.

And yet development is speciﬁc. How, then, is this mechanism of inheritance to be conceived of? It is to Hans Driesch that we are indebted for an exhaustive attempt to think out the whole problem. In his Aualytiec/Le T/teorie tier organise/len Entwic/clzmg Driesch starts with the facts with which we are already acquainted, the similarity of the nuclei, the dissimilarity of the cytoplasm in the several regions of the developing germ.‘

The arrangement of these dissimilar substances determines ﬁrst of all an axis in the egg, an axis with unlike poles ; around this axis the cytoplasm is radially symmetrical or isotropic, but in the direction of the axis it is not ; or, as Boveri puts it, there is a ‘stratiﬁcation ’ of the substances of the egg at right angles to the axis, the concentration of the animal substance decreasing towards the vegetative pole, that of the vegetative substance in the contrary direction. There are cases (Coelenterates, Sponges, Ctenophora) in which this radial is the only symmetry; but in other types (Bilatcralia) a third point may be established by the disposition of some special substance (the grey crescent in the Frog"s egg or the yellow pigment in the

1 It may be mentioned that in 1887 Platner had already denied the existence in development of any qualitative nuclear division. V GENERAL REFLECTIONS AND CONCLUSIONS 281

Ascidian Cyntﬁia, for example), or by the arrangement of the blastomeres (as by the large posterior cell of Annelids and Mollusca), and this point, together with the axis, determines a plane about which the ovum is bilaterally symmetrical. The axis and the plane of symmetry of the egg are deﬁnitely related to the axis and symmetry of the embryo, the substances to its primary organs.

Further, in very many, though not in all, instances the parts of the ovum—-blastomeres or egg-fragments——are totipotent ; and the same is true of the parts of elementary organs like the archenteron of Echinoderms or the optic vesicle of Amphibia. The totipotence is, however, sooner or later lost, and this limita

FIG. 165.——Diagram to illustrate Driesch’s conception of the minute structure of the (Echinid) ovum. It is supposed to be composed of particles all similarly polarized and oriented to the whole. In A onemeridional—half is shown. In B this has become spherical and the parts have been disturbed. In 0 they have regained the original orientation. (From Korschelt and Heider.)

tion is apparently due to the way in which the substances are distributed in the ovum, an explanation which seems to be accepted by Driesch for most cases. But in accounting for the phenomena in the Echinoderm egg, the form with which he himself has chieﬂy experimented, he urges a different hypothesis. Here he conceives of the egg as composed of like particles, each of which is polarized and oriented in the same manner as the egg itself (Fig. 165), and consequently the only limitation to totipotence is due to size ; any isolated part that is not too small can develop into a whole as soon as its polarized particles have reassumed a similar orientation. Again, it is stated that the blastomeres can be disarranged to any extent without interfering 282 DRIESCH’S THEORIES or “DEVELOPMENT v

with the normal development of the larva; they are all equivalent, without limit, at least until the ectoderm and endoderm have been diﬁerentiated ; any part can contribute to the formation of any organ: ‘ jeder Theil kann jedes.’

This conception, of the absolute equipotentiality of the parts, as we have already had occasion to remark, is erroneous; but for Driesch it is of the ﬁrst importance, for it dominates, as we shall see, all his theoretical speculations.

The position of the embryonic axes and primary org-ans being thus determined in the whole egg (or its isolated parts), it is still left to inquire into the causes which decide the destinies of the remainder. Driesch’s answer to this question is twofold: there

FIG. 166.—Diagram to illustrate the possible part played by stimuli (‘inductions’) in ontogeny, and by ‘position '. A, B, and C’ are three larval organs (ectoderm, endoderm, stomodaeum). C may exert a stimulus on that part of B nearest it; this part, reacting to the stimulus, becomes (3. Were the position of C’ altered the position of B in the cquipotential system B might be altered too, so that the fate of any part of B would be a function of its position relative to 0. So, under the inﬂuence of B, part of A may become a. (After Driesch, 1894.)

are two possible factors, one is ‘ position ’, the other ‘induction ’. In the case of the ﬁrst, the destiny of a part is imagined to be determined by its distance from the system of points already established, ‘ its fate,’ so runs the famous formula, ‘ is a function of its position in the whole.’ It would, however, be absurd to suppose that the behaviour of any one of a number of precisely similar bodies could depend upon its mere geometrical position. The points already diifcrentiated——the animal pole, for instance— must be supposed to exert an inﬂuence with a force which is some function of the distance upon the parts which are at present equivalent, and so to excite their differentiation 1 Fig. 166). V GENERAL REFLECTIONS AND CONCLUSIONS 283

Properly speaking, therefore, this factor of ‘ position ’ belongs to the second category of ‘ induction ’.

An ‘induction’ is simply an effect produced upon the parts that are developing by other parts, or possibly by some factor in the external environment. These inductions are, according to Driesch, of the nature of those events which are brought about by the addition of a single antecedent condition, an ‘occasion’, to an assemblage of antecedent conditions, as a spark ﬁres a rocket or the movement of a lever sets a piece of machinery in operation; or, in the language of physiology, the causes of these inductions are stimuli, the eﬁects, reactions or responses,

50

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2

FIG. 167.—Diagram to illustrate Driesch's hypothesis of the part played by the nucleus in development. A is the cytoplasm, '1‘ its totipotent nucleus, ml, ac, are stimuli.

A is sensitive to a:, and becomes A,, T is sensitive to A, and is set in activity (Ta). A‘ being sensitive to Ta becomes A1, and A, is sensitive to .L‘._., and so on. (After Driesch, 1894.)

the quality and the quantity of which are determined not only by the stimulus but also by the nature of the reacting body itself. The analysis of these ‘ inductions ‘is not, however, yet complete. Occurrences of this kind imply the stimulus, the reception of the stimulus and the response. The first of these is some other organ or some external agent; the second and third are functions of the organ which is to be produced; and Driesch imagines that while it is the cytoplasm which is receptive of the stimulus, it is the nucleus which makes the response. Each totipotent nucleus is supposed to contain ferment-like bodies; under the inﬂuence of. some stimulus received through the cytoplasm some 284 DRIESCH’S THEORIES OF DEVELOPMENT V

one of these is called into action; this works on and alters the cytoplasm; this then becomes receptive to a new and dilferent stimulus, and so on, as shown in the accompanying diagram (Fig. 167). The cytoplasm thus becomes perpetually altered, while the nucleus retains the entirety of the potentialities of the organism, a supposition which is held in reserve to account for the phenomena. of budding and regeneration.

The progressive limitation of potentialities, therefore, which we observe in ontogeny is a limitation of sensibilities to stimuli, equality of potentialities an equality of such sensibilities. In an ‘ equipotential system’ (the blastula or the archenteron of Ec/linus) all (meridional) parts are equally susceptible, and what stimulus each will as a matter of fact receive is a function of its position in the whole, that is to say of its distance from the centres of force situate in the dissimilar points ﬁxed ab initio in the system of symmetry of the egg (Fig. 166).

Development then proceeds from the comparatively simple organization given in the structure of the fertilized ovum by the creation of ever-increasing complexity by the action and reaction of the parts on one another. Each ontogenetic eifect produced becomes in turn the cause of further eifects, the possibility of fresh speciﬁc action, for it becomes the seat of a new speciﬁc stimulus and response, and so on until the complexity of the ‘ ultimate organs ’ of the adult is achieved.

In order, however, that differentiation may be normal it is clear that these stimuli and these responses must be accurately coordinated: the right stimulus must be ready at the right time and at the right place for the right organ to respond to. This indispensable temporal and spatial co-ordination is the ‘ Causal Harmony ’ of development; and it is given by the initial structure of the egg and the constitution of the external environment. ’

This is, however, not the only kind of harmony involved in development. Potentialities become restricted as differentiation progresses, and the development of the primary, secondary, tertiary and subsequent systems of organs successively produced is a process of ‘ self-diiferentiation ’, depending on factors residingin each system itself. Nevertheless these independent systems V GENERAL REFLECTIONS AND CONCLUSIONS 285

frequently unite later on to form complex organs, and the necessary co-ordination of these is a ‘ harmony of composition ’.

Such, in brief, is the idea worked out in the Analytiec/ze T/leorie. On this hypothesis development is an epigenesis ; it involves a super-additio partium, an increase of structure, a creation of fresh form out of the simple cytoplasmic organization of the egg, and the causes which operate in this process of morphogeny are just that initial structure and the stimuli which the parts respond to, stimuli which may proceed from the parts themselves or from the world outside.

These responses are primarily physico-chemical and only secondarily structural, and they happen in accordance with a pre-established harmony. Only in this sense can development be described as evolution; but this is not the evolution of the preformationists of the eighteenth century, nor even that of the school of Roux and Weismann, less gross, more subtle, but still morphological; rather it is the realization of a form which is physically and chemically predetermined, not structurally preformed, in the simple organization of the germ.

The hypothesis is worthy of the attention of every serious embryologist. It is scientiﬁc in the strict sense of the word, for in employing Herbst’s suggestion to bring the events of ontogeny into the category of physiological responses to stimuli it gives meaning and precision to Hertwig’s somewhat vague idea of the ‘ mutual relations of the parts’, and makes thereby a genuine effort to think the particular under the universal, to bring the facts of embryology under wide general laws of causation.

Its chief Weakness is at present lack of evidence; the development of every organ requires to be examined by the touchstone of experiment before the theory can rise beyond the rank of a working hypothesis. That, however, is the common lot of all working hypotheses, and this one has certainly some counterbalancing advantages. Its conception of the réle of the nucleus has been to some extent borne out by Boveri’s recent researches and adopted by their author, a conception which, as we have observed before, is not irreconcilable with the facts of budding and re2'eneration. Secondly, only the very simplest original 286 DRIESCH’S THEORIES OF DEVELOPMENT V

organization of the germ is demanded. This organization is not, however, to be looked for in the similar orientation of similarly polarized particles. On the contrary, it is to be found in the existence of deﬁnite cytoplasmic organ-‘forming substances whose arrangement must be simple enough to allow of the divisibility of the whole into totipotent parts, complex enough to account for that limitation of the potentialities of these parts which, sooner or later, inevitably ensues. And lastly, when modiﬁed in this important respect, Driesch’s views are no longer hopelessly at variance with Roux’s position, provided that we discard the vicious fallacy of qualitative nuclear and cell—division, and substitute for the idea of complete morphological preformae tion at origine the successive preformation by the action of the parts on one another in the organs of each stage of the structures that are to be developed out of them in the next. To use Roux’s own terms, development, in its early stages at least, is both a process of ‘ self- ’ and of ‘ dependent ’ differentiation ; or, as Nag-eli expressed it, it is by the continued combination and permutation of a few original elements that inheritance is brought about.

Driesch, however, is not merely a scientiﬁc thinker who is fully alive to the prime importance of pushing the causal analysis to its extreme limit. He is also a philosopher; and as a philosopher he realizes that when science has said all it has to say the account may still need to be completed from a new and distinct point of view. In the case of living organisms this new standpoint is the teleological. The harmony—causal harmony and harmony of composition—deduced from develop: ment, the functional harmony exhibited by the organs of the adult, all appear to be directed to an end, which is the reproduction or the preservation of the speciﬁc form, and it is only when this end is understood that the mere reference to beginnings which a knowledge of the mechanism gives acquires a genuine signiﬁcance. Purposiveness, in a word, is a characteristic of all organic functions and cannot be ignored.

This principle is borrowed avowedly from Kant’s Kritik of Me Teleological Judgement. Like the scientists of to-day, Kant lays it down as a rule that the mechanical method, by which natural phenomena are brought under general laws of causation and V GENERAL REFLECTIONS AND CONCLUSIONS 287

so explained, and without which ‘ there can be no proper knowledge of Nature at all,’ should in all cases be pushed as far as it will go,‘ for this is the principle of the determinant judgement.“

There are cases, however, in which this alone does not sufﬁcef‘ The possibility of the growth and nutrition, above all of the reproduction and regeneration of organisms is only fully intelligible to human reason through another quite distinct kind of causality, their purposiveness. Organisms are not mere machines, for those have merely moving power. Organisms possess in themselves formative power of a self-propagating kind which they communicate to their materials. They are, in fact, natural purposes, both cause and eifect of themselves, in which the parts so combine that they are reciprocally both end and means, existing not only by means of one another but for the sake of one another and the whole. The whole is thus an end which determines the process, a ﬁnal cause which ‘ brings together the required matter, modiﬁes it, forms it, and puts it in its appropriate

' §§ 70, 78, 79, 80. _

’ It should be mentioned, perhaps, that Kant employs the terms ‘determinant’ and ‘reﬂective’ judgement in two senses. In the Introduction to the Kritik of Judgement judgement is deﬁned as that faculty which thinks the particular as contained under the universal, and is stated to be of two kinds. It is ‘determinant’ when the universal is given (as in the mathematical sciences). When, however, only the particular is given, for which the universal has to be found (as in the inductive sciences), it is ‘ reﬂective ’, and Kant insists that the ‘ reﬂective’ judgement requires a principle which it cannot borrow from experience, and this principle is, in brief, the ultimate intelligibility of Nature by us. Only so far as this holds good can we hope to gain a knowledge of general, empirical laws of causation.

The ‘reﬂective’ judgement is again of two kinds, for which Kant, however unfortunately, employs the same two terms, ‘ determinant‘ and ‘reﬂective’. The duty of the former is, assuming that all phenomena are explicable in mechanical terms (‘ causal’ terms in the usual sense of the word), to push the analysis of these efﬁcient causes (nexus e;ﬁ'eclit'us) to its extreme limit. The latter, on the other hand, is concerned with a kind of causality after the analogy of our own causality according to purposes, in order that it may have before it a rule according to which certain products of nature, namely organisms, must be investigated. The whole argument of the Kritilc is directed to proving that these two uses of the judgement are mutually supplementary and both indispensable, though in the last resort the ﬁrst has to be subordinated to the second.

In the text the terms ‘ determinant’ and ‘reﬂective’ are used in the second sense. The ‘determinant judgement‘ of the deductive sciences

does not of course come into this discussion at all. 3 §§ 64-66.‘ 288 DRIESCH’S THEORIES OF DEVELOPMENT V

‘ place ’.1 Such purposiveness is internal, for the organism is at once its own cause and an end to itself, not merely a means to other ends, like a machine whose purposiveness is relative and whose cause is external.

Such is the principle of the teleological judgement. It is a ‘heuristic principle’ 2 rightly brought to hear, at least problematically, upon the investigation of organic nature, ‘ by a distant ‘ analogy with our own causality according to purposes generally,’ 3 and indispensable to us, as anatomists, ‘ as a guiding thread if we ‘ wish to learn how to cognize the constitution of organisms ‘ without aspiring to an investigation into their first origin.’ 4

For ‘ we cannot adequately cognize, much less explain, ‘ organized beings and their internal possibility, according to ‘ mere mechanical principles of nature ’, and it is therefore absurd ‘ to hope that another Newton will arise in the future who shall ‘make comprehensible by us the production of a blade of grass ‘ according to natural laws which no design has ordered ’.5

Could our cognitive faculties rest content in this maxim of the reﬂective judgement it would be impossible for them to conceive of the production of these things in any other fashion than by attributing them to a cause working by design, to a Being which would be ‘productive in a way analogous to the ‘ causality of an intelligence ’.°

Natural science, however, needs not merely reﬂective but determinant principles which alone can inform us of the possibility of ﬁnding the ultimate explanation of the world of organisms in a causal combination for which an Understanding is not explicitly assumed, since the principle of purposes ‘ does ‘ not make the mode of origination ’ of organic beings ‘ any more ‘comprehensible ’.7 And then, in a passage“ remarkable for its prophetic insight, Kant proceeds to show how this might be. ‘ The agreement of so many genera of animals in a common ‘ scheme . . . allows a ray of hope, however faint, to penetrate into ‘ our minds, that here something may be accomplished by the aid ‘ of the principle of the mechanism of nature (without which ‘there can be no natural science in general). This analogy of

1;8s6. ‘@978. 3§e5. 4§72. v§7.5. °§-15. V GENERAL REFLECTIONS AND CONCLUSIONS 289

‘forms, he says, ‘which with all their differences seem ‘ to have been produced according to a common original type, ‘strengthens our suspicions of an actual relationship between

.‘ them in their production from a common parent, through the

‘ gradual approximation of one animal-genus to another—from ‘ those in which the principle of purposes seems to be best authen‘ ticated, that is from man, down to the polype, and again from ‘this down to mosses and lichens, and ﬁnally to the lowest ‘ stage of nature noticeable by us, namely, to crude matter. ‘And so the whole Technic of nature, which is so incompre‘hensible to us in organized beings that we believe ourselves ‘ compelled to think a different principle for it, seems to be ‘derived from matter and its powers according to mechanical ‘ laws (like those by which it works in the formation of crystals).’

A purposiveness, however, must be attributed even to the crude matter, otherwise it would not be possible to think the purposive form of animals and plants.

Although there are doubtless in the Kritil: many obscurities and apparent inconsistencies, to which we cannot allude now, the general meaning of Kant’s reﬂections upon organisms is perfectly clear. He who would ‘complete the perfect round’ of his knowledge must think not only in beginnings but in ends. The end in the case of a living being is plain-—it is the maintenance and reproduction of its form; the end in the case of the cosmic process, though perhaps not so plain, is to be sought in the ethical, or, in Kantian phraseology, the ‘practical’ concept of the freedom of the moral consciousness of man.

Such a position is quite intelligible, philosophically; and it can only be a matter of surprise that Driesch has not been able to abide by it. In his later writings he has indeed ‘executed acomplete change of front and repudiated the philosophical doctrine laid down in the earlier treatise; and the principal reason for this voZte- ace is that there are cases in which the localization of

ontogenetic effects cannot be explained by any theory of formative stimuli. In the theory we have already considered the causal harmony which secures the due co-ordination in space and

in time of the stimuli and responses into which the process of JINKIHIOH U 290 DRIESCH’S THEORIES OF -DEVELOPMENT V

differentiation is resolved is held to result from the initial structure of the germ and to be maintained by the constitution of the environment. Now, however, it is urged that no materia factor can possibly account either for this harmony or for the secondary harmony of composition or the functional harmony seen in the activities of the adult. When, for example, the gastrula of a sea-urchin is transversely divided into two, each develops into a diminished whole larva in which the gut becomes divided into the characteristic three regions, and all the other organs are formed in correct proportion. For each of these acts in the whole uninjured larva an explanation may conceivably be given in terms of stimuli or forces emanating from the originally distinct parts of the egg and producing effects which vary with the distance upon other parts, as suggested before. A mechanism may be thought of which, when set in motion, will achieve a certain end in accordance with its own pre-established harmony, but a mechanism which can be subdivided ad libitrm, or almost arl libitzcm, and the parts of which will still achieve the same end, will still behave as wholes with their parts co-ordinated in the same ratio, temporally and spatially! Such a mechanism is an inconceivability, for to ensure the result which does happen the working distance of the forces imagined must be altered in each case according to the size of the fragment removed. Something is therefore required to superintend, to co-ordinate, to harmonize the causes of development in the case not only of the part but of the whole egg as well; and this something is not material. A corroborative proof of the inadequacy of the purely material explanation——-the causal explanation in the ordinary sense of the word—may be derived from a consideration of certain other vital processes. The facts of acclimatization and immunity betray an extraordinary adaptability of the organism to a change in its environment; an organ will adapt itself structurally to an alteration, quantitative or qualitative, of function (Roux’s ‘ functional adaptation ’) ; lost parts can be regenerated; and then there is the physiology of the nervous system!

In all these cases of ‘ regulation ’—and indeed'in all other responses to stimuli—-the same element, inexplicable in chemical ‘ V GENERAL REFLECTIONS AND CONCLUSIONS 291

and physical terms, exists as must exist in development. This entity is not a form of energy but a vital constant, analogous to the constants or ultimate conceptions of mechanics and physics and chemistry and crystallography, but not reducible to these, just as these cannot be translated into one another. Driesch describes it as rudimentary feeling and willing, as a ‘ psychoid ’, as ‘ morphaesthetic ’, or perceptive of that form which is the desired end towards which it controls and directs all the material elements of differentiation. Its activities are thus verae causae— unconditional and invariable antecedents—psychical factors which can intervene in the purely physical series of causes and eﬁects, and for it he revives the Aristotelian term ‘Entelechy ’. Such is the ‘vitalism’ introduced by Hans Driesch, a teleological theory clearly, but not the ‘static ’ teleology of the Analytiac/ze T/zeorie ; rather it is a ‘ dynamic’ teleology which not only sees an end in every organic process but postulates an immaterial entity to guide the merely mechanical forces towards the realization of that end.

This theory would seem to be open to serious criticism, and from two sides, the scientific and the philosophical.

In the first place, we must remind Driesch that on his own showing a comparatively simple structure is all that is necessary to form the starting-point of a developmental process, however complex that may be, and that there is no reason why such a structure should not be divisible into portions, each of which will possess all the parts of the structure in correct proportions, and be therefore totipotent. But such division cannot continue indeﬁnitely, for as we know, and as Driesch knows too, there is always, sooner or later, a restriction of potentialities, and this is due to the manner of distribution at or2'gz'ne of the constituent parts of the whole. When Driesch asserts that this restriction is due to size alone, to mere lack of material, and not lack of speciﬁc material, when he tells us that the blastomeres can be dislocated indeﬁnitely without prejudice to a normal development, when he exclaims that ‘ J edes jedes kann ’, he is manifestly led away by the reaction against the theory of the preformation of as many units as there are inheritable characters on the one hand, and on the othe; by his own erroneous presuppositions as to the con U 2 292 DRIESCH’S THEORIES OF DEVELOPMENT V

struction of the egg out of like particles all similarly polarized and all oriented in the same way.‘

This being so, the first argument based on the ‘ causal harmony ’ will fall to the ground; for this will be given in the initial structure of the egg, and if that may be divided, then the ‘ causal harmony ’ may be divided too. The correct proportionality of the organs of partial larvae, then, offers no peculiar difficulty. The corroborative argument is founded on a consideration of responses to stimuli. This is not a question for the embryologist, but it may be pointed out that there are still physiologists who maintain that even the complex phenomena presented to us in the activities of the nervous system are susceptible of a purely mechanical explanation.

The second series of objections to the new ‘vitalism’ is philosophical. Driesch has quoted the authority of Kant and Aristotle in support of his doctrine. The former is, however, rather a difficult witness, as Driesch is well aware. He complains, indeed, that Kant’s teleology is descriptive or ‘static ’ rather than ‘dynamic’, as is perfectly true, except in the case of man, a point of which Driesch naturally makes the most. There are no doubt passages where Kant speaks of ‘a cause ‘ which brings together the required matter, modiﬁes it, forms it ‘and puts it into its appropriate place ’,2 but against these must be set the explicit statement that if the body has an alien principle (the soul) in communion with it, ‘the body must either ‘ be the instrument of the soul—which does not make the soul ‘a whit more comprehensible’3— or be made by the soul, in which case it would not be corporeal at all. ‘ Vitalism ’ can glean small comfort from this.

Let us turn, then, to the second authority.

Aristotle’s matured reflections on the soul (xpvxﬁ), its nature, functions, and development, are to be found in the treatises De Anima and De Generatione Animalium.

Soul is deﬁned in the most general way as an activity of

‘ C. M. Child (Biol. Centralbl. xxviii, 1908) has recently published a similar criticism of Driesch’s absolutely equipotential systems. Child points out in particular that in the _regeneration of the head of Tubularia the pro ortionality of the parts d1ﬁ'ers in different regions of the stem, and un er different conditions, and cannot therefore always be exact. This §eq1éipotentiality osf gh6e5system is therefore not absolute.

6 . . V GENERAL REFLECTIONS AND CONCLUSIONS 293

a natural organic living body, life being autonomous nutrition and growth and decay. The activity (évrekéxaa) may, however, be latent or patent, passive or active, sleeping or waking, without losing its peculiar characters. This activity is substance (ofmia), but substance as ‘ form ’, as opposed to the material substance of the body; the living body is therefore also a substance, in a double sense.

Soul is not, however, identical with the body, but as form, proportion ()\o'yos‘), activity (e’ve’p~/eta), essence (16 ref 1312 Juan), it is related to the body, mere matter (z'5)\n) and potentiality (bzfuams), in just the same way as the seal is related to the wax, and the body is the instrument whereby it effects its purposes; though subsequent in time it is prior in thought to the body, as all activities are to the materials with which they operate.

At the same time neither it nor its parts are separable from the body, with the exception, possibly, of mind (voﬁs); it is indeed the actual or possible functioning of the body, like the seeing of the eye or the cutting of the axe, and with the disappearance of the capacity of this functioning the soul itself also perishes.‘ Lastly, it is a cause (dpxip xal atria) in a triple sense: ﬁrst, as the source of motion, secondly, as that for the sake of which the body exists, and thirdly, as its essence (oﬁafa), or formal cause.”

The soul is of several kinds, which form together an ascending series, each member of which is necessarily involved in those above it.“

The lowest is the nutritive soul (ﬁpsvrrmf), found in all living things, and the only soul possessed by plants. It is defined as motion in respect of nutrition, decay and growth, processes which involve alteration (&Mo[aoa-is‘) in the body, and its functions (épya) are to utilize the food’ for the maintenance and reproduction of the form of the body, and to control and limit growth.

The second is the perceptive soul (aicrﬂm-uni), the possession of which distinguihes animals from plants. Perception is a kind of alteration (dkkoiwcts 119),‘ and consists in being moved and affected. The fundamental and indispensable perception is touch (écjni),

1 De Au. II. 1. ’ Ibid. II.4. 3 Ibid. II. 3,4. ‘ Ibid. II. 5. 294 DRIESCH’S THEORIES OF’DEVELOPMENT V

for it is concerned in the acquisition of the food. It is invariably present; the others may or may not, some or all, be present.

Some animals are also possessed of a capacity of locomotion, and the performance of this function requires again a special kind of soul.

Lastly, there is the reasoning soul (ecauonnmi), or mind (voﬁs). This is found in man alone, unless there be other beings similar to him, or even nobler than he. Mind alone is eternal and separable from the body.

In all reproduction (except in generafio eguivoca) the startingpoint of a new individual is what Aristotle calls a a--rréppa. In plants, in which he does not recognize the sexes, this is the seed; in sexually produced animals it is the result (mﬁmua) of the mingling of the male (yomi, or anépua in a narrower sense) and female elements ; the latter is an egg or, in Mammals, the catamenia.‘

This avréppa, he holds, does not come from all the organs of the body by a kind of pangenesis, but is a tissue, homogeneous like bone and flesh, and separated out from the food in its ﬁnal stage of digestion, when it is in the form of blood and ready for assimilatiou, and hereditary resemblance is explained by the fact that the food which is about to be assimilated by the organs is naturally like that set aside to form the <r1re'pp.a."

In the matter of the o-n-éppza all the parts of the organism that is to be formed are indeed present potentially, but this means no more than that the material is there.“ Actually (évepyetq), they cannot be present until the soul has been developed, and in particular the soul that is characteristic of animals, the perceptive.‘ Out of this matter the organs are differentiated successively, the heart first, not only as a matter of observation (on the chick), but as a. theoretical necessity, since in it is the principle of growth, then the blood, the blood-vessels, the tissues gathering about these by a process of condensation and coagulation, the foreparts of the body first, and then the hinder.5 The anéppa, then, or xiinua, is a material cause of development, but it is also a cause in other senses ,- it is the eﬂicient cause, since it must

‘ De Gen. I. 18-20. ’ Ibid. I. 18, 19; IV. 1. 39. I 3 Ibid. II. 4. ‘ Ibid. 1. 19; II. 3, 5. ‘ Ibid. II. 6. , V GENERAL REFLECTIONS AND CONCLUSIONS 295

contain the source of motion, and it is further the ﬁnal and the formal cause. These several causes are not, however, all contributed by both parents. The teaching of Aristotle is that the matter is provided by the female, and the female alone.‘ The egg (or catamenia) is described as being matter (z'5)\n), body (a63p.a), potentiality (bﬁuams), passive'(1ra0nru<6v), and merely quantitative, although it is true that a sort of soul, the nutritive, is somewhat grudgingly conceded to it, since unfertilized eggs appear in some sense to be alive.“ The male element, on the other hand, provides the principle of motion (dpxi; 1-ﬁr xwﬁo-ems) and the form (eZ6os) ,- it is qualitative, it is activity, it produces the perceptive soul, if it is not itself that soul, and it is responsible for the ‘ correct proportionality ’ (Myos) of the organization.3

The male element contributes only motion; it acts upon the female element as rennet acts when it coagulates milk, except that the analogy is incomplete, since the yomi brings about a qualitative, and not merely a quantitative, change in the material on which it operates.‘ To this it imparts the same kind of motion which itself possesses, the motion which was present in the particles of the food in its ﬁnal form from which it was itself derived.5

The communication of this motion is enough to set going the machinery (az’;ro';ua1-ov) ; the rest then follows of itself in proper order.“ To impart this necessary motion is the function of the nutritive soul, which is primarily associated with the male, only somewhat doubtfully with the female, element; the perceptive soul which is, and therefore presumably also imparts, motion of a kind (dmofwms) is found in the former alone.7 As to the third kind of soul, mind, Aristotle says little, but it is not introduced in the male element: it is separable and comes in from outside.

Lastly, the sperm of the male acts like a cunning workman who makes a work of art, using heat and cold as its implements as the workman uses his tools; 3 for this heat and this cold could never of theme1ves—by coagulations and condensations——produce the form of the body, as the older naturalists had supposed,

‘ De Gen. I. 20, 21; II. 1, 4. 3 Ibid. II. 5. 3 Ibid. I. 20, 21; II. 1, 4. ‘ Ibid. I. 20; IV. 4. “ Ibid. II. 3. “ Ibid. II. I, 5. 7 Ibid. II. E’); De An. II. 5. ‘ De Gen. II. 4. 296 DRIESCH’S THEORIES OF DEVELOPMENT V

regarding only the material and eﬂicient, and ignoring the formal and the ﬁnal cause ; for the organic body is not what it is because it is produced in such and such a fashion, rather it is because it is to be such and such that it must be developed as it is.‘ And here lies the kernel of the whole matter. For while Aristotle has made it perfectly plain that according to his idea, the soul, at least its nutritive and perceptive faculties, is to be regarded as a function of matter and that this function may be ultimately expressed in terms of movement, and further that development is a mechanism which is set going by the communication of motion proceeding from the ‘soul’ of the male element, and derivable in the last resort from the ‘ motions’ into which the ‘ functions ’ or ‘ soul ’ of the parent can be resolved, to the mere matter which the female provides, it is equally evident that he does not regard this mechanical explanation—in terms of material and eﬂicient causes—as satisfactory or complete. But when we inquire why, he gives us no certain and consistent answer. On the one hand there are passages in which he tells us that there must be something which controls the material forces and imposes upon them a limit and proportionality of growth,” that the soul makes use of these forces as the artist makes use of his implements,“ and such passages are naturally interpreted by Driesch in the sense of a ‘ dynamic ’ teleology ; it is the xpvxrf (not, of course, voiis, but the two lower kinds) which superintends and controls, and the \[tux1f is ‘ Entelechy ’. Elsewhere, however, we are informed that even the proportionality of the developing parts is simply the outcome of the motion imparted by the male, which is actu what the female material only is 12otent2'a”.4 Moreover it may be questioned whether Aristotle ever intended to imply more than an ‘ analogy with the causality of purpose’ when he uses the ﬁgure of the workman and his implements to illustrate his meaning of the formal cause. The formal cause of a work of art is an intelligible ‘ vera causa ’, it is the idea in the mind of the artist antecedent to the execution of the work, but the formal or ﬁnal cause of an organism, the end which it apparently strives to attain, is only metaphorically prior in time to the existence of the organism itself. Prior in thought, how ‘ De Gen. V. 1. 9 De An. II. 4. ’ De Gen. II. 4. ‘ Ibid. II. 1. 44V GENERAL REFLECTIONS AND CONCLUSION S_ 297

ever, it certainly is, for it is only the performance of its functions (iv-rehéxeca) by the organism complete in all its parts that makes the mere mechanism of development comprehensible to us; the process, therefore, exists for the sake of the end. Only as eﬂicient cause is the soul prior in time ; only so far as it is prior in thought can it be said to be a ﬁnal cause.‘

Such a teleology is, it is obvious, indistinguishable in principle from the position in which Kant leaves us. It is the position adopted by Driesch, as we have seen, in the Analytisc/le T/zeorie, but abandoned in the Vitalismue in favour of a theory of ‘pyschoids ’.

Now, quite apart from the meaning which Aristotle may or may not have intended to convey, there appear to be grave objections to this belief.

This ‘ psychoid ’, to which the name ‘ Entelechy’ is surely misapplied, this rudimentary feeling and- willing, which is aware of the form it desires to produce, must be, psychically, at least as complex as the phenomena it is designed to account for, and stand, therefore, as much in need of explanation as they, which will involve us at once in an inﬁnite series of such entities. In fact, to borrow the epithet which Driesch himself has bestowed on the nuclear architecture imagined in the Roux-Weismann hypothesis, it is only a photograph of the problem, and not a solution at all. Again, when we ask what the modus opcramli of this cause is we get no reply either from Driesch or from any other neo-vitalist, though this is just the knowledge that we so urgently stand in need of. The objection that the intervention of a psychical cause in a physical process is unintelligible, an objection which would probably appeal to many, may be waived, for in the last resort the connexion between any——even simply mechanical—-causes and eﬁects is equally hard to understand. It may, however, be seriously doubted whether these entities are not being ‘ multiplied beyond necessity ’, and whether the progress of science would not be better served by an adherence to a simpler philosophy.

‘Vere scire est per causas scire.’ The maxim of the great founder of modern inductive science is the watchword of embryologists to-day. By exact observation and crucial experi ‘ De Part. II. 1. 7. 298 DRIESCH’S THEORIES OF DEVELOPMENT V

ment, utilizing every canon of induction, the facts of development are to be brought under wide general laws of causation, which will be in the ﬁrst instance physiological laws—of response to stimuli, of metabolism, and of growth: by means of these laws we can predict, and our predictions can be veriﬁed. The thought process cannot, however, rest here. Ultimately—-—as we believe—it may be possible, no more than that can yet be said, but it may be possible to state the widest generalizations of biology in chemical and physical, and these again in purely mechanical, terms. Thus evolution of form in the individual as well as the larger evolution of form in the race, become but the ﬁnal terms in a far vaster cosmic progress, from ‘ homogeneity to heterogeneity ’.

The idea, of course, is perfectly familiar ; it is the analysis of purely physical causes carried to its extremest limit. Phenomena are thought out in terms not of origins merely but of one origin, and that one origin is the only mystery that remains. This uniﬁcation of the sciences always has been, and must still remain, the dream and the faith and the inspiration of the scientiﬁc man, and could such an ediﬁce of the intellect ever be realized the task of science would have been completed.

But where science leaves off there philosophy begins, and it is for philosophy to attempt the solution of this last mystery of all.

Philosophy cannot rest content in an endless regress of cause and eﬁect, and a ﬁrst supreme cause, first in time that is, is metaphysically out of the question. An original homogeneity is equally unthinkable, for out of a system all whose parts are absolutely alike, by no imaginable process could any heterogeneity ever be evolved.

That ﬁrst simplicity must have contained potentici all that has since developed out of it, it must have possessed a structure, an arrangement of parts such that the end which it has realized, or is to realize, would be what it is or will be, and to regard the end as well as the beginning is the duty of philosophy, a duty which Aristotle and Kant have both impressed on us. The outlook of pure science, an outlook to which, gm? science, it cannot too rigidly conﬁne itself, is thus supplemented and enlarged. V GENERAL REFLECTIONS AND CONCLUSIONS 299

Knowledge through material and eﬂicient causes is rounded into a whole through a knowledge of the ﬁnal cause, which, in the last analysis, is just as much a rem cauea as they are; for in our total ignorance of what constitutes the invariable connexion we observe between antecedent and consequent it is as true that the second causes the ﬁrst as the converse. Only because of the inherent desire of the human mind to predict from the past, which is known, to the future, which is not, have we come in ordinary usage to restrict the term to the antecedent.

The ultimate end of the human or any other race we cannot tell, the ultimate end of the universe we cannot tell, any more than we can imagine its absolute beginning, unless we ﬁnd these ends in the freedom of the moral consciousness of man. But the end of an organism, the production of speciﬁc form and the maintenance of that form which itself has produced, does seem self-evident and plain, though we must never forget that form is variable and subject to change, that species are not immortal any more than individuals, and that the effort to achieve that form does not invariably succeed.

Here, however, we touch on the fringe of a problem—the problem of evil—too large to be discussed in this place. Putting this aside, a purposiveness is an unmistakable characteristic of the functions of living things, of the production and preservation of form, a characteristic which still remains when those functions have been expressed in terms of the chemistry of the proteids. It is only, however, by a remote analogy with our own ‘causality according to purposes’ that we can speak of organic functions as purposive ,- it is only as they were guided and controlled by an intelligence; their purposiveness is indeed only the expression of our inability to comprehend their beginnings except in terms of their ends; it is relative to us, though not, therefore, any the less real.

Biology, then, although built upon the ultimate conceptions of chemistry and physics, has yet peculiar features of its own. Its relation, indeed, to these lower sciences is just what their relation is to one another. A survey of the whole hierarchy displays to our view a series in ascending order of complexity; each member of this series has its own ultimate conceptions, the 300 DRIESCH’S THEORIES OF DEVELOPMENT V

most general expression for the facts with which it deals, but the ultimate conceptions of each, as in Aristotle’s series of ‘ souls ’, are necessarily involved in the one next above, while conversely each endeavours to translate its own ultimates into those of the science below: a translation, however, which, be it never forgotten, leaves the reality of the original undestroyed.

Thus, mechanics expresses molar motions in terms of pure numbers, physics explains forms of energy-—heat and light and electricity—as the motions of molecules; chemical affinity is to be reduced to the mutual attractions of intramolecular atoms; purposive responses to stimuli may be stated in terms of chemical reaction, and the psychical phenomena of mental and moral science—understanding and feeling and will—are a form of these.

The cosmic process thus takes place in a succession of stages, and the peculiar features which mark each individual stage are simply the outcome of an increase in complexity of the peculiarities characteristic of the stage below. To establish this is the ﬁnal achievement of science.

Nevertheless, the facts with which each science starts, the facts which come ﬁrst in the order in which knowledge is acquired, do not become wholly merged in those simpler facts into which they are at each stage translated ,- when the translation has been accomplished the original still remains.

The ‘secondary qualities’, as well as the other properties of those bodies whose behaviour the physicist investigates, are as real as, no more and no less relative to our intelligence than, those ‘primary qualities ’ of impenetrability and extensionexhibited by those bodies whose behaviour forms the subjectmatter of mechanical science—into which it is his endeavour to translate them, just as the primary qualities themselves obstinately refuse to be reduced to mere number. Chemical afﬁnity remains as a phenomenon sui generis after it has been reduced to the operations of intramolecular forces, and the purposiveness of responses to stimuli is something over and above the chemical reactions to which they are rightly referred. Last of all, the ﬁnal term in the series, "the mental and moral consciousness, the ‘other side’ of certain purely physical functionings of the organism, is as real as any’ of those qualities of matter which V GENERAL REFLECTIONS AND CONCLUSIONS 301

have been step by step involved in its evolution. In a word, the increase in the complexity- of the phenomena which marks the transition from each of these stages to the next is itself a new phenomenon and cannot be ignored.

And herein we may perhaps discover the essence of the relation between ‘mind’ and ‘ matter’, whether in ordinary function or in development. The mind is not matter, not even living matter; rather it is the new quality constituted by an increase in the complexity of living matter, immaterial and as distinct from that matter as is ‘ blueness’ from vibration of a certain wave-length. Dependent on and inseparable from matter, however, it is; when that matter, whether in the individual or in the race, attains a certain degree of complexity, then and then only does mind appear; and with the disappearance of that complexity it perishes.

While, therefore, we have no reason for supposing that the mind ‘ comes in from outside ’, we are at the same time saved from that somewhat extravagant ‘ psycho-physical parallelism ’ which, to explain the evolution of consciousness, postulates a complete psychical, accompanying the complete physical series of causes and eifects, and credits, of necessity, the merest matter with the rudiments of feeling, thought and will. On the other hand, if the mind cannot be said to intervene in the physical series, there seems to be no alternative but to suppose that the operations of the one, when they exist, are parallel to those of the other.

Such a system of philosophy as that which we have here ventured to suggest can give no countenance to a Vitalism which interpolates an unnecessary psychical element into the complete causal chain of physical events. But it is not for that reason to be condemned as materialistic ; for the mind, developed out of and conditioned by matter, the last term and ﬁnal cause of the whole process, is not itself matter but an accompaniment of certain material complexes, and still remains when they have been resolved into simple mechanical expressions. And in this mind, last in time but ﬁrst in thought, a larger philosophy will perceive not only the end towards which, in time and space, matter strives, but the Undertanding which, itself eternal, imposes the forms pf space and time upon that Nature which it makes. 302 DRIESCH’S THEORIES OF DEVELOPMENT V

LITERATURE

ARISTOTLE. De Generations Animalium, ed. Bekker, Oxford, 1837. ARISTOTLE. De Partibus Animalium, ed. Bekker, Oxford, 1837. Anrsrorm. De Anima, ed. Trendelenburg, Berlin, 1877.

H. Dnnzscn. Analytiache Theorie der organischen Entwicklung, Leipzig, 1894.

H. Dawson. Entwicklungsmechaniache Studien. X. Ueber einige allgemeine entwicklungsmechanische Ergebnisse, Mitt. Zool. Stat. Neapel, xi, 1895.

H. DRIESCH. Resultate und Probleme der Entwicklungsphysiologie der Tiere, Anat. Hefte, 2“ Abt., viii, 1898.

H. DRIESCH. Die Lokalisation morphogenetischer Vorgiinge: ein Beweia vitalistischen Geschehens, Arch. Ent. Mech. viii, 1899.

H. DBIESCH. Der Vitalismus a.ls Geschichte und ale Lehre, Leipzig, 1905.

I. KANT. Ktitik der Urteilskmft, Eng. Trans. by J. H. Bernard, London, 1892.

C. VON NAGELI. Mechanisch-physio]ogische Theorie der Abst:uumungslehre, Mﬁnchen und Leipzig, 1884.

Gr. PLATNER. Kern und Protopla.sma., Breslau, 1887.

Review in Jahresber. yes. Medicin, xxii, 1888.

W. Roux. Zu H. D1-iesch‘s ‘Analytischer Theorie der orgzmischen Entwicklung’, Arch. Ent. Mech. iv, 1897. APPENDIX A

FURTHER REMARKS ON RELATION BETWEEN THE SYMMETRY OF THE EGG, THE SYMMETRY OF SEG MENTATION, AND THE SYMMETRY OF THE EMBRYO IN THE FROG.

IN the measurements, referred to above (pp. 165-8), of the angles between the plane of symmetry of the egg (as determined by the position of the grey crescent), the ﬁrst furrow and the sagittal plane of the embryo, it was found (1) that there was a certain tendency for the ﬁrst furrow and the sagittal plane to coincide, since in a. large number of cases small angles preponderated over large ones, the standard deviation of this angle from the mean (which was practically = 0°) being a- = 40-39° i-65 ; (2) that there was a much greater tendency for the plane of symmetry and the sagittal plane to coincide, the standard deviation of the angle between these two planes being o'=29-75° _-J; -63 ; (3) that the first furrow tended either to coincide with or to lie at right angles to the plane of symmetry, the standard deviation about 0° being 18-70° i -60, that about 90° being 23-29° j-_ -86, the value of 0' for all the observations being 47-90° 1- 1-19. The correlation between the first furrow and the sagittal plane was found to be p=-138i -031, that between the plane of symmetry

and the sagittal plane p=-372i -025, that between the plane of symmetry and the first furrow p=-O87 i -032.

These results may be tabulated as follows : rr :0 40-39° + -65. -138 i -031.

tal Plane.

Plane of Symmetry and Sagittal Plane. l 2975 -t '63’

Plane of Symmetry and First Furrow.

First Furrow and Sagit- }

.372 i .025.

} 47.9oi1.19. .os7¢_.o32.

Full details of these results will be found in a paper in Biometrika V. 1906.

For the purpose of making these measurements the eggs were placed in rows parallel to the [mat]; of glass slides, and the angles measured between the various planes and lines ruled across the slide. Such eggs compress one another by their jelly 304 APPENDIX A

coats; further, the eggs taken ‘from the uterus were placed haphazard on the slides with the axis making any direction with the vertical. The egg takes about half-an-hour to turn into its normal position with the axis vertical, and during this interval gravity may possibly act upon the yolk and protoplasm, of different speciﬁc gravities, and impress a plane of bilateral gravitation symmetry upon the egg, as occurs when the egg is permanently inverted (see above, pp. 82-87). This obliquity of the axis may possibly aﬁect the relations between the planes, and the mutual compression may also be a disturbing factor, since it is known that in compressed eggs the nuclear spindle is perpendicular to the direction of the pressure (pp. 34-36).

These angles have therefore now been measured under four different conditions:

(a) The eggs are close to one another in the rows and the axis is horizontal.‘ (Since the rows are parallel to the length of the slide the pressure, if any, must be in the same direction, while the surfaces of compression or contact are across the slide. The eggs were always so placed that the vegetative poles faced in one direction and the planes of ‘ gravitation symmetry ’ were at right angles to the length of the slide. This holds good of all the following experiments.)

(/3) The eggs close, but the. axis vertical with the white pole below. In these there can be no gravitation plane of symmetry.

(y) The eggs spaced, but the axis horizontal. In these the jellies do not touch.

(6) The eggs spaced and the axis vertical. In these, therefore, both the supposedly disturbing factors are removed. The results are given in the following table :—

A B C

First Furrow and Plane of Symmetry Plane of Symmetry Sagittal Plane. and Sagittal Plane. and First Furrow.

(.7) .7 = 38-42 g._ -70. .7 = 31-86: -56. .7 = 41-591-_-84. ,7 = -201;-028. ,7 = -263;:-_-027. ,7 -= -118;-029. (.9) .7 = 33-443-_-56. .7 = 30-17:51. .7 = 39-7l_-1;-61. ,7 = 3523-021. .7 = .27si.o22. ,7 = .o23¢.o24. (-,) .7 —_- 33-49;:-_-96. .7 = 27-53¢-84. .7 = 36-60: 1-108. ,. = .292:-039. —_— -399:-036. p = .075:-043. (a) .7 = 31.45133. .7 —_— 26-80¢-82. .7 = 34-46:1-065. ,7 —_- -364:-033. ,7 = -451 1.035. ,7 -_- -186;-043.

It is evident from this that gravity and ‘ mutual compression ’ (as I will for the moment term it, though it is doubtful whether the pressure has anything at all to do with the result) do affect the

magnitude of the angles between these three planes,_for in each APPENDIX A 305

case the standard deviation falls, while the correlation coeﬂicient rises, when they are both removed. It will be observed that, while gravitation (y) has less eﬂect than compression (3) upon the angles B and C, the reverse is the case with the angle A. We may be able to ﬁnd a reason for this later on.

There is one point worth noticing. It is quite clear that gravity is not indispensable for the development of a grey crescent and plane of symmetry, though it is true that the position of this plane may be aifected by gravity even in the short interval that elapses before the egg turns over.

The values for the compressed eggs with horizontal axes (or) compare fairly well with those previously obtained, except in the case of the plane of symmetry and the first furrow. In the former series the latter tended either to coincide with or to lie at right angles to the former. In the present series this is not the case. This diﬂerence is probably to be attributed to the fact that many of the eggs in the ﬁrst series must have been placed on the slide with the white pole upwards: possibly also the ‘ compression ’ was greater then than now.

It is fortunate that the same data enable us to study exactly the relation between the ﬁrst furrow and the plane of symmetry on the one hand, and the direction of ‘compression’ and of the gravitation symmetry plane on the other. It must be remembered that these two are at right angles to one another.

Consider first the ﬁrst furrow.

(a) When the eggs are close but the axis horizontal the ﬁrst furrow tends to lie at right angles to the slide, that is, in the direction of compression, but at right angles to the gravitation symmetry plane. (a-=38-16 i -69.)

(ﬂ) When the eggs are close but the axis vertical this tendency is not quite so marked. (a'=46-67 i -7' 1.)

(y) When the eggs are spaced and the axis horizontal it is still there, but slight. (o-=49-32 ;l-_ 1-40.)

(6) When the eggs are spaced and the axis vertical the direction of the first furrow is random. (zr=52-76¢ 1-17.)

VVe may conclude, therefore, that the ﬁrst furrow tends to lie in the direction of the ‘ compression’ and at right angles to the plane of gravitation symmetry. The latter tendency, we know, exists in forcibly inverted eggs, together with a tendency to lie in the plane of symmetry and at 45° to it (above, p. 84). Pressure experiments alo show that division is in the direction of pressure (p. 34 sqq.).

The direction taken up by the plane of symmetry under these different circumstances is-quite distinct from that of the ﬁrst furrow. It appears to be determined in the ﬁrst instance by

JINKINIOI ° X 806 APPENDIX A

gravitation, as it usually lies in the gravitation symmetry plane. It is not, however, only so determined, for if the eggs (compressed and with axi horizontal) be allowed to develop in the light the plane of symmetry lies either in the gravitation symmetry plane, or in the direction of the incident light (parallel to the length of the slide in the experiment , while in the dark it lies only across the slide. That this secon effect is due to the light and not to the pressure is shown by the fact that it occurs when the eggs are spaced, and that it may be made to vary in position by varying the position of the slide with regard to the light. Light, therefore (ordinary daylight), as well as gravity, can help to determine the -position of the plane of symmetry, and when the latter is excluded it appears that this plane is placed either in or at right angles to the source of light.

Light appears to exert no effect u on the ﬁrst furrow.

It is now intelligible why, when 1 these factors are operative, the relation between the first furrow and the planes of symmetry of egg and embryo should be disturbed, since, in the conditions of the experiment, those factors which determine the position of the former are at right angles to those on which the direction of the latter depends.

It still remains for us to inquire into the internal causes of the direction of these planes in the egg. Roux, as has been pointed out, has asserted that the grey crescent appears on the opposite side of the egg to that on which the spermatozoon has entered (pp. 80, 165), and further that the point of entry of the sperm also determines the meridian of the first furrow, since this either includes the sperm-path, or is parallel to it, or, when it is crooked, includes or is parallel to the inner portion or ‘ copulation ’ path, which is taken to represent the line of approximation of the two pronuclei; the outer part being simply the ‘ penetration’ path. Roux also arbitrarily selected a fertilization meridian (meridian of the sperm-entry), and showed that this became the ventral side (opposite the grey crescent) later on, as well as the ineridian of the first furrow (p. 248).

I have been able to accurately investigate—by means of sections-—the relation between the fertilization meridian, ﬁrst furrow, and sperm-path in a number of eggs in which the direction of the symmetry plane had been previously determined, and the results of the measurements of these angles are given here. The eggs fall into two series, those which were compressed and had their axes horizontal (a), and those which were spaced and had their axes vertical, the white pole being below (6). In (a) the gravitation symmetry plane and the direction of compression were at right angles to one another, as before. APPENDIX A 307

8 a Meridian of sperm entry a- = 21-02° 1-_ 1-63. o- = 31-04°: 1-34. and ﬁrst furrow. p = -435 3 -074. 9 =-613 i -038.

Meridian of sperm entry 0' = 25-67° i 1-35. 0 = 41-01° 3-_ 1-78. and symmetry plane. p = -302 i -083. / P = -006 1 -061.

SP§;§§f:;l3(g;§‘}ff1,§§;“ } . .. .—. 17.94° : 1.15. o‘ = 21.47° 1 -93.

From this it is clear that there is a very close relation indeed between the point of entry of the spermatozoon and the direction of the ﬁrst furrow, especially when the disturbing eﬁects of pressure and gravity are removed. There is, however, little relation between the sperm meridian and the plane of symmetry even under the most favourable circumstances, and when the conditioﬁs are not favourable the correlation is negligible. There is however (in the 6 series) a considerable correlation (p = -479 i '070) between the sperm-pat/l and the plane of symmetry. It should be remembered, however, that all these eggs were exposed to the light. From what we know of the eifect of this agent upon the direction of the symmetry plane, it would not perhaps be too hold a hazard to surmise that in darkness there would be a correlation between the sperm entrance and the plane of symmetr .

Eiien after the removal of this disturbance there remain factors which interfere with the completeness of the correlation between these planes; these must probably be looked for in the incomplete radial symmetry of certain eggs—due possibly to pressure in the uterus—and to the slight squeezings and distortions the eggs may be subjected to when they are being taken from the Frog.

It will be seen that the relation between the sperm-path and ﬁrst furrow is closer than that between the latter and the sperm entrance. This is because though the furrow may be placed to one side of the entrance point, it may still be parallel to the path , or, if not to the ‘penetration ’ path then to the inner or ‘copulation ’ path, as observed by Roux. This ‘ copulation’ path is usually observed when the penetration path is turned away from the first furrow, that i, when it has not been directed towards the egg-axis.

The same data give the position of the point or of entrance with regard to the direction of ‘pressure ’ and ‘gravitation symmetry’. In the (a) series the sperm tends to enter in the direction of ‘pressure’, that is, on that side of the egg on which it is in contact with its neighbours. Hardly a single spermatozoon enters on that side of the egg on which the white pole had been turned up, and very few on the opposite side.

X2 308 APPENDIX A

It is scarcely possible to suppose that either the compression of the egg or the gravitation plane brings the spermatozoa round to the side of compression, but it may be imagined that either by capillarity or by some chemotactic stimulus the spermatozoa are especially attracted to the point where the rapidly swelling coats of adjacent eggs come into contact, and that therefore fertilization is principally effected upon this side. This explains why the ﬁrst furrow lies so often in this direction. The pressure may of course aﬁect the position of the planes in the egg later on.

When the eggs are spaced the sperm enters on any side at random. The deviation of the sperm entrance from the egg-axis (the angle between sperm-entrance radius and egg-axis) varies in the two series of observations. When the eggs are spaced and the axes vertical, the sperm enters mainly near the equator, never near the animal pole; when the eggs are compressed and the axis horizontal, usually at about 45° from the axis, though it may enter near the pole or near the equator. This diﬁerence obviously depends on the diiference in the initial position of the eggs on the slide. The deviation has apparently very little effect on any of the planes we have been considering.

Finally, let us try and gain some conception of the mechanism by which the direction of the furrow depends on the point of sperm entry. It is apparently quite simple, for the sperm-path is directed usually towards the axis, the sperm nucleus travels along that path to meet the female nucleus, which is also in the axis, the centrosome of the sperm divides at right angles to that path, the fertilization spindle is developed between the diverging centrosomes and cell-division takes place in the equator of the spindle ; the first furrow includes therefore the sperm-path. Should, however, the ‘penetration ’ path not be exactly radial, for whatever reason, the sperm nucleus turns aside to meet the female pronucleus, there is a ‘ copulation’, as distinct from a ‘ penetration’ path, the centrosome divides at right angles to the former, and this, then, is included in or parallel to the plane of the furrow. In those cases in which the sperm-path is parallel to the furrow it is always quite close to it, and we may suppose perhaps that the ﬁrst division "has not been quite equal. (The division of the centrosomes has not, I believe, been observed in the Frog, and the foregoing description has been taken from the Axolotl. In this genus the deﬁnitive centrosome is formed from the sperm nucleus, when the latter has already penetrated some little way into the egg.) .

The causes of the formation of the grey crescent which marks the symmetry plane are not so clear. APPENDIX A 309

Roux describes it as being due to the immigration of superﬁcial pigment. Now we have strong reason for believing that both the entrance-funnel——produced when the spermatozoon ﬁrst touches the egg-—and the sperm-sphere are local aggregations of watery substance. The accumulation of what appears to be a more watery substance about the middle piece which has been observed in the Axolotl,appears also to occur in the Frog: at least the same formation of large clear vacuoles in the sperm-sphere may be seen in the latter as in the former. Should this be actually so, we may suppose that the streaming movement centred in the entrance-funnel and sperm-sphere is responsible for drawing away the pigment from a certain region of the surface; hence the grey crescent. The sperm-sphere is on the inner side of the sperm nucleus: hence the grey crescent would appear on that side of the egg which is opposite to the entrance of the spermatozoon, should no disturbance of the streaming movement have taken place, and, since the sperm-path is radial, would be symmetrically disposed with regard to it. In this case, fertilization meridian, sperm-path, grey crescent and plane of symmetry, first furrow, and, later on, sagittal plane, would all coincide. There is, as we have seen, a very fair correlation between the sperm-entrance and the ﬁrst furrow, and again between the sperm-path and the grey crescent. But should some other streaming movement of the cytoplasm be set up by the gravitation of the heavy yolk particles, or by pressure, or by light, then the relation between the two processes, the division of the centrosome which determines the direction of the ﬁrst furrow, on the one hand, and on the other, the streaming movement towards the sperm-sphere which determines the position of the grey crescent, would be disturbed, and while the entrance point of the sperm might still continue to determine, though not so completely, the position of the furrow, it might come to be without relation to the symmetry of the egg and of the embryo; and this is what is actually observed.

Though it is diﬂicult to assign the exact cause of each and every deviation from the rule, this much is certain, that however they may coincide in ‘typical’ development (I use R0ux’s expression), the factors which determine cell-division, and those which determine differentiation, may be influenced by different external causes in widely diifering ways, and are therefore presumably distinct. Nor does this artiﬁcial separation of the two processes in any wise prejudice the complete normality of the

development of the embryo". 310 APPENDIX A

Lillie has shown (Jozmz. Esp. Z002. iii. 1906) that in the egg of C’/Iaetopterus there are granules of diﬁerent kinds which pass, in segmentation, into deﬁnite cells. By means of the centrifuge some of these--the endoplasmic—-may be driven to one side of the egg, but in whatever position these organ-forming granules may be thus artiﬁcially placed, the cleavage has the same relation to the egg axis (as determined by the polar bodies) as in the normal egg. The factors of cell-division are thus separable from those of differentiation.

To the cases quoted in the summary on pp. 245, 246 might be added the various instances in which an egg may be made, by heat or pressure or shaking, or in artiﬁcial parthenogenesis, to segment abnormally and yet give rise to a normal larva. APPENDIX B

ON TIIE PART PLAYED BY THE NUCLEUS IN DIFFERENTIATION

(i) BOVERI has more recently (Zellen-Studim, vi, Jena, 1907) published a very elaborate account of the irregularities produced by dispermy in Echinoid eggs, in which are brought forward

still more facts in proof of the qualitative difference of the chromosomes.

As has been stated above, p. 263, dispermy is induced by the simple expedient of adding a large quantity of sperm to the eggs. The following types of dispermy are distinguished.

A. Tetracentric, i. e. each sperm centre divides. (i) 'I‘etraster, with four spindles.

(ii) Double spindle, i. e. the female and one male pronucleus lie in one spindle, the other male lies aside in its spindle.

B. Tricentric, one sperm centre remaining undivided. (i) Triaster, a tripolar ﬁgure with three spindles.

(ii) Monaster-amphiaster, the undivided sperm centre remaining apart with one sperm nucleus.

C. Dicentric, neither sperm centre dividing. (i) Amphiaster, a spindle is formed between the two centres.

(ii) Double monaster: the centres remain apart, one with one male, the other with the other male and the female pronucleus.

The segmentation of these eggs is as follows.

The tetraster divides simultaneously into four, which may either lie in one plane if the divisions are meridional, or be tetrahedrally arranged. In the first case another meridional division ensues, followed by an equatorial, then ‘eight micromeres are formed, eight macromeres, and sixteen mesomeres. In the latter case not more than three cells can share in the micromere region and only four or six of these are produced. The triaster eggs, having divided simultaneously into three (meridionally), subsequently show six micromeres, six macromeres, and twelve mesomeres.

The segmentation of the double spindle eggs is interesting and important. Usually the egg divides across the two spindles 312 APPENDIX B

into two binucleate cells, but it may divide at once into four, or into three, one of which is binucleate. The interest lies in the binucleate cells, for they continue to produce uni-nucleate and binucleate cells until the latter divide simultaneously into four, and this simultaneous division may sometimes involve an irregular distribution of the chromosomes, with fatal consequences to the cell. Bovcri had already produced evidence of the evil effects of an irregular distribution of the 3 n x 2 chromosomes present in triasters and tetrasters. A more detailed account is now given.

Of the tripartite (triaster) ova about 8 % on an average produced Plutei. In these larvae three regions may be distinguished in the egg by the size of the nuclei (proportional to the number of chromosomes) and the boundaries between them may be shown to correspond to the divisions between the three blastomeres. The form is asymmetrical in skeleton and pigment, but Bovcri shows that both sides are normal, as though the larva had been compounded of two types such as occur, as individual variations, in any culture. It is suggested therefore that the slight differences in the two sides are due to diﬁerences in the two sperms.

Some of the larvae have partial defects in skeleton or pigment, or the skeleton may be much reduced on one side, or one-third of the cells may be pathological, i. e. disintegrate in the segmentation cavity, while the remaining two-thirds are sound and sometimes symmetrical. In this case it is supposed that the degenerate cells had separated from the others at an early stage, and that the remainder had had time to recuperate. In others two-thirds are degenerate, one-third normal, or all three degenerate. When the three blastomeres are isolated and allowed to develop independently, segmentation is partial, with two micromeres, two macromeres, and four mesomeres, and often all three develop normally up to the blastula stage. After that only one or two, rarely all three, become Plutei, the rest giving rise to stereoblastulae or stereogastrulae, full of degenerating cells.

The isolated quarters of tetrasters also segment partially and normally, but few give rise to Plutei. The whole simultaneously quadripartite eggs only rarely give rise to what may be called a Pluteus (2 cases in 1500) ; but very degenerate larvae are found, with masses of disintegrating cells inside, which are assigned to one of the four blastomeres. Stereogastrulae-—with nuclei of all the same size--are frequent.

As has been alread mentioned, Bovcri points out that the probability of each cell’ of a triaster receiving a complete set of the 71. chromosomes of the species when there are 3 n x_2 to be distributed must be greater than‘ that of each cell of tetraster obtaining a full complement, and the probability for one isolated APPENDIX B 313

cell must be greater than that for the whole egg. What the mathematical values of these probabilities are Boveri does not know, though he makes an attempt to reckon them—not theoretically, but by means of a mechanical apparatus; the attempt is not quite successful. The fact, however, remains that eight per cent. of the triasters produce normal Plutei, only -06 per cent. of the tetrasters. This does not depend on the cells receiving too much or too little chromatin (see p. 265), nor again on the fact that the ratio between size of nucleus and size of cytoplasm (see pp. 268, 269) can only be satisfied by certain deﬁnite numbers of chromosomes, and the only explanation remaining is that for normal development of each and every part the nucleus of each cell must contain a complete set of the speciﬁc chrosomomes ; from which it follows that the chromosomes are qualitatively unlike.

A word may be said about the double-spindled eggs (Type A. i). The larvae from these sometimes show abnormal regions, and this is attributed to one or more of the binucleate cells having divided with a tetraster and irregular distribution of chromosomes. Of all such eggs 50 % gave rise to normal Plutei.

The degenerative changes undergone by the nuclei of these larvae are of several types, to be associated again with differences in the combinations of chromosomes.

(ii) Boveri’s experimental proof of the qualitative difference of the chromosomes does not of course of itself involve a belief in the individuality of these bodies, for if the chromatin is concerned in inheritance, it is necessary to suppose that the number of qualitatively distinct bodies is far greater than the number of chromosomes, and these bodies may be differently grouped during each successive resting stage.

The hypothesis of the individuality of the chromosomes, i.e. of a constancy in the manner of grouping of these particles, rests in the first instance on such facts as those observed by Sutton in B2-ac/:3/stola, where in the spermatogonia the chromosomes are of dilferent sizes, which may however be arranged in pairs, together with an odd one or accessory chromosome. 1 In the resting stage the accessory chromosome remains apart in a separate vesicle, while the large chromosomes lie in separate pockets of the nuclear membrane, the small ones, each as a separate reticulum, in the main body of the nucleus. In the spermatocyte a number of bivalent spiremes appear, which show the same dilferences of sizes a the pairs of chromosomes previously, and the accessory chromosome.

The accessory chromosome passes into two only of the four spermatids and is supposed to be a sex-determinant. 314 APPENDIX B

Similar facts have been reported by Wilson for several Insects (see Joum. Esp. Zool. ii, iii, 1905, 1906). '

Wilson ﬁnds constant size differences between pairs of chromosomes, and either an accessory odd chromosome (which passes into only one half of the germ cells) or a pair of idio-chromosomes of unequal size (one of which goes to one half, the other to the other half of the spermatozoa), or both the accessory and the idio-chromosomes (giving four kinds of spermatozoa). The idiochromosomes are supposed, again, to play a part in sex-determination. Several other observers have found these accessory chromosomes, idio-chromosomes, and pairs of chromosomes of diﬁerent sizes in various Insects (Boring, Journ. E211. Zool. iv. 1907 ; Stevens, ibid. ii. 1905, v. 1908; McClung, Biol. Bull. iii. 1902, ix. 1905; Montgomery, Biol. Bull., vi. 1904; Baumgartner, Biol. Bull. viii. 1904-5 ,- Zweiger, Zool. Anz. xxx. 1906; Nowlin, Jomw. Exp. Zool. iii. 1906); in Spiders (Wallace, Biol. Bull. viii. 1904»-5 ; Berry, Biol. Bull. xi. 1906); and in Myriapods (Blackman, Biol. Bull. v. 1903 ; Medes, Biol. Bull. ix. 1905).

It is a noteworthy fact that the accessory chromosome retains its individuality in the resting stage (looking like a chromatin nucleolus), while the others break up. The belief in the individuality of these others rests therefore on the constancy of the relative sizes from generation to generation.

Further support for the hypothesis may be derived from theoretical speculations. VVe know that only 2; (one-half the normal number) chromosomes are necessary for normal development provided that they comprise a complete set. In sexual reproduction n maternal unite with n paternal. A study of the reducing division shows that 1: whole chromosomes first pair with and are then separated from or whole chromosomes, and that when they dilfer in size those of the same size pair together, and it looks as though paternal were here separated from maternal, though the distribution of paternal and maternal to the two cells will diﬁer, almost certainly, in diiferent cases.

If the particles of which the chromosomes are composed are also to be paired and separated, it would appear to be necessary that their groupin should be constant, in other words that the chromosomes shou d retain their individuality.

(iii) A case of heterogeneous fertilization between eggs of Seaurchins and the sperm of Anletlon has been described above (p. 262). Loeb has recently succeeded in rearing Plutei from the eggs of Slrongylocmlrolue fertilized by the sperm of a Mollusc (0/lloroaloma). Cytological details are not given (Arc/E. Eul. Mecﬁ. xxvi. 1908). ‘ INDEX OF AUTHORS

Agassiz: effects of fertilization in Ctenophors, 250.

Aristotle: theory of development, 13.

— the soul in function and development, 292 sqq.

— mechanism and teleology, 296.

Auerbach :' segmentation of Ascuris nigrovenosa, 33.

von Baer, 16.

Balfour: effect of yolk on segmentation, 29, 88.

Bataillon: monstrosities osmotic pressure, 120, 135.

—- artiﬁcial parthenogenesis, 124.

Bergh: cell-division in germ-bands of Crustacea, 34.

Berthold: surface-tension and celldivision, 41, 42.

Bischoﬂ‘, 16.

Blane: effect of light upon the development of the Chick, 94, 96.

Boas: rate of growth in man, 63.

— change of variability, 73, 74.

— diminution of correlation coeﬁicient, 75.

Bonnet : emboitement, 14.

— preformation, 15. Bonnevie : diminution of chromosomes in Ascaris lumbricoidcs, 258. Born : gravity and development, 18, 88-85.

— pressure experiments on Frogs’ eggs, 34, 35.

Boveri : early development of Slrongylocentrotus, 23, 183-185.

— egg of Strongylocentrotus stretched, 39.

— suppression of micromeres in Strongylocentrotus, 186.

-— causes of the pattern of segmentation, 197.

— karyokinetic plane, sperm path,

11 ng first furrow in Strongylocentrotus,

8 .

— potentialities of? animal and vegetative cells, 192.

— stratification of cytoplasmic substances, 242, 280.

-- characters dependent on cytonlmam in Flnhinnid larvae. 261.

due to

Boveri : diminution of chromosomes in Ascaris megalocephala, 252, 255-257.

— due to a difference in the cytoplasm, 257.

— hybrid larva from enucleate egg fragment with characters of male parent, 253, 258-260.

— irregular distribution of chromosomes a cause of abnormality, 253, 263-266.

— individuality of chromosomes and chromatin, 256, 263.

—part played by nucleus in differentiation, 266, 285.

—possiblo signiﬁcance of reducing divisions, 266.

— number of chromosomes, size of nucleus, and size of cell, 68, 267, 268.

—2méclear division not qualitative,

6 .

Bowditch: rate of growth in man, 63.

-- change of variability, 73.

Brauer : Branchipus, 22, 24.

Brooks: Lucifer, 22.

de Butfon : Preformation, 15.

Bullzt: artiﬁcial parthenogenesis, 12 .

Bumpus: change of variability in Litlorina, 71, 72.

Bunge: respiration of Ascaris, 112.

Castle : see Davenpoﬂ: and Castle.

Chabry: segmentation furrows and embryonic axes in Ascidians, 229.

—- development of isolated blastemeres in Ascidians, 229, 230.

Child : critique of Driesch’s vitalism, 292, note.

Chun : isolated blastomeres of Ctenophora, 209.

Conklin: maturation, fertilization, and development of Cynthia, 230236.

— development of isolated blastemeres in Oyntlzia, 237.

— development of pieces of gastrula in Cynthia, 238.

— streaming movements of protonlnsm. 40. 316 INDEX OF

Crampton : isolated blastomeres of Ilycmesaa, 215, 216.

— eﬁeot of removal of the polar lobe, 217.

Dareste: mechanical agitation of the Hen’s egg, 89.

— electricity, 91.

Davenport : catalogue of ontogenetic processes, 4 sqq.

— deﬁnition of growth, 58.

— rate of growth, 69.

— the role of water in growth, 58, 59, 115, 116.

- and Castle : acclimatization of eggs of Bufo to heat, 100.

Delage : causes of artificial parthenogenesis, 124.

-- number of chromosomes in artiﬁcial parthenogenesis and in merogony, 125. De Vries : importance of potassium for turgor of plant-cells, 146.

Doncaster: hybrid Echinoid larvae, 26].

Driesch: effect of light in development, 94.

— abnormal segmentation in Erhinus produced by heat, 105.

— Anenteria, produced by heat, 106.

—- segmentation made irregular by dilution of sea-water, 118.

—— pressure experiments on Echinoid eggs, 37, 38, 185, 240.

—- cell-division suppressed by pressure and dilute sea-water, 55; and by heat, 105.

—nuclear division not qualitative, 186.

— blastomeres disarranged, 187, 188.

— isolated blastomeres of Echinoids, 190, 191, 193, 194.

— potentialities of animal and vegetative cells, 193, 194, 201, 242, 243.

— fragments of blastulae and gastrulae in Echinoderms, 194.

— potentialities of ectoderm and agghenteron, and their limitations, 1 .

— development of egg fragments of Echinoids, 195, 196.

— germinal value, surface-area of larvae, and number of cells, 197199, 269.

— one larva from two blastulae, 202.

— and Morgan : isolated blastomeres of Ctenophora, 210, 211.

—2e1gg-fragments of Ctenophora, 30,

2!

AUTHORS

Drgggchz development of Myzostoma,

— isolated blastomeres and parts of larvae in Phallusia, 288, 289.

— first furrow and sagittal plane in Echinoids, 250.

— characters which depend on cytoplasm in Echinoid larvae, 261, 262.

— number of organ-forming substances in cytoplasm, 246, 284, 286.

—— theory of egg-structure, 281, 286, 292.

— reason for limitation of potentialities, 192-194, 201, 212, 242, 243, 281, 282, 284, 291.

--fate a function of position, 188, 282.

—- return of displaced mesenchyme cells in Echinus, 274.

- stimuli in ontogeny, 20, 277, 28"284.

— part played by nucleus in differentiation, 266, 284, 285.

—— equipotential and inequipotentiul systems, 176, 277, 285.

— rhythm of development, 3.

—- harmony of development, 284.

—- composition in development, 3, 285.

— self-diﬁerentiation, 284.

—- teleology, static, 286, 291, 292, 297.

— —- dynamic, 291, 292, 297.

— vitalism, 20, 289 sqq.

Edwards : physiological zero for Home egg, 102.

-- growth without differentiation, 104.

Endres and Walter : post-generation of missing half-embryo, 171.

Eycleshymer: ﬁrst furrow sagittal plane in Necturus, 168.

and

Fabricius : views on development, 13.

Fasola : electric currents, 91.

Fehling : growth of the human embryo, 59, 60, 63.

Feré : effect of sound-vibrations upon the Chick, 90.

_ ._ of light, 96.

— malformations due to high temperatures, 105. .

—- need of oxygen for the Chick, 109.

—— monstrosities produced by various chemical reagents, 18,2. INDEX OF AUTHORS

Fischel, A. : hybrid Echinoid larvae, 261.

— variability of Duck embryos, 71.

Fischel, H. : isolated blastomeres of Ctenophora, 210, 211.

-— derangement of blastomeres in Ctenophora, 211.

Fischer: artiﬁcial parthenogenesis, 124. ’ Foot : polar rings in Allolobophom,

251.

Garbowski : function of pigment ring in Strongylocentrotus egg, 192. — ﬁrst furrow and sagittal plane in

Echinoids, 260.

— grafting of blastulae fragments of Echinus, 202.

Gerassimow: size of nucleus and cells in Spirogyra, 269.

Giacomini: need of oxygen for the Chick, eﬁect of low atmospheric pressure, 109, 110.

Giardina : diﬁerentiation of chromatin in female cells of Dytiscus.

Godlewski : the respiration of the Frog’s eg, 110, 112, 113.

-— heterogeneous cross-fertilization, 262.

Graf : fusion of blastomeres, 56.

Greeley: artiﬁcial parthenogenesis produced by cold, 108.

— low temperatures and absorption of water, 108.

Grobben : Cetochilus, 22.

Groom : effect of fertilization in Cirripedes, 250.

Gigiber: regeneration in Protozoa,

54.

Gurwitsch : monstrosities produced in Amphibian embryos by chemical reagents, 120, 123.

Hacker : Cyclops, 22.

Haeckel: recapitulation, 16.

— development of fragments of blastulao of Crystallodes, 181, note. Hr;ller : preformation and epigenesis,

5.

Harvey: epigenesis, 13.

— metamorphosis, 14.

Hecker: growth of the human embryo, 62, 63.

Hansen: growth of guinea-pig embryos, 62.

Herbst : potassium, sodium, and lithium larvae of Echinoderms, 136-140.

—- signiﬁcance of monsters for origin of variatiops, 141.

317

Herbst : necessity of elements present in sea-water for normal development of Echinoid larvae, 141 sqq.

—— separation of blastomeres of Seaurchins in calcium-free sea-water,

45.

— stimuli in ontogeny, 20, 272, 273, 285.

— formation of Arthropod blastederm oxygenotactic, 114.

—— arms of Plutous due to presence of skeleton, 187, 138, 144, 149, 274, 275.

I-Ierl itzka, development of half-blastomeres of Newt, 173.

Hertwig, 0. : centrifugalized Frog’s egg, 29, 87.

—- rules for nuclear and cell division, 31, 32, 85.

— — conﬁrmed by pressure experiments, 34-36.

— gravity and Echinoderm eggs, 78.

—— insemination of Frog's egg, 79.

— cardinal temperatures for Rana

fusca. and csculenta, 97.

— monstrosities produced by high and by low temperatures, 99.

— temperature and rate of development, 100.

—— monstrosities produced in Amphibian embryos by sodium chloride, 119, 135.

— ﬁrst furrow and sagittal plane in Frog's egg, 165.

— compressedeggs: disproof of qualitative nuclear division, 34—86, 168, 169, 240.

— development of half-blastomere of Frog’s egg, 169.

— mutual interactions of developing parts, 271, 285.

Hertwig, 0. and R. : fertilization processes altered by heat and cold, 107.

— — by alkaloids, 126 sqq., 263.

His: mechanical explanation of development, 3.

—- germinal localization, 17, 158.

— the blastoderm oxygenoti-opic,114.

Hunter: artiﬁcial parthenogenesis by concentrated sea-water, 124.

Iijima: spiral asters in Nephelis egg, 40.

Jenkinson: pressure experiments on eggs of Antedon, 37, note.

— abnormalities of Frog embryos produced by various solutions not due to increased osmotic pressure, 120, 133-136. 318

Jenkinson: plane of symmetry, ﬁrst furrow and sagittal plane in Frog's egg, 165-168.

Jennings: fertilization spindle in Asplanclma, 34.

Kaestner: cardinal temperature points for the Hen‘s egg, 102.

— malformations due to low tem~ peratures, 104. '

Kant : teleology, 286-289, 292, 297.

Kastschenko: injuries to blastoporic lip in Elasmobranchs, 178.

Kathariner: gravity and the gray crescent of the Frog's egg, 86.

King : cause of differentiation of lens, 276, 276.

Knowlton : sec Lillie and Knowlton.

Kolliker: 16.

Kopsch : ﬁrst furrow and sagittal plane in Frog's egg, 165, 168.

—— eﬂ'ect of injuries to blastoporic lip, 178.

Korschelt: fusion of ova in Ophryotmcha, 202.

— nucleus of egg-cell in Dyﬁscus, 252. .

Kostanecki and Wierzejski: eﬁ'ect of fertilization in Physa, 250.

Kowalewsky: 16.

Kraus : the role of water growth of plants, 58.

Lang : effect of fertilization in Polyclads, 250.

Leibnitz : preformation, 15.

Lewis: causes of formation of lens and cornea, 275, 276. Lillie and Knowlton: eﬂect of low temperatures in Amphibia, 100. — temperature and rate of development, 101.

Lillie: effects of salts on ciliary movement, 135.

— ghysiologically balanced solutions, 1 6.

in the

— toxicity and valency, 136.

Loeb : suppression of cell-division in Echinoids and Fishes, 56, 117. -— eﬂect of light in development, 94. —the respiration of Otmolabrua and

Fundulua eggs, 111.

—— the respiration of the ova of Echinoids, 112.

— function of oxygen in regeneration of Tubular-ia head and other processes, 114, 278, 274.

-— eﬁ'ect of hypertonic solutions on Fundulus and Arbacia eggs, 117.

--exovates produced by dilute seawater, 118, 190, 194, 195.

INDEX or AUTHORS

Loeb: artiﬁcial parthenogenesis, 121, 124.

—- etfect of potassium cyanide in prolonging life of ova, 131, 132.

— eﬂect of certain salts on Fundulus embryos and on Plutei, 135.

— toxicity and antitoxicity functions of valency, 186.

-— effect of alkalies, 151.

— effect of gravity on Anmmularia, 272, 273.

-gégterogeneous cross-fertilization,

Lombardini : electric currents, 91.

Lyon : need of oxygen for the eggs of Arbacia, 112.

— action of potassium cyanide, 132.

Malebranche : preformation, 15.

Malpighi: preformation, 14, 15.

Marcacci : mechanical agitation of Hen's eggs, 90.

Mark: spiral asters in eggof Lz‘maac,40.

Mathews: artiﬁcial parthenogenesis by mechanical agitation, 90.

—— effects of atropine and pilocarpine on Echinoderm eggs, 131.

—toxicity and decomposition tension, 136.

— see also Wilson (E.B.)and Mathews.

Mencl : formation of lensin SaImo,276.

Metsclinikoif : separation of blastemeres of Oceania, 181.

-—fusion of blastulae in Mitrocoma, 202.

Minot : rate of growth deﬁned, 60.

—— change of rate of growth of guineapigs, 61.

— - of rabbits, 62, 68.

— — ofchickens, 67.

— coeﬂicients of growth, 65.

— senescence, 65.

-- increase of cytoplasm, decrease of mitotic index, 65.

— change of variability in guineapigs, 71. _ — genetic restriction, 246, 277. Mitrophanow: malformations due to low and high temperatures, 104. — necessity of oxygen for the Chick, 109.

Moore : sodium sulphate an antidote to sodium chloride, 135, 186.

Morgan : suppression of cell-division in Arbacia, 56, 118.

- gravity and the gray crescent of the Frog's egg, 86.

-— monstrosities produced by low temperatures in Ranapaluslris, 100.

— need of oxygen for the Frog's egg, 110. INDEX OF AUTHORS

Morgan :lithium salts used to produce alzlgéiormalities in Frog's eggs, 120,

— attempts to induce parthenogenesis, 124.

— number of chromosomes in artiﬁcial parthenogenesis, 125.

— artiﬁcial parthenogenesis produced by cold, 108. — first furrow, plane of symmetry, and sagittal plane in Frog's egg, 165,168.

— development of half-blastomere of

Frpg's egg ; post-generation, 170,

17 .

— development of vegetative cells of Frog’s egg, 173.

— potentialities of half-blastomeres in Teleostei, relation of ﬂrstfurrow tn sagittal plane, effect of removal of yolk, 178.

— effect of injuries to blastoporic lip, 179.

— number of cells in partial larvae of Amphioxus, 181.

— potentialities of ectoderm in Echinoids, 195.

— development of egg-fragments of Echinoids, 197.

— number of cells in partial larvae of Echinoids, 198.

— fusion of blastulae of Sphaerechinua, 201.

— and Driesch: isolated blastomeres and egg-fragments of Ctenophora, 210-212.

— micromercs of Ctenophore egg, 30.

—- characters of hybrid Echinoid larvae, 260.

Moscowski : gravity and the gray crescent of the Frog's egg, 86.

Miihlmann : prenatal growth-rate in man, 64.

artiﬁcial

Nﬁgeli : permutations of original elements in development, 286.

Pander: 16.

Pearson : variability in man, 73.

Pﬂiiger: isotropy of the cytoplasm, 18, 158.

—--inﬂuence oi’ gravity on development, 18, 78, 81-83, 168.

-- rule for direction of nuclear division, 32, 85.

Plateau : principle of least surfaces, 41, 43.

Platnerz 280.

Pott : growth of the Chick, 59, 60, 67.

319

Pott and Preyer: respiration of the Chick, 112. — loss of weight of Hen’s egg due to evaporation from albumen, 115. Preyer : rate of growth, 60.

Quetelet: change of rate of in man (weight), 68.

— — (stature), 69.

— — (other dimensions), 90.

growth

Rauber : eﬁect of reduced atmospheric pressure on the Frog’s egg, 110.

— elfect of pure oxygen on the eggs and tadpoles of the Frog, 118, 114.

Reichert: 16.

Remak : 16.

Robert : mechanics of spiral segmentation, 45-47.

— rate of growth in man, 68.

—-— change of variability, 73.

Rossi : eﬁ‘ect of electricity on Amphibian eggs, 91.

Roux : aims of experimental embryology, 13.

— ‘Mosaik-Theorie ’ of self-differentiation, 17, 158, 279, 286, 297.

— qualitative nuclear division abandoned, 19, 159, 240.

— idioplasm and reserve-idioplasm, 159, 266.

— a half-embryo from one of ﬁrst two blastomeres and post-generation of missing half, 159, 162.

— coincidence of ﬁrst furrow and sagittal plane in Frog's egg, 17, 159, 165. '

— the spermatozoon and symmetry of the Frog's egg and embryo, 80, 165, 247, 248.

— meaning of karyokinesis, 252.

— dependent diﬂerentiation, 17, 158, 277, 286.

— functional adaptation, 290.

-— speciﬁc gravity of contents of Frog’s eg, 79.

—- gray crescent of Frog's egg, 80, 165.

— inﬂuence of gravity on the Frog's egg, 85-87.

— effect of electricity upon the Frog’s egg, &c., 92.

— light and development, 93.

— segmentation of Rana esculenta, 26.

—- Frog's eggs compressed in small tubes, 39, 40.

— comparison of systems of oil drops and segmenting ova, 49-58.

— cytotropism, 55, 278. 320

Roux: cytotaxis, 55.

— cytochorismus, 45.

-— cytarme, 45, 53.

— cytolisthesis, 58.

— ‘ Framboisia’, 135.

Ruseoni : electric currents, 91.

Sachs : law of direction of cell division, 28.

Sala: fertilization processes altered by cold, 108.

- fusion of the eggs of Ascaris, 202.

Samassa: effect of pure oxygen at

pressures on the Frog's egg,

— effect of lack of oxygen on the Frog's egg, 119.

— effect of various gases on the eggs of Ascaris, 112.

—development of animal cells of Frog's egg, 173.

— Schaper: development of tadpoles after removal of brain and eyes, 175.

—- cause of differentiation of lens, 275.

Schulze, F. E. : Sponges, 22. Schulze, 0.: gray crescent of Frog’s

eg, 80, 247.

—— gravity and the Frog’s egg, 86.

—- effect of low temperatures on the Frog's egg, 100.

—— ﬁrst furrow and sagittal plane in Frog's egg, 165.

— double monsters from Frog’s egg, 171.

Seeliger : hybrid Echinoderm larvae, 260, 269.

Selenka: first furrow and sagittal plane in Echinoids, 250.

Semper: rate of growth in Limnaea, 67.

Smith: Peltogaster, 24.

Sollmann : after effects of hypertonic solutions, 124.

Spemann : development ofconstricted Newt's eggs, and embryos, 174, 175.

— causes of formation of lens and cornea, 275, 276.

Sumner: injuries to blastoporic lip of Teleostei, 178, 246.

Sutton {individuality of chromosomes in Brachyslola, 256.

Swammerdam : preformation, 14, 15.

segmentation of

Vejdovsky : unequal centrosomes in dividing pole-cells, 31.

— polar rings in Rhym.-hclmis, 251.

Vernon: rate of growth in Strongmlocmtrotus, 67, 70.

INDEX or AUTHORS

Vernon : alteration of variability in Echinoid larvae, 71, 74.

-— effect of light on Echinoid larvae, 95, 96. '

— effects of change of temperature on Echinoid larvae, 106, 107.

-— change of variability produced by heat, 107.

— and by chemical agency, 141, 156.

—poisonousness of carbon dioxide to Sea-urchin eggs, 112.

— characters of hybrid Echinoid larvae, 261.

Verworn : behaviour of Protozoa in an electric current, 93.

— regeneration in Protozoa, 254, note.

Walter, sec Endres and Walter.

Weber : law of stimuli, 272.

Weismann: qualitative division, 19, 297.

— idioplasm, and reserve—idioplasm, 159.

Weldon : growth-rate in Carcinus, 71.

— change of variability in Carcinus, 72.

— — in Clausilia, 73.

Wetzel : double monsters Frog’s egg, 172, 245.

Whitman : polar rings in Clepsine, 251.

Wierzejski, see Wierzejski, 250.

Wilson, 0. B. : malformations of Amphibian embryos, 120.

— acclimatizution to salt-solution, 136.

Wilson, E. B. : phioxus, 26.

—— segmentation of Renilla, 55, note.

— unequal centrosomes in dividing pole-cells, 31.

—pressure experiments on eggs of Nareis, 39, 213, 240.

- cytology of artiﬁcial parthenogenesis, 124.

— development of isolated blastemeres in Amphioxus, 179, 180.

—— isolated blastomeres of Oerebratulus, and fragments of blastulae, 205, 206.

— isolated blastomeres of Patella, 218-222.

—- of Dentalium, 225, 226.

—— removal of polar lobe, 224.

— effect of fertilization, 222, 223.

— development of egg-fragments, 226, 227.

nuclear

from

Kostanecki and

segmentation of AmINDEX OF

Wilson (E. B.) and Mathews : spermpath, egg axis, ﬁx-st furrow, and embryonic axes of Toacopneustes, 185, 249, 250. ‘

Windle: effect of magnetism and electricity on development, 91.

Wolff : epigenesis, 16. '

Yatgu: egg-fragments of Cerebratulus,

2 7.

Yung: effect of light on tadpoles, etc., 94.

Zeleny : egg-fragments of Cerebratulus, 206, 207.

Zelinka : fertilization Callidma, 34.

spindle in

Jnxntsonr’ Y

AUTHORS 321 Ziegler : heterodynamic centrosomes, 80.

.— formation of micromeres in Cteno phora, 209, note.

-— pressure experiments on egg gaéiinoids and Ctenophora,

— fertilization of Diplogaster, 84.

— egg and embryonic axes, 250.

Zoja : isolated blastomeres of Hydromedusae, 181, 182.

—— animal and vegetative cells of Strongylocentrotus, 198.

Zur Strassen : segmentation of Asoaiis, 81.

— fusion of the eggs of Ascaris.

s of 88, INDEX OF SUBJECTS

Abnormality, 1; produced by centi-ifugalizing, 88; by shocks, 89; by perpetual rotation, 90; by sound vibrations, 90; by electric currents and magnetism, 91; by heat and cold, 98 sqq. ; by desiccation, 115; by sodium chloride and other substances, 199 sqq., 132; signiﬁcance of forstudy ofvariation, 155, 156; produced by irregular distribution of chromosomes, 263266.

Absorption of water, sea Water.

Absorption, by cells, 5.

Abstraction of water, withdrawal of.

Acanthocystis, 48.

Acceleration of development by weak electric currents, 91; in violet light, 94, 95; at high temperatures, 97, 105; in pure oxygen, 110; by pilocarpine, 151 ; by sodium hydrate, 151.

Acclimatization, 290; to heat, 100; to hypertonic solutions, 117; to salt, 136.

Acids, 122.

Acrosome, 123.

Adaptation, 290.

Aggregates of cells, 5 sqq.

Aggregation of cells, 5.

Agitation, mechanical, 89.

Albumen, as amedium, 45, 53 ; drops of, 54 ; of Hen’s egg, 115.

Alcohol, 132.

Alcyonaria, 55.

Alkalinity, need of, 150, 151.

Alkaloids, in Amphibian eggs, 120, 123 ; on Sea-urchin eggs, 126 sqq. ; on Hen's egg, 132.

Allantois. fused with somatopleure, 9; sticks to yolk sac in eggs that are not turned, 89.

Allelomorphs, 266.

Allolobophora, 251.

Alteration of potentiality, see Potentiality.

Alteration in shape of cells, 5, 29, 48.

Alternation in direction of division in ‘ spiral’ cleavage, 25, 28.

Amblystoma, absorption of water by

see Water,

tadpole, 58; effects of cold, 100; eifect of solutions, 120; acclimatized to salt, 186; formation of lens, 275.

American Periwinkles, 71.

Ammonia, on Hen’s egg, 132; effect on nuclei, 135.

Ainnion, not formed at low temperatures, 104; folds of, 9; abnormal, 110.

Amniota, archenteron of, 8; oviduct of, 9.

Amoeboid processes in blastomeres, 53 ; movements of polar lobe, 227.

Amphiaster, see Spindle.

Amphibia, oviduct in, 9; absorption of water by tadpoles, 58, 115; inﬂuence of electric current on, 91; effect of various solutions on, 119—123, 133-135; efects of heat and cold on, 97-101 ; acclimatized to heat, 100 ; to salts, 136 ; isolated blastomeres of, 159 sqq.

Amphikaryotic, 267.

Amphioxus, club-shaped gland, 6; medullary plate, 8, 133 ; segmentation of, 22, 26, 28, 47; in certain solutions, 140; potassium necessary to adult, 147; isolated blastemeres of, 179, 180 ; double embryos, 180.

Amphitrite, segmentation of, 26; parallel polar furrows, 46; artiﬁcial partheuogenesis, 124.

Anachronism of first furrow in Frog, 161, 165.

Analytical theory of Driesch, 280286.

Anaistomoses of cell-aggregates, 6, 2 3.

Ancncephaly, 119.

Anenteria, 106.

Angles between contact surfaces in systems of lamellae, 43; in segmenting ova, 47, 53; between egg axis and first furrow in forcibly inverted Frog's eggs, 82; between plane of symmetry, first furrow, and sagittal plane, 165-167.

Animal cells, see Cells.

Animal pole, near anterior end of INDEX OF

embryo, 165 ; apical organ formed at, 244.

Anion, 136.

Annelids, germ-bands of, 5 ; nephridia and coelom, 6; nerve-ganglia of, 11; segmentation of, 25, 48; symmetry of egg and that of embryo, 241.

Antecedent conditions, 283.

Anledon, pressure, 37 ; heterogeneous cross-fertilization, 262.

Antennularia, 272, 273.

Antidotes to poisons, 135, 136.

Antitoxicity, 136.

Apical body, 123.

Apical organ of Echinoids, 144; of Pilidium, 204-206 ; of Ctenophora, 211; of Patella, 218; of Dentalium, 224-225 ; of Molluscs and Annelids, 244.

Aplysia, 46, 47.

Aplysia depilans, 30; limacina, 30, 45.

Arachnids, 24.

Arbacia, suppression of cell-division, 56, 117 ; artiﬁcial parthenogenesis, 108 ; need of oxygen, 112 ; effect of atropine, 131 ; of sodium hydrate, 151 ; hybridization experiments, 261.

Archenteron, of Echinoderms, 3, 185; in lithium larvae, 137; of Amniota, 8; in I-Iemigastrula, 159; everted, sec Exogastrula; in double monsters of Frog, 172; tip of, removed, 195 ; of partial larvae, 198, 205, 221.

Architecture of nucleus in Mosaic Theory, 19.

Area vasculosa, 110.

Arenicola, segmentation of, 26, 30, 47 ; effect of ions on larvae, 135. Arm, rate of growth of, 70; varia bility, 74; of Pluteus, see Pluteus.

Arrangement of systems of drops or bubbles, 43, 44.

Arrest of development at low temperatures, 97, 99, 102, 104.

Arrhenokaryotic, 267.

Arthropods, blastoderm of, 5, 114; muscles of, 5 ; somites of, 11 ; nerve-ganglia of, 11 ; segmentation of, 24.

Artiﬁcial parthenogenesis, see Parthenogenesis.

Ascaria lumbricoides, 258.

Ascaris me-galocephala, segmentation of, 27, 28, 34, 47; cell-movement, 29; simultaneous division of related cells, 31 ; eggs poisoned by oxygen, 112 ; fusion of eggs to form double monsters, 202; symmetry of egg

SUBJECTS 323

and embryo, 245; diminution of chromosomes, 252, 253, 256-258; individuality of chromosomes, 256; dispermy, 257.

Ascaris nigrorenosa, 33, 34.

Ascidians, segmentation of, 24, 26, 47, 54, 229,235 ; eﬁ'ect of solutions, 140; isolated blastomeres, 230, 237 ; symmetry of egg and that of embryo, 229, 235, 245.

Ascidiella, egg- and embryonic axes, isolated blastomeres, 229, 230.

Asplanchna, unequal asters, fertilization spindle, 34, 48.

Asters, spiral, 40; unequal, 80; induced by salts, 124, 125; altered by alkaloids, 128-130.

Aster-ias, ﬂattening of blastomeres, 45; artiﬁcial parthenogenesis, 108 ; effect of atropine, 131 ; exogastrulae, 140 ; necessity of sulphate, 144; necessity of potassium, 146; blastomeres without magnesium fall apart, 148; blastula cut up, 194; tip of archenteron removed, 195 ; coelom-sacs removed, 195 ; ochracea, heterogeneous cross-fertilization, 262.

Asteroids, segmentation of, 24, 26, 28, 47, 64.

Atmospheric pressure as affecting growth, 2; reduced, 109, 110; increased, 114.

Atrium, 229.

Atropine, 131.

Attachment of cells to other bodies, 5.

Auditory vesicle of Vertebrates, 6.

Axes of embryo, see Embryonic axes.

Axial organs, correlations between, 75.

Axis cylinder, 273.

Axis of eg, see Egg axis.

Axolotl, 119.

30;

Bacterial toxines, 132.

Barium, 150.

Bases, 122.

Ba/tmchus, 178.

Beginnings, knowledge of, 289, 298.

Belgians, growth of, 63.

Benzole, 42.

B6759, pressure experiment, 39; isolated blastomeres and egg-fragments, 209-211.

Bilateral egmentation, see Segmentation.

Bilateral symmetry, see Symmetry.

Bile-capillaries, 6.

Bipinnaria, necessity of sulphate, 144.

Y2 324

Birds’ eggs, effect of electric current on 91.

Blastema, metanephric, soc Kidney.

Blastococl, see Segmentation cavity.

Blastocyst of Mammalia, 2, 6, 7, 115.

Blastoderm, of Arthropods, 5, 114, 273 ; of Cephalopods, 6 ; of Sauropsida, 6; growth of, without formation of primitive streak, 104; growth of oxygenotropic, 114, 273 ; isotropic, in Teleostei, 179.

Blastomeres, separate, in Triclads and Salps, 5; in Oceania, 181 ; grouping of, in Salps, 11 ; size of, 29 ; separation of, 45; before division, 45; in calcium-free sea-water, 45, 150; without magnesium, 148 ; ﬂattened against one another, 45, 53 ; movements of, 29, 42, 53.

— ‘isolated’ development of, in the Frog, 159-161, l69—l71,173 ; in the Newt, 173 ; in Fishes, 178 ; in Amphioxus, 179, 180; in Coelenterata, 181, 182; in Echinoderms, 190 sqq.; in Nemertines, 204, 205 ; in Ctenophora, 209-211 ; in Mollusca, 215 sqq. ; in Ascidia, 229, 230, 237, 239 ; reason for diﬁerences in behaviour of, 241-244.

Blastopore, dorsal lip of, in Frog, 80; in forcibly inverted eggs, 82, 84; rim of, thickened by heat, 99 ; fails to close, 99, 119 sqq., 133; in half-blastomere of Frog, 170; in double monsters of Frog, 172; injury to lip of, in Fishes, 178; in Ascidia, 235; in Molluscs and Annelids, 244.

Blastula, of Echinoderms, 2, 6, 115, 185 ; in lithium salts, 137 ; opaque, without potassium, 146 ; opaque, in neutral media, 150; crumpled, without carbonate, 148 ; of Coelenterates, 6 ; of Sponges, 6 ; pieces of, in Siphonophora, 181 ; in Echinoids, 194 ; in Nemertines, 206 ; long-ciliated, from animal cells of Echinoids, 192; united to form one embryo, 201, 202.

Bleu de Lyon, 94, 96.

Blood,the first to be formed (Harvey), 14.

Blood-vessels, growth of, 5; branching of, 6 ; lumen of, 11 ; distended at low temperatures, 104 ; muscles of, 278.

Blue light, 94-96.

Balina, 210.

Bombinator igneus, Pﬂilger’s experiments on eggs, 82, 88.

INDEX OF SUBJECTS

Bone, 2 ; and tendon, 5.

Bonnetty machine, 91.

Boston school-children, growth of, 63.

Boys, sea Human being.

Brachystola, 256.

Brain remains open solutions, 119 sqq.

Branchipus, 22, 24, 47.

British Periwinkles, 71.

Bromides, 128, 132,133, 134.

Bromine, 141, 145.

Budding, a form of development, 1 ; nucleus in, 284, 285.

Buds of Doliolidae, 5; of plants, 11 ; of Vertebrate limbs, 11.

Bu,/‘o, absorption of water by tadpoles, 58; acclimatization to heat, 100; rate of growth affected by temperature, l02 ; effect of solutions, 120.

Butyrate of sodium, 140.

in certain

Caesinm, 146.

Caffein, 120.

Calcium, eﬂ‘ect of absence of, 45, 148 ; effect on cilia and muscles, 135, 150; acetate, 49; phosphate, 58, 149; chloride, 123, 133; nitrate, 135; hydrogen phosphate, 148, 150; carbonate, 148, 150; seawater without, for isolating blastemeres, 45, 150, 190.

Callidina, 34.

Cane-sugar, effect of, on Frog's eggs, 120, 121, 133, 185, 136; artiﬁcial parthenogenesis, 123.

Capillaries, 5; anastomoses of, 6; absence of, without oxygen, 110. Capillarity, in cell-division, 41 sqq.,

145.

Cupitellu, 26.

Carbon dioxide, fatal to Frog‘s eggs, 110; to Teleestean ova, 111; to Echinoid eggs, 112 ; not fatal to Ascaris eggs, 112; quantitative determination of, 112 ; excess pro~ duced by Chick in pure oxygen, 112; neutralized by hydroxyl, 151.

Carbonate, necessity of, 148-150.

Carbon-ion, see Carbonate.

Carcinus mocnas, growth-rate, 71 ; decrease of variability, 72, 74.

Cardinal temperature points, sec Temperature.

Cartilage, 2.

Catamenia, 294.

Cation, 136, 145.

Causal, see Cause; causal harmony, 284, 285, 292.

Cause, explanation in terms of cause and eifect, 12, 20, 286-289, 290, 292, INDEX OF SUBJECTS

296, 297, 298; formal, 13, 293, 296 ; eillcient, 13, 296, 297, 299; final, 20, 296, 297, 299.; material, 294, 296, 299.

Cavities, development of, 11.

Cavity perivitelline, 79 ; and sagittal plane in Frog, 165-168; segmentation, see Segmentation cavity.

Cell~plate, 41.

Cells, animal and vegetative. differences in potentiality, 191, 204206, 242.

—— division of (see also Segmentation), a factor in development, 2, 22 sq.; not necessarily a process of differentiation, 22, 166, 245, 246, 279; meridional, 22, 32; in ‘compressed eggs, 34, 36; equatorial, 22; latitudinal, 22, 32; in compressed egg, 34, 36; dexiotropic, 26, 28; laeotropic, 26, 28; alternation of direction of, 25, 28; direction of, 29, 54; depends on that of nucleus, 31, 48, 55; pressure experiments on, 34-39; and yolk, 24, 29, 48; and surface-tension, 41 sqq.; influence of gravity on, 78 ; inﬂuence of electricity on, 93 ; unequal, 29, 30, 168; rate of, 29, 261; suppressed, 55, 56, 106, 117; simultaneous division of the ovum into several cells, 55, 56, 118, 128, 130, 264, 265; in plant-cells, 41; qualitatively unlike, 158, 286; power of, restored in fertilization, 252.

— follicle, of Tunicatn, 5; and cell aggregates, movements of (Davenport), 4 sqq. ; movements of, in sogmentation, 29, 42, 222; number of, in partial embryos or larvae: in the Newt, 173; in Amphiorcus, 181; in Coelenterata, 182; in Echinoids, 197, 269; in Nemertines, 206; separation of, 45, 105, 145.

Cellulation, 162.

Cellulose, 58.

Centrifugalized eggs, 29, 87, 88.

Centrosomes, heterodynamic, 30 ; in artiﬁcial parthenogenesis, 124,125; direction of division, 40; division prevented, 265, 267.

Cephalopods, blastoderm of, 6; segmentation of, 27 ; symmetry of egg and axes of embryo, 245.

Cerebratulus, isolated blastomeres and egg-fragments, 204-208.

Cestoda, 28.

Cctochilus, 22.

Chaetopods, artiﬁcial pertinenc 325

genesis, 124; pressure experiments, 39, 213.

Ghaetopterus, 214. .

Chemical agents, effect of various, 126 sqq. ; need of, 142-152.

Chemically equivalent solutions, 140.

Chemotactic, 55, 273.

Chest-girth, change in rate of increase of, 70.

Chick, Harvey, 14; Malpighi, 15; do Buifon, 15 ; percentage of water in, 59, 60 ; decline of growthrate, 67; effect of electric current, 91 ; inﬂuence of light of various colours, 96 ; respiration of, 109, 110, 113; effects of heat and cold, 102-105 ; eﬁects of poisons, 132.

Chloral hydrate, effect on fertilization, 128-130.

Chlorates, 145.

Chlorides, 123, 133; see also Sodium chloride, and Chlorine.

Chlorine, necessity of, 145.

Chloroform, 42; and vitelline membrane, 129.

Chondrin, 58.

Chorion, of Teleostei, 35.

Choroid, 273; choroid ﬁssure, 273.

Chorophilus, 120 ; acclimatized to salt, 136.

Chromatin, the vehicle of inheritance, 252; individuality of, 256,

265,266. Chromosomes, number of, varied artiﬁcially, 267; in merogony,

267; in artiﬁcial parthenogenesis, 125, 267; and cell-volume, 268; and size of nucleus, 268; and number of cells, 268; irregular distribution of, in poisoned eggs, 128-130; in dispermic eggs, 253, 265; individuality of, 252, 256; diminution of, 252, 256-258.

Cilia, apical tuft of, in Plutei. without sulphate, l44; in Pilidium, 204-206.

—' movement of, 135; independent of sulphates, 144; without magnesium, 147 ; in neutral media, 151.

Ciliated ring of Pluteus, without sulphate, 144; without calciumcarbonate, 148; of Mollusca and Polychaeta, 213, 220.

Cirrhipedes, segmentation of, 22, 24 ; fertilization and egg-structure, 250.

Olausilia, decrease of variability, 74.

Cleavage, see Segmentation.

Clepsine, germ-bands of, 6; rings, 251.

Clove-oil, 54.

pola r326 INDEX OF

Clymanella, 46.

Clytia, 181.

Coca(i)n, effect on fertilization, 12818 .

Coeﬁlcient of growth, 65; of variability, 7l, 72, 78; of correlation, 75.

Coelenterata, skeleton in, 2; unipolar immigration in, 11 ; segmentation of, 22, 28, 47, 54; development, of isolated blastomeres, 181.

Coelom, of Echinoderms, 8; of Annelids, 6; epithelium of, and gonads, 7; lumen of, 11; sacs removed, 195.

Cohesion of blastomeres, 45, 148, 150.

Cold, effects of, on segmentation and development, 99 sqq.; on fertilization, l07, 108; arrest of development due to, 97, 99, 102, 104; together with potassium cyanide, 132; produces artiﬁcial parthenogenesis, 108, 124; segmentation of an isolated blastomere becomes total, 216.

Combination of original elements, 286.

Composition, organs formed by, 3, 277, 285.

Concentration of material of blasteporic lip in axis of embryo, 178; of sea-water, 124.

Concrescence of lips of blastopore, 178; of layers, sea Cells, movements of.

Conditions of development, see Development.

Cone of entrance, 128.

Connective tissue, 273.

Consciousness, of the process, in development, 20, 291, 297.

Constriction, see Cells, movements of.

Contact stimuli, 137, 272, 275-277.

Contractility of cytoplasm, 130, 131.

Contraction of muscles, inhibited by magnesium, 135, 148 ; sodium necessary for, 135, 145 ; potassium necessary for, 147.

Co-ordination, 284.

Copper sulphate, 96.

Cornea, causes of formation, 275, 276.

Corpus luteum, 5.

Corpuscles, concentric, of thymus, ll.

Correlation, 75; decrease in, 75, 277; between symmetry plane, first furrow and sagittal plane, 167.

Cosmic process, 298.

Costae of Ctenophora, 211, 212.

SUBJECTS

Cotylorhiza, necessity of potassium, 146.

Crab, see Carcinuﬁ

Cramgon, 24.

Gnepidula, segmentation of, 26, 30 ; asters and protoplasmicmovement, 40; ﬂattening of blastomores, 45; parallel polar furrows, 46.

Crescent, gray, see Gray crescent.

Crinoids, 22.

Cross-fertilization, see Hybrid.

Cross furrow, sec Polar furrow.

Crustacea, segmentation of, 22, 24, 47; eggs of, and gravity, 78; structure of egg, and embryonic axes, 245.

Orystallodes, 18].

Ctenolabrus, cell-division suppressed, 56, ill; ﬁrst furrow and sagittal plane, 178 ; need of oxygen for, 111, 262.

Ctenophora, segmentation of, 26, 208; micromeres, 30; pressure, 39; inﬂuence of gravity on eggs, 78; micromercs displaced, 211 ; isolated blastomeres, 209, 210; cg,-;—frug~ ments, 212; effect of fertilization, 250.

Gyclas, 26.

Cyclops, 22.

Cynthia, development, 230; isolated blastomeres, 237 ; symmetry of egg, 232, 250.

Cytarme, 45, 53.

Cytastors, 125.

Cytochorismus, 45.

Cytolisthesis, 53.

Cytoplasm, organ-forming substances in the, 19, 208, 212, 213, 217, 224, 225, 228, 233, 236, 240, 241, 246, 280, 286.

-—nucleus and, 31, 131; ratio between, 65, 269; inter actions of, 266, 283, 284.

-— increase of, in differentiation, 65, 66 ; ascent of, in inverted eggs, 84 ; receptive to stimuli, 283; paralysed by poisons, 127; contractility of, 130, 131; determines diminution of chromosomes, 257.

Cytotaxis, 55.

Cytotropism, 55, 273.

Daphnella, 24.

D_aphm'a, 24.

Darkness, Fundulus embryos, 94 ; tadpoles of Frog, 95 ; eggs of Limmwa, 95 ; Echinoid larvae, 96.

Death-rate, selective, 74 ; in various lights, 94-96; in desiccators, 115; of heterogeneous hybrids, 262. INDEX OF SUBJECTS

Decay, senile, 65. , Decapoda, segmentation of, 24. Decline of growth-rate, 61-71. Decomposition, tension, 135. Defects, local, due to local injuries, 17, 159.

Degeneration, gray, of medullary groove, 119, 135.

Dehydration, see Water, withdrawal of.

Dendritic ﬁgure, 130.

Dentalium, development, 222-224 ; removal of polar lobe, 224; isolated blastomeres, 225, 226 ; egg-fragments, 226.

Dependent differentiation, see Differentiation.

Descemet's‘ membrane, 276.

Desiccation of Hen‘s egg, 115; of Frog’s egg, 123.

Detachment of parts, 8.

Determinant judgement, 287, 288.

Deutoplasm, see Yolk.

Development, deﬁned, 1 ; component factors of, 2; speciﬁc, 1, 15, 17, 280 ; and inheritance, 1 ; rhythm of, 3 ; external conditions of, 18, 20, 152, 155-157; a predetermined process, 19; internal conditions of, 20, 158 sqq., 279286; rate of, inﬂuenced by temperature, 100, lol ; without potassium, 146, 147; mgnesium, 148; and calcium, 150; in partial larvae, 200, 204; acceleratcdhsea Acceleration ; retarded, see Retardation ; arrested, see Arrest ; suspended, see Suspension.

Deviation, standard, 71.

Dexiotropic division, see Cell.

Dextrose, Frog's egg, 121 ; Hen's egg, 132.

Dicephalus tetrabrachius, 175.

Dicyemidae, 11.

Differentiation, see also Cytoplasm and Nucleus; defined, 2; histological, 3; self-differentiation, 17, 18, 158, 240, 286; increase of, 76, 176, 277, 284 ; dependent, 18, 158, 162, 277, 286; progressive, 247; and cell-division, 22, 166, 213, 236, 245, 246, 279; and growth, 62, 65, 279; and increase of cytoplasm, 65 ; cytoplasm in, 19, 208, 212, 213, 217, 225, 227, 237, 240, 241, 246, 261, 266; nucleus in, 251 sqq.; interactions of the parts in, 271277.

Dilution of sea-water, as a means of preventing cell-division, 55; ex 327

ovates produced by, 118, 189, 190, 194; causes irregular segmentation, 118, 187, 194.

Dimensions, see Size.

Diminution of chromosomes, 252, 256-258.

Diphtheria toxine, 132.

Diplogasler, rotation of fertilization spindle, 34; egg- and embryonic axes, 250.

Diplokaryotic, 267.

Diprosopus triophthalmus, 175.

Directive stimuli, 162, 272.

Discocelis, segmentation of, 25, 80, 47; crossed polar furrows, 46; fertilization and egg-structure, 250.

Disintegration of epithelium, 119, 120, 135.

Dispermy, in Ascaris, 257; in Echinoids, 263--265.

Dispersion of cells, 11.

Division of nucleus and cell a factor in development, 2; of nucleus, sea Nucleus; of cell, sea Coll; of oildrops, sec Oil-drops; of centrosomes, see Centrosomes.

Doliolidae, buds of, 5.

Dorsal lip of blastoporc, sec Blastepore.

Double monsters, see Monsters.

Dreissensia, 45, 47.

Drops of ﬂuid, 42, 43; of oil, 43, 49 sqq.

Duck embryos, decline of variability in 71.

Ducts, see Gland, Segmental; lumen of, 11.

Duplicitas anterior, 175.

Dytiscus, 258.

Earth-worm, ectoderm of, 3; nephridia, 5.

Echinoderm, blastula, 2, 110, 185; spicules of larva, 2; archenteron divides into gut and coelom, 3; mesenchyme, 5; gastrula, 5; segmentation of, 22, 24, 26-28, 54, 188 ; pressure experiments on eggs, 39, 185, 186; inﬂuence of gravity on eggs, 76; artiﬁcial parthenogenesis, 123 ; eifects of sodium, potassium, lithium, on larvae, 137-140 ; blastomeres disarranged, 187 ; blastomeres isolated, 190 sqq.; blastulae and gastrulae cut up, 195; egg-fragments, 195-197; one embryo from two blastulae, 201, 202; number of cells, rate of development and, size of partial larvae and their organs, 198-200. 328

Echinoids, egg, segmentation of, 22, 27 ; position of nucleus in egg, 81 ; normal development of, 183-185; pressure experiments on, 37, 88, 185, 186 ; blastomeres disai-ranged, 187, 194; isolated blastomeres, 190 sqq. ; egg-fragments, 195-197; fragments of blastulae and gastrulae, 199, 195; one embryo from two blastulae, 201, 202; larvae, inﬂuence of light on, 95. 96 ; require oxygen, 112; spicule cells in, 114; inﬂuence of potassium, sodium, and lithium, 137-140; partial larvae, size of, rate of development of, number of cells of, 198-200; hybrids, 258-262.

Echinus, pressure experiments, 37, 38, 185, 186; suppression of celldivision, 55; inﬂuence of light, 94; ciliary movement in larva, 135; potassium, sodium, and lithium larvae, 137; necessity of certain elements for, 142 sqq.; blastomeres disarranged, 187, 188, 194; blastomeres isolated, 190 sqq.; segmentation of egg-frag ments, 195-197; blastula cut up, 194; grafting of blastula fragments, 202 ; size of partial larvae, 199, 201; hybridization experiments, 258-262 ; number of chromosomes varied, 267.

Ectoderm, of Earthworm, 3; growth of, in Amphioacus and Vertebrates, over medullary plate, 9; of Simmculus, 9 ; of lithium larvae, reduced, 139, 140; of Echinoid, gastrula, separated,I95 ; ofpartial larvae,198.

Ectoproctous Polyzoa, segmentation of, 22.

Elﬁcient cause, see Cause.

Egg axis, how determined, 79, 163, 183, 204, 208, 232, 249, 250, 280; and gravity, 78; and embryonic axes, 81, 165, .166, 174, 204, 209, 229, 238, 236, 244, 281; and segmentation furrows, 81, 85, 229, 235; secondary, in inverted eggs, 85.

Egg-fragments, enucleate, fertilized, sea Merogony; segmentation of, see Segmentation.

Elasmobranchs, lower layer cells, 5 ; segmental duet, splitting of, 6 ; yolk-nuclei, 88; injuries to germring. 178.

Elastic plates and tubes, layers of Chick compared to, by His, 3.

Electricity, 91 sqq.

INDEX OF SUBJECTS

Elementary organs, see Organs.

Emboitement, 14.

Embryology, experimental, 1, 18; descriptive. 12; comparative, 12, 16 ; aims of, 13, 156, 157, 297, 298.

Embryonic axes, and gravity, 78; and symmetry of egg, 8], 165, 166, 174, 204, 209, 229, 233, 236, 244, 266, 281 ; inﬂuence of light on, 93.

Embryonic plate, of Mammals, 9.

Embryos, double, 201, 202; partial, see Blastomeres, isolated, and Eggfragments.

End, knowledge of the, 286, 289, 296, 297, 299.

Endoderm of Echinoid larvae, separated, 195.

End—organ and nerve, see Nerve.

Energy, life not a form of, 291.

English, growth of, 63.

Entelechy, 291, 297.

Environment, physical and chemical, adaptation of developing organism to, 157.

Epidermis of Earthworm, 3.

Epigenesis (Harvey), 13, 15; (Wolff), 16; not a theory, 17, 19.

Epithelium, degeneration of, 120.

Equatorial division, 24.

Equilibrium, osmotic, 118.

Equimolecular solutions, 140.

Equipotential, 176, 277.

Essential oils, 132.

Ether, 132.

Ethical, 289.

Eucharis, 209.

Evaginations, see Cells, movements of.

Eversion of gut, see Exogastrula.

Evil, problem of, 299.

Evolution, theory of (of the individual), 14 ; modern form of, 19, 158 ; (of the race), 16.

Excretory, see Kidney.

Exocoefus, 178.

Exogastrula, in lithium salts, 137; without sulphate, 143; of Ascitlitt, 237.

Ex-ovates, of A rbacia, 118.

Experimental method inembryology, 1, 13, 297, 298.

External conditions of development, see Development.

Eyes, width of face between, growth of, 70.

Fat, 58. 1

Fate a function of position, 188.

Ferments, in nucleus, 288.

Fertilization in Ascaria nigrorenosa, 38; altered by heat, 107, 108; INDEX OF SUBJECTS

altered by alkaloids, 126 sqq. ; independent of sulphates, 144; without potassium, 146; without magnesium, 147 ; in neutral media, 151 ; alteration of egg structure in, 80, 163, 222, 232, 247251 ; two processes involved in,252.

Film, at surface of blastomeres, 45, 64, 150; of drops of ﬂuid, 42, 45, 54.

Final cause, sea Cause.

First furrow, coincidence of, with sagittal plane, see Furrow, first.

Fissure, choroid, 273.

Flattening of blastomeres against one another, 45, 53.

Flexure, cranial, 110.

Folds, medullary, see Medullary.

Follicle cells, in Tunicata, 5.

Follicles, of hair, 7.

Food, growth dependent on, 2, 58, 59.

Foot, rate of growth, 70. Form, production of, 1 ; nucleus necessary for, 252, 254.

Formal cause, sec Cause.

Formate of sodium, 145.

Formative stimuli, 273. Fovoa germinativa, 79.

Fragments of egg, sec Egg-fragments ; of hlastulae, and gastrulae, see Blastulae, Gastrulae.

Framboisia, 135.

Freedom, 289, 299.

Fresh water, Funduluseggs in, 117,136.

Frog, ovarian and uterine egg, 79; eﬁect of fertilization, 163; eggs forcibly inverted, 18, 78,158; inﬂuence of gravity upon. 18, 78 sqq. ; eggs placed on vertical wheel, 85 ; eggs kept in constant motion, 86 ; egg centrifugalized, 29, 87, 88 ; pressure experiments, 34-37, 40, 168; absorption of water by tadpoles, 58; eggs exposed to electric currents, 92, 93; eggs, influence of light on, 94 ; eggs, inﬂuence of heat on, 97-99; of cold on, 99-100; eggs, respiration of, 110-112; effect of sodium chloride on eggs, 119, 120, 276 ; of other substances, 120, 121 ; acclimatizod to sodium chloride, 136 ; ﬁrst furrow and sagittal plane, 17, 159, 165-168; half—embryo from half-blastomere, 18, 159 sqq., 169-171 ; double monsters, 171, 172; development of animal and vegetative blastomeres, 173 ; formation of lens,275,276 ; of cornea, 276, 277.

Frontal breadth in Carcinus, growth of, 71 ; variability of, 72.

329

Fuchsin, 94.

Functional adaptation, 290.

Fundulus embryos, in light and darkness, 94 ; need of oxygen for, 111, 114, 274 ; in fresh water, 117 ; eifect of ions on, 135, 136; in sodium chloride, 117; in magnesium chloride, 276; first furrow and sagittal plane, 178; injuries to blastopore, 178.

Furrow, first, and sagittal plane in Frog, 17, 159, 165-168 ; in Necturus, 168; in Newt, 175; in Fishes, 178; in Ascidia; 229, 233, 235 ; in Echinoids, 250 ; and plane of symmetry, 167, 233; and horizontal plane in Newt, 175 ; first and entrance point of spermatozoon, 40 ; in Frog, 247, 248; in Toazopneustes, 250.

Furrows, polar, parallel, and crossed, 25, 26, 46,47; in Roma, 53 ; in systems of oil drops, 50.

Fusion of cell-aggregates, see Cells, movements of; of larvae to form double monsters, 137, 201 ; or one larva, 202.

Gulvanotaxis, 272.

Galvanotropism, 272.

Gammams, 24.

Ganglia, see Nerve.

Ganoids, 26.

Gastrula, of Echinoderms, 5; opaque in neutral media, 151 ; of animal and vegetative cells, 191194; pieces of, in Echinoids, 194 ; of Ascidia divided, 238.

Gastrula wall of lithium larvae, 138-140.

Gastrulation, need of sodium for, 145 ; of isolated blastomeres, 191-194.

Gemmule of Sponges, 5, 11.

Genetic restriction, 246, 277; see also Potentialities, limitation of.

Geophilus, 4.

Geotropism, 272.

Germ-bands, 5; of Clcpsim-, 6; from teloblasts, 29 ; of Crustacea, 34. Germ-cells, liberation of, 11 ; the vehicles of inheritance, 1, 252; of

Ascaris, 253.

Germ-layers, 2.

Germ-ring, injuries to, 178.

Germinal localization, His's principle of, 17.

Germinal value, and loss of potentiality, 181; and number of cells, 182, 197, 206, 269; and size of 330

larvae, 198, 199, 269; and rate of development, 200, 204.

Geryonia, 181.

Gill slits, abnormal, 104.

Girls, see Human being.

Glands, growth of necks of, 5, 273; branching of, 6; club-shaped, of Amphfoxus, 6.

Globules, of Wolff, 16.

Glycerine, 132.

Gonads, from coelomic epithelium, 6.

Grafting of pieces of blastulae, 202.

Gravity, inﬂuence on segmentation of Frog's egg, 18, -78, 158; influence on development in general, 78 ; Plliiger’s views, 83 ; directive inﬂuence of, experimentally eliminated, 85 ; replaceable by a contrifugal force, 87; eifect on Anfennu(aria, 272, 273; on Serlzdaria, 272.

Gray crescent of Frog's egg, 80; said to be due to gravity, 87; caused by spermatozoon, 247, 248.

Gray degeneration, see Degeneration.

Gray patch in inverted eggs, 85, 87.

Green light, 94-96.

Growth, a factor in development, 1, 279; conditions of, 1, 58, 59; local inequalities of, 2, 3, 11; in length, 5; of spherical and plane surfaces, 6; defined, 58; without differentiation, l04; dependent on absorption of water, 58, 59, 68, 115, 116,146; and other substances, 58 ; change of rate, 61-71; change of variability during, 71-75; change in correlations during, 75, 76; rate of, deﬁned, 60 ; variable, 74; inﬂuenced by temperature, 101.

Guinea-pig, change of rate of growth, post-natal, 61, 62, 65; pre-natal, 62; decline of variability in, 71.

Gut, muscles oh 5 ; of Echinoderms, 3; imaginal, of Insecta, 5; of Pluteus, in potassium, 137 ; without sulphate, 143; without magnesium, 147 ; without potassium, 146.

Haemoglobin, not formed in absence of oxygen, 110.

Haemorrhage, due to cooling, 104.

Hair, follicles, 7.

Half-embryo, depends on position of egg, 170, 171.

Hand, rate of growth, 70; variability, 74. Harmony, causal, 284, 285, 292; of composition, 285.

Head, length, rate of growth, 70;

INDEX OF SUBJECTS

variability, 74; correlations with other parts, 75; breadth, rate of growth, 70; variability, 74; correlations with other parts, 75.

Heart, the first organ to be formed, according to Aristotle, 13, 294; -beat of Chick at low temperatures, 104; halves of, remain apart, 104; distended, 104; beat of Fundulus in hypertonic solution, 117.

Heat, 97 sqq., as affecting growth, 59; effect of, on Frogs’ eggs and embryos, 97-99; acclimatlzation to, 100; effect on Sphaerechinus ova, 105, 106, 187; on fertilization, 107, 108 ; stimuli, 272.

Heliotropism, 272.

Hemiblastula, 159; superior, 161.

Hemicrania, 119.

Hemi-embryo laternlis, anterior, 161.

Hemigastrula, 159.

Hemikaryotic, 267.

Hemimorula, 159.

Hemitheria anteriora, 18.

Hen’s egg, axis vertical,‘-78 ; need for being turned, 89; exposed to shocks, 89; perpetually rotated, 90; exposed to sound-vibrations, 90 ; placed between poles of a magnot, 91 ; inﬂuence of light on, 93, 91; loss of weight of, 115; effect of poisons on, 132.

Heterocentron, 58.

Heterodynamic centrosomes, 30. _

Heterogeneity of ovum, 2, 157; and homogeneity, 298.

Heterogeneous hybridization, 262.

Hips, rate of growth, 70. Holoontoblastia, 139.

Holothurians, segmentation of, 22.

Homologies, sought for in descriptive embryology, 12.

Human being, decline of growth—rate after birth, 63, 64, 65 ; before birth (rte Human embryo) ; rise of growth-rate at puberty, 65 ; decline of variability, 73; increase at puberty, 73; decline of correlations, 75. ‘

Human embryo, percentage of water in, 59, 60; decline of growth-rate, in weight, 63; in stature, 68; of organs, 70.

Humid atmosphere, necessary for Hen’s egg, 115.

Hybrid Pluteus from enucleate eggfragment, 253, 258-260; Plutei, various, 262; heterogeneous, 262.

Hz/dractinia, 47.

159, 171 ; INDEX OF

Hydranth, see Hydroids.

Hydrochlorate of morphine, 128-180.

Hydrogen, effect of, 109-111, 132.

Hydroids, growth of tolons and bydranths, 5 ; attachment of, 272 ; from isolated blastomeres, 181.

Hydroxyl, 150, 151.

Hyoid of tadpoles, enlarged, 114.

Hypertonic solutions, see Osmotic pressure.

Hypotonic solutions, pressure.

see Osmotic

Idioehromatin, 258.

Idioplasm, 159.

Idioplasma, in the nucleus, 19.

Ilyamassa, segmentation, 26, 214, 215; isolated blastomeres, 215-217 ; removal of polar lobe, 217.

Immigration, see Cells, movements 0 1

Immunity, 290.

Increase of mass, 58, 115; of selfdifferentiation, 75, 176, 277, 284; of structure, 2, 279.

Increments, percentage of growth, 60.

Index, mitotic, 65; of development, 103.

Individuality of chromosomes and chromatin, 252, 256, 265, 266.

Induction, 282, 283.

Inequalities, local, of growth, see Growth.

Inequipotential, 176, 277.

Inheritable qualities, represented by units in the germ, 19, 159, 246, 247, 279, 280.

Inheritance, material basis of. 1, 19, 252; Aristotelian explanation of, 294 ; mechanism of, 1, 19, 280.

Injuries, local, the cause of local defects, 17, 159.

Insects, gut of, 5; pupae, phagocytosis in, 5; segmentation of, 24; polar nuclei, 29 ; eggs of, and gravity, 78 ; symmetry of egg and embryo, 245.

Insemination, ﬁrst elfoct of, in Frog, 79.

Insufficiency of material, sec Potentialities, limitation of.

Ingeractions of the parts, 271-277,

84.

Internal conditions of development, see Development.

Interruptions of continuity, see Cells, movements of.

Invaginations, see Cells, movements of.

SUBJECTS 331

Investment by cells, see Cells, movements of.

Iodides, 183.

Iodine, 141, 145.

Ion-proteids, 136.

Ions, 135, 186, 140.

Iris, 276.

Iron, 141.

Irregular distribution of chromosomes, see Chromosomes.

Irregularities of segmentation, see Segmentation.

lschnocliilon, 25, 46.

Isobilateral segmentation, see Segmentation.

Isopoda, 24. '

Isotonic solutions, 120, 123, 124, 140, 142.

Isotropy of egg, 18, 158, 202, 280; of blastoderm, 179.

J elly of Frog’s spawn, 42, 43, 79. Judgement, Kritik of Tcleological, 286-289.

Karyokinesis,without calcium, 150 ; signiﬁcance of, 252.

Karyokinetic plane, 185.

Kidney, larval, of Lamellibranchs, 5 ; tubules, branching of, 6; anestomoses of, 6; tubules (mesonephric).fusionwithvasaefferentia, 6; tubules in metanephric blastema, ll.

Labrax, 145.

Lacertilia,inner wall of pineal vesicle, 9.

Laeotropic division, see Cell-division.

Lamellae, systems of, 41 sqq.

Lamellibranchs, larval kidney of, 5.

Laodice, 181. ,

Lappets of Pilidium, 204, 207.

Larva, of Sponges, 6; of Dicyemidae, 11; of Coelentcrates, 6 ; characters of, developed early, 3; partial, see Blastomeres, isolated, and Egg-fragments.

Latitudinal division, 22, 24, see also Cell-division.

Law, Biogenetic, 12, 16; of alternation of direction of cleavage in ‘spiral’ eggs, 25; Sachs’, 28, 40; Weber's, 272.

Laws of causation, 13, 285, 298.

Leg, rate of growth of, 70.

Lens, outer layerof, 7 ; absent, in certain solutions, 134; causes of formation, 275, 276. 332

Lepidonotus, 25, 46.

Lepidosteus, 26.

Light, general effect on developing organisms, 93 ; on fern prothallus, 272; on Serpula, Sormlarella, 272; effect on direction of embryonic axes, 93 ; of various colours on eggs of Echinus, Planarbis, 94 ; Rana, 94, 95 ; Trout, 95 ; Limnaea, 95; Echinoid larvae, 96 ; promotes oxidation, 112.

Limax, segmentation of, 25, 26, 30; spiral asters, 40.

Limbs, buds of, in Vertebrates, ll.

Limitation of potentialities, see Potentialities.

Limnaea, 67.

Linear cell aggregates, 5.

Liriope, 181.

Lithium chloride, eggs of Amphibia, 120, 135; salts, on Amphibian eggs, 120, 138, 135; on Echinoid larvae, 137-140, 146.

Liltorina, 71, 72, 74.

Lobe, polar, 214, 228; removal of, 217, 224, 227.

Localization of ontogenetic effects, 289.

Loss of potentialities, soc Potentialities.

Lucifer, 22, 47.

Macromercs of Echinoids, 22, 30, 185, 192; of Ctenophora, 211; of Mollusca, 215, 221.

Magnesium in artiﬁcial parthenogenesis, 123; Frog embryos, 133, 135; Fundulus, 135, 276; Arenicola larvae, 135; effect on ciliary and muscularmovement, 135; necessity of, 147.

Magnetism, 91 sqq.

Male element, function of, according to Aristotle, 13.

Mammalia, blastocyst, 2, 6. 7, 115; truncus arteriosus, splitting of, 6 ; trophoblast of placenta, 7 ; embryonic plate, 9 ; segmentation in, 24.

Man, see Human being.

Massive cell aggregates, 11.

Material basis of inheritance, 1, 19, 252; cause, see Cause ; explanations of ontogeny, 12, 20, 286-289, 290, 296, 298.

Materialism, 301.

Matter and material cause, 13.

Matter and mind, 801.

Maturation, nuclei of germ-cells in, 252, 254.

INDEX OF SUBJECTS

Maximum temperature, see Temperature.

Mechanical, see Mechanism; Mechanical agitation, 89. Mechanism of inheritance, 1, 280; mechanism, explanation of ontogeny as a, 12, 20, 286-289, 290, 292, 296, 298.

Medulla, roof of, 7; solid, in Frog's embryos grown in certain solutions, 133.

Medullary folds, 9; not formed at low temperatures, 104 ; divided at high temperatures, 99; remain open in various solutions, 119 sqq.; in Hemiembryo, 160.

Medullary plate of Amphioxus, 8.

Mcidusa from isolated blastomere,

81.

Megalecithal eggs, 24.

Membrane of egg, 45, 46; absence, due to poisons, 128; formed by chloroform, 129 ; at surface of cells, see Film ; of shell , 104 ; of Descemet, 276.

Mendelian allelomorphs, 266.

Mercury, 132.

Meridian, streaming, in inverted Frog's eggs, 84.

Meridional division, 22,32; see also Cell-division.

Megridional polarization in Frog's egg,

3.

Meroblastic segmentation, mentation.

Merogony, 131 ; male nucleus only in, 254

Mescnchyme, of Echinoderms, 5; of Echinoids, 185, 26], 262, 269; of partial larvae, 193, 194, 198. 237; of Ascidia, 235 ; of Echinoids, displaced, 274.

Mesoderm, segmentation of, 11 ; in Hemiembryo, 160; deﬁcient, after imury to germ-ring, 179; dependent on presence of polar lobe, 217.

Mesomeres in Echinoids, 27, 185, 192.

Mesonephric tubules, see Kidney.

Migtgibolism, nucleus needful for, 252,

4.

Metamorphosis (Harvey), 14.

Metanephric blastema, see Kidney.

Methods of embryology, descriptive, 12 ; experimental or physiological, 13.

Micromeres in Echinoids, 22, 30, 185, 192; suppressed by pressure, 37, 186; by heat, 186 ; in ‘spiral’ eggs, 25; suppressed by pressure, 39, 213,- rotation of, 29, 47 ; of Mollusca

see SegINDEX OF

isolated, 215, 221 ; of Ctenoplxora, 39, 209-211.

Micropyle, 183.

Migration of cells, 4, 5; of spicule cells in Echinoids, 114.

Mind and matter, 301.

Minimum temperature, see Temperature.

Mitosis,quadripolnr,tripolar,268-265.

Mitotic index, 65.

Mitrocoma, isolated blastomeres, 181 ; fusion of blastulae, 202.

Mbina, 24.

Molecules, meridional rows of, in Frog’s egg, 83.

Mollusca, muscles of, 5; segmentations of, 25; eggs of, influence of gravity on, 78; isolated blastemeres, egg-fragments, 215 sqq.; symmetry of egg and embryo, 244.

Monaster, 267.

Monsters, double, in lithium salts, 138; of Frog, 171, 172; of Newt, 174, 175; of Ampliioxus, 181; of Asterias and Echinus, 194; of Sphaerechiuus, 201.

Monstrosity, 1 ; natural, 18, 159; signiﬁcance of, 155, 156; produced by shocks, 89; perpetual rotation, 90; by sound vibrations, 90; by electric currents and magnetism, 91 ; by heat and cold, 98 sqq. ; by chemical agents, 120-121, 132; signiﬁcance for variation, 141.

Moral consciousness, 289, 299, 300.

Morphaesthetic, 291.

Morphine, effect on fertilization, 128130 ; on Hen's egg, 132.

Morphography, 12.

Morphology, method of, 12; experimental, 1.

Mosaik~Theorie, sea Theory, mosaic; segmentation, see Segmentation. Motion imparted by male element,

295, 296.

Mouth, growth of, 70.

Movements, of cells and oell-aggrcgates, 4 sqq., 272 ; in segmentation, seecell;oforgan-formingsubstances, 207, 208, 226, 227; protoplasmic, 40, 41.

Mucin, 58, 79.

Muscles, in Arthropods, 5 ; of Ascidia, 235, 237; in Mollusca, 5; of gut, 5; of blood-vessels, 273; striated, 55; abnormal, in rotated I-Ien’s eggs, 90; contractions of, sec Contraction.

Mutations, 141.

Myotomes, 11.

SUBJECTS 333

Myriapods, 24. Mysis, 24. Myzostoma, 214, 228.

Nature, 801.

Neck, rate of growth, 70.

Necturus, 168.

Nematocysts in Planula, 8.

Nemertines, segmentation in, 25 ; isolated blastomeres and egg-frag ments, 204-208.

Nee-vitalism, 20; see also Vitalism.

Nephelis, 40.

Nephridia of Earthworm, 3, 5: of Annelids, fusion with coelom, 6. Nereis, segmentation in, 25, 47; unequal asters, 80; egg compressed, 39, 213 ; crossed polar furrows, 46; change of shape of cells, 48; arti ﬁcial parthenogenesis, 124.

Nerves, growth 01', 5; branching of, 6; anastomoses of roots, 6; plexuses of, 6; fusion with end-organ, 6; crest, ll; ganglia, fusion of, 11; ﬁbres, 273; physiology of, 290.

Nervous system of Earthworm, 3; oi".l‘eleostei, 6, 11, 133; of Invertebrates, 11 ; of Ascidia, 235, 237.

Neural, sec Nerve.

Newt, development of half-blastomeres, 173; double monsters, 174, 175. .

Nickel nitrate, 94.

Nicotine, and eggs of Bufo, 120; effect on fertilization, 126,130; on Hen’s egg, 132.

Nitrates, 123.

Nitric oxide, eggs of Ascaris in, 112.

Nitrogen, eggs in, 110, 112.

Nomelectrolytes, poisonous effects of, 136 ; see also Cane-sugar, Dextrose, Urea.

Normal temperature, seo'1‘emperature.

Notochord, vacuoles of, 2 ; early differentiation of, 3; application of skeletal cells to, 5, 11 ; formation of in Urodela, &c., 8 ; from archanteron, 11; divided at high temperatures, 99; in cane-sugar, 133 ; whole, in Hemi-embryo, 160; of Ascidia, 285, 287.

Notoehordal tissue, in neural tube and archenteron, 134.

Nucleus, division of, a factor in development, 2, 22 sqq. ; division of, direction determines that of cell, 31, 55; rules of: Hertwig for, 31, 82; rule of Pﬂiiger for, 82; polar divisions of Insects, 29; polar divisions with equal asters, 81 ; division 334 without cell-division, 55, 56, 118 ; indirect division, see Karyokinesis.

Nucleus, position in cell, 81, 32, 79; injured by poisons, 128-130, 135; of yolk, sea Yolk-nuclei; qualitative division of, 19, 159, 168, 186, 213, 240, 265, 279, 280, 286 ; rapidity of, 65; idioplasm in the nucleus, 19, 159; the vehicle of inheritable characters, 253, 258262; in diﬁerentiation, 251 sqq.; essential for life of cell, 251, 252, 254 ; of germ-cells alike, 252, 254, 256; both male and female, not necessary, 254; decrease in size of, 65; variability in size of, 269; size of, and number of chromosomes, 268; number of, and number of chromosomes, 268; and cytoplasm, ratio between, 65, 269; volume of, and cell-surface, 269 ; responsive to stimuli, 283; reaction of, on cytoplasm, 266, 288, 284.

Number of cells, see Cell; of nuclei, see Nucleus.

Oakland, Mass., growth of, 68.

Obelia, 147, 148.

Obliquity of spindles in ‘spiral ' eggs, 32 ; of egg axis in Rana esculenta.

Occasion, 283.

Oceania, 54.

Oil-drops, 43 ; systems of, 49 sqq.

0ils,essential, 132.

Oniscus, 24.

Ontogeny and phylogeny, 12; causal explanation of, 13, 297, 298.

Operculum of tadpole, not formed after removal of brain, 176.

Opﬁnglia, artiﬁcial parthenogenesis,

4.

Ophiuroids, segmentation in, 24, 26, 28, 47.

Ophryotrocha, 202.

Optic cup, and lens, 134, 275, 276; and cornea, 276, 277.

optic vesicles, absent, 110; injured or removed, 275, 276.

Optimum temperature, see Temperature.

Orchcatia, 24, 26, 28, 47.

Organ-forming substances, see Substances.

Organs, elementary, of ﬁrst, second, &.c., orders, 3.

Orientation, similar, of egg-particles, 281, 286.

Origin and fate of cells, 246; knowledge of, 288, 298.

school-children,

INDEX OF SUBJECTS

Osmotic pressure, as aﬂecting growth, 2, 115, 116; as aifeeting celldivision, 56, 117 ; increased by addition of salt, 116, 117 ; the asserted cause of malformations, 120-122; the cause of artiﬁcial parthenogenesis, 128, 124 ; decrease of, 118; equilibrium, 118.

Otolith, 239.

Oviduct, in Amphibia and Amniota,9. Oxidation, agents of, 181 ; promoted by light, 112; by alkalinity, 151. Oxygen, lack of, cell-division suppressed, 56, 111, 112 ; effect on pigment of Fundulus, 111, 114; quantitative estimation of amount absorbed, 112; effect of pure, 112 114 ; need of, 109 sqq., 182.

Oxygenotactic, Oxygenotropic, 114, 273, 274.

Oxyhaemoglobin, excess of, in Chick, in pure oxygen, 112.

Papilla, ﬁxing, 229.

Paraderm, of Amniota, 8.

Paramn oil, 49.

Parallelism, psycho-physical, 301.

Paralysis of cytoplasm, 127.

Pars prima genitalia (Harvey), 14.

Parthenogenesis, artiﬁcial, produced by mechanical agitation, 90, 124; by cold, 108, 124 ; by increased osmotic pressure, 123, 124; cytology of, 124, 125; maternal nucleus only in, 254; number of chromosomes in, 125.

Patch, gray, in inverted eggs, 85.

Patella, isolated blastomeres of, 218222.

Pelagic ova, 89.

Pellogaster, segmentation, 24.

Penetration by cells, 5.

Peptones, 132.

Percentage increments of growth, 60 sqq. ; of water, 58-60, 115, 116.

Periods, spiral, radial, bilateral, of segmentation, sec Segmentation; variation in, 74.

Peripatus capcnsis, 24.

Perivitelline cavity, 79, 87 ; fluid, 79.

Periwinkle, 71, 72, 74.

Permeability of tissues to salts, &c., 119, 121, 124, 145.

Pegmutation of original elements,

86.

Petromyzon, notoehord in, 8, 183; segmentation, 47 ; artiﬁcial parthenogenesis, 124.

Phagocytosis, 5. INDEX OF SUBJECTS

Phallusia, 288.

Pharmacology, 136.

Phuscolosoma, 25.

Phosphorus, effect on Chick, 132; necessity of, 141.

Physa, crossed polar furrows, 46; effect of fertilization, 250.

Physico-chemicalresponsestostimuli, 285. ‘

Physiological zero, see Temperature, minimum.

Physiologically balanced solutions,‘

136.

Physiology of development, physiological laws, 18, 298. Pigment, change of position in isolated blastomeres of Frog, 53; of Frog's egg. 79; disappearance of, in Frog's egg, 80, 163 ; in Fundulus, in darkness, 94; without oxygen, 11], 114; distributed through cell-body (gray degeneration), 135; of secondary mesonchyme in Pluteus, 144; without magnesium, 148; without calcium, 150; in neutral media, 151; of Strongylocentrotus, 183-185; function in gastrulation, 184, 192; of Dontalium, 222; of Cynthia, 230 sqq.,

250.

Pilidium, 205-207.

Pilocarpine, 131.

Pineal vesicles, inner wall of, split in Lacertilia, 9.

Pisces, egg-axis and embryonic axes, 178; half-blastomeres, 178; injuries to lip of blastopore, 178.

Placenta, of Mammals, 7.

Placental Mammals, segmentation, 24.

Plunorbis, segmentation in, 25; crossed polar furrows, 46; light of various colours, 94.

Plant, division of cells, 41 ; absorption of water by, 58.

Planula, nematocysts in, 3; from blastula by unequal growth, 6; from isolated blastomeres, 181; giant, from fused blastulac, 202.

Plate, medullnry, see Medullary; plate, white, in inverted eggs, 85.

Platyhelmia, excretory tubules of, 6; segmentation, 54.

Pluteus, in light of various colours, 96; ciliary movement of, 135; effect of potassium, sodium, lithium, 137; necessity of various elements for, 142 sqq.; partial, 194-196, 199, 200; hybrid, 258 13;

335

262; arms of, dependent on skeleton, 188, 144, 150, 195, 275; growth rate of arms, 70, 71.

Podarke, crossed polar furrows, 46.

Pole cells, see Teloblast.

Polar areas, in Frog's eggs exposed to electric currents, and in other structures. 92, 98; bodies, asters of same size in, 31 ; in Ascaris m’grovenosa, 33; in Callidina, 34; division, violate Hertwig’s rule, 84 ; furrows, see Furrows; lobe, see globe; nuclei, see Nucleus; rings,

51.

Polarity of Echinoid eg, 183; see also Egg-axis.

Polarization, meridional, in F1-og‘s egg, S3; of egg-particles, 281, 286.

Polyaster, 128.

Polyclads, 25.

Polychaeta, inﬂuence of gravity on eggs, 78; pressure experiments, 89, 213.

Polyspermy, pathological, 107; due to poisons, 128; irregular distribution of chromosomes in, 253, 264, 265.

Polyzoa, 22.

Position, changes of position in development, 3; Davenport's cata logue of, 4 sqq. ; fate a function of, 188, 282.

Post-generation of missing half-embryo in Frog, 161, 162; denied by Hertwig and Morgan, 170, 171; conﬁrmed by Endres, 171.

Post-trochal region, 224.

Potassium bichromate, 94; bromide, 123, 132; nitrate, 123; sulphate, 123; iodide, 132; chloride, 123, 133; cyanide, prolongs life, 131, 132 ; larvae of Echinoids, 137, 146 ; necessity of, 146.

Potentialities, altered by chemical reagents, 140; limitation of, 175, 176, 180, 181, 191, 194, 201, 204, 242-244, 246, 277, 281, 284, 286; due to lack of material, 182, 194, 201, 225, 242, 281, 291; due to lack of speciﬁc material, 180, 201, 217, 225, 242, 243, 281, 291.

Practical concept of freedom, 289.

Predetermination of embryonic axes, see Embryonic axes; physicochemical, 285.

Prediction, the function of science, 299.

Preformation of organ-forming stances, 207, 227, 241, 286.

sub336

Preformation hypothesis, sec Evolution.

Prelocalization, 207, 222, 227, 241.

Pressure, atmospheric, see Atmospheric ; osmotic, see Osmotic ; experiments on eggs of Frog, 84-37, 40-168; on eggs of Echinoderms, 37, 38, 185, 186; on eggs of Nereis, 39, 213; on eggs of Beriie, 39; as a meansof preventing cell-division, 55, 56.

Primary qualities, 300.

Primitive streak absent, 104.

P1-i1;ciple of germinal localization, 1 .

Principle of least surfaces, 41-44; in spiral segmentation, 45-47; disobeyed in radial and bilateral types, 49.

Proctodaeum, 9.

Pronuclei, causes of union of, 130; ggﬁh male and female, not needed,

Proportionality, 293, 295; of parts in partial larvae, 198-201, 290 ; in one larva from two blastulae, 202.

Prototroch, 218, 224.

Protozoa, alteration of shape during division, 48; senile decay, 65 ; enucleate fragments of, 254; in electric current, 93.

Pseudo-gastrula, 287.

Pseudo-triaster, —tetraster, 108, 129.

Psychoid, 291, 297.

Puberty, increase in growth-rate at, 65 ; increase in variability at, 73 ; increase in value of correlation coeﬁicient at, 75, 76.

Purposiveness, 286-289, 296, 299.

Purposes, see Purposiveness.

Pyrasoma, 24.

Quadripolar mitosis, 263-265, 267.

Qualities, inheritable, see Inlieritable.

Qugaggties, primary and secondary,

Qualitative division of nucleus, see Nucleus.

Quartettes of xnicromeres in ‘spiral’ eggs, 25, 80, 213; isolated, 215, 221.

Quliéigne, effect on fertilization, 128 Rabbit, decline of growth-rate, postnatal, 62, 65; prenatal, 68.

Radial segmentation, see Segmentation; radial symmetry, sea Symmetry.

Rana esculenta, bilateral segmentation, 26, 28; obliquity of egg-axis,

INDEX OF SUBJECTS

81, 168; cardinal temperature points, 100; eifects of salt, 118; brain and eyes of tadpole remo\ ed, 175; fusca, segmentation, 28, 47, 53; cardinal temperature points, 97; inﬂuence of heat and cold, 97-100; effect of salts and other substances, 119—123; artiﬁcial parthenogenesis, 124; symmetry of egg, 8], 163, 247, 248; formation of lens, 275; palustris, symmetry of eg, 81, 247; inﬂuence of cold, 100; temporaria, symmetry of egg, 81, 163, 247, 248; virescans, influence of cold, 100.

Rate of cell-division, see Cell—division ; of Development, see Development ; of growth, see Growth.

Ratio between nucleus and cytoplasm, 65, 268, 269.

Rearrangement of cells, 11.

Recapitulation theory, 12, 16.

Red light, 94-96.

Redistribution of yolk and protoplasm in inverted eggs, 84; of organ-forming substances, 207, 208, 226, 227.

Reducing agents, 131.

Reflective judgement, 287, 288.

Reﬂex movements of tadpoles deprived of brains, 176.

Regeneration, 290; a form of development, 1 ; of head of Tubularia, 114, 144 ; and Reserve Idioplasson, 159; of missing half-larva in Ctencphora, 210; in Ascidia, 239; necessity of nucleus for, 254; of lens, 276; nucleus in, 284, 285.

Regulation, 290.

Rsnilla, 55.

Reproduction, sexual.1 ; Aristotelian view of, 13, 294-296.

Reserve-idioplasm, 159, 161, 266.

Resistance to injury, capacity for, alters, 104.

Respiration of Chick, 109, 113; of Frog's egg, 110, 111, 112 ; of Telecstean ova, 111; of Echinoid ova, 112.

Responses to stimuli, 12, 20, 114, 272-277, 288-285.

Restriction, genetic, 246, 277 ; see also Potentialities, limitation of.

Retardation of development, by sound vibrations, 90; by light of certain colours, 94-96; at low temperatures, 97, 99, 100;- without oxygen, 110, 111 ; by desiccation, 115; by increased osmotic pressure, 117; in absence of potassium, 146 ; and INDEX OF SUBJECTS

magnesium, 148; and calcium, 150 ; with decrease of germinal VJI16, 200, 204.

Rhumkorﬂ coil, 91.

Rhynchelmis, 251.

Rhythm of development, 3.

Rind at surface of blastomeres, 54.

Ringer's solution, 120.

Rotatiomperpetual, of Hen’s eggs, 90 ; of pronuclei in Ascaris nigrovenasa, 34: in Diplogasler, 34; not in Oallidina, 34; of tirst two blastemeres in Oallidina, 34; of micromeres on macromeres, 47.

Rotation structure, ‘of ova, 79.

Rotifera, 26.

Rubidium, 146.

Rule, of Balfour, 29, 88; exceptions to, 80; of Hertwig, 31, 32, 85; conﬁrmed in Ascaris nigrovenosa, 34; and in Diplogaster, 34; by pressure experiments, 34 sqq. ; violated, by polar divisions, &c., 34 ; of Pﬂiiger, 82, 85.

Sagittal plane and first furrow in Frog, 17, 159, 165-168; in Cteno— phora, 209; in Newt, 175; in Ascidia, 229, 235; and plane of symmetry, 80, 165, 229, 235; determined by gravity, 82, 84; and streaming meridian, 84.

Salmo lens, 276; see also Trout.

Salps, blastomeres of, 5, 28, 54; rearrangement of cells, 11.

Salt-solution as a medium, 45, 53; see also Sodium chloride.

Salts of various kinds, malformations in, 119 sqq., 182, 276; artiﬁcial parthenogenesis, 123, 124.

Salvelin-Is, 178.

Sauropsida, blastoderm of, 6.

Science, the scope and functions of, 297 sqq.

Sciences, relation of, to one another, 299, 300.

Scleroblasts of Sponges, 3.

Sclerotome, 11.

Sea-urchin, sec Eohinoids, Pluteus, Echinus, Sphaerechinus, Strongmacentrotus, Arbacia.

Sea-water, calcium-free, used to separate blastomeres, 45, 190.

Sea-water, composition of, 141; artiﬁcial, 142.

Secondary qualities, 800.

Secretion of substances in growth, 258.

Segmental duct, splitting of, 6.

JIIKIIIOH

growth of, 5 ;

N

337

Segmentation of the ovum, a factor in development, 2, 279; the beginning of development, 22; and differentiation, see Cell-division, Differentiation; a mosaic, 17, 19, 158, 213, 236, 245, 246; types of, 22

sqq. ; radial, 22, 49, 53; spiral, 25,‘

30, 45-47, 52, 204; bilateral, 26, 49, 52, 229, 285 ; isobilateral, 26, 208; irregular, 28, 54, 55; periods of spiral, radial, bilateral, 26; types pass into one another, 28; meroblastic, in Vertebrata, 24; in Arthropods, 24; in Cephalopods, 27; in centrifugalized Frog's egg, 29, 88; as a result of heat, 97; in salts, solutions of, 119; causes of pattern of, 29 sqq., 197, 246, 266; and egg—axis, 81; in compressed eggs, 34-40; inﬂuence of gravity on, 78 sqq. ; effect of electricity on, 92; of heat, 97, 105; of cold, 100 ; of altered osmotic pressure. 117, 118 ; of chemical poisons, 119, 121, 128-130; of lack of oxygen, 111, 112; without sulphate, 143; without chlorine, 145; without sodium, 145; without potassium, 145; without magnesium, 145; in neutral media, 151 ; without calcium, 150 ; irregularities of, produced by heat, 105, 187; by shaking, 188; altered osmotic pressure, 117, 118; by dilution of sea-water, 187 ; alkaloids, 128-130; by calciumfree sea-water, 45, 188, 190; by double fertilization, 264, 265; of isolated blastomeres, total. 173, 178, 180, 181,230; partial, 190, 204, 205, 210, 211, 215 sqq., 229, 237 ; of eggfragments, I95-197, 207, 212,226,227.

Segmentation cavity, 11; absent in Stereoblastulae, 128; reduced without potassium, 146; in Hemiblastula, 159.

Segmentation of mesoderm, 11; of neural crest, 11.

Selective death—rate, 74.

Selenium, 144.

Self-differentiation, sec Differentiation.

Senescence, 65.

Senile decay, 65.

Sense-organ of Ctenophora, 211 ; see also Apical organ.

Separation of blastomeres, see Blastemeres.

Septa, of corpus luteum, 5.

Serosa, of Sipzmculus, 9.

Serpula, 272. 338 INDEX OF

Serranus, 178.

Sertulareln, 272.

Sertula-ria, 272.

Sexual reproduction, 1 ; Aristotelian view of, 13, 294-296.

Shaking eggs, a means of disarranging blastomeres, 188 ; or separating them, 190; prevents division of centmsome, 265, 267.

Shape of cells, 29; altered by spindle, 48.

Shocks given to eggs, 89.

Silicon, 141.

Silk-worms, exposed to inﬂuence of magnetism, 91.

Sipunculoids, 22.

Sipunculus, 9.

Siphonophora, 181.

Size, increase of, see Growth; of organisms, 65; of cells, determined by yolk, 29, 48; in partial larvae, 199; of bubbles, 43, 46; of nucleus, sea Nucleus; of partial larvae and their organs: in the Newt, 173; in Fishes, 178; in Amphioxus, 181; in Echinoids, 198, 199; in Nemertines, 206.

Skeletal cells, and notochord, 5.

Skeleton, of Chick, malformations of, 90; of Pluteus, in potassium, 137; and lithium salts, 138; Without sulphate, 144; without potassium, 146; without magnesium, 147 ; without calcium, 149; without carbonate, 150; in bromides, 145; in neutral media, 151; number of arms depends on number of spicules, 138, 144, 150, 195, 275; dependent of nucleus, 261; of hybrids, 259-262.

Soap ﬁlms, 41, 42; bubbles, 43-47.

Sodium, chloride, used to suppress cell-division, 56, 117; to produce abnormalities, 119, 120, 133-135; larvae of Echinoids, 137; bromide, 120, 134; sulphate, 135; nitrate, 135; butyrate, 140 ; formate, 145; hydrate, 151, 262; acclimatization to, 136; effect on ciliary and muscular movement, 185; necessity of, 145.

Solenoid, Tesla's, 91.

Solutions, chemically equivalent, 140; equimolecular, 140; hypertonic, 116, 117, 120-122, 128, 124; hypotonic, 118; isotonic, 120, 123, 124, 140, 142.

Somatic cells of Aacaris, 252, 258.

Somatopleure, fused with allantois, and trophoblast, 9.

SUBJECTS

Somites, fusion of, 11.

Soul, Aristotelian doctrine of, 292297; nutritive, 18, 293; perceptive, 13, 293; reasoning, 294 ; and body, 292.

Sound-vibrations, effect of, 90.

Speciﬁc gravity of yolk, 78, 79.

Spermatozoon, entrance of, and direction of first furrow, 40, 185, 247, 248; entrance of, and gray crescent, 80, 165, 247, 248; entrance of, and alteration of eggstructure, 80, 165, 222, 232, 247251 ; sperm-sphere and aster, 123; nature of stimulus imparted by, 124; effect of alkaloids on, 129; without potassium, 146; without magnesium, 147.

Sphaerechinus, effect of heat on, 105, 106, 187; potassium and lithium larvae, 137-140; necessity of cortain elements for, 142 sqq.; ‘pressure experiments, 186; isolated blastomeres, 191 ; blastomeres disarranged, 187; pieces of blastulac and gastrulae, 194, 195; size of partial larvae and their organs, 198, 200; one embryo from two blastulae, 201, 202; hybridization experiments, 258-262.

Spina biﬁda, in centrifugalized eggs, 88; as a result of heat, 99; ofocold, 100; of lack of oxygen, 11 .

Spindle, nuclear, direction of elongation of, 31, 32, 34, 39, 48, 248; oblique in ‘spiral' eggs, 32; rigidity of, 48; in artiﬁcial parthenogenesis, 124, 195; in karyokinetic plane, 185.

Spiral segmentation, see Segmentation ; asters, see Asters.

Spirogyra, 269.

Spleen of Vertebrates, 5.

Splitting of cell-aggregates, 6, 9, 11.

Sponges, larval skeleton, 2, 3; gemmule of, 5, 11 ; segmentation of, 22, 47; larva cf, 6; unipolar immigration in, 11.

Spongilla, 47.

Sports, 141.

Stages, series of, in development, sec Rhythm.

Standard deviation, 71.

Static teleology, 291, 292.

Stature, human, change in rate of increase, 68-70; variability of, 74 ; correlations with other magnitudes, 75.

Stereoblastulae, 128. INDEX OF

sternum, girth round, rate of growth, 70.

Stisiuli, 12, 20, 272-277, 288-285; of spermatozoon in fertilization, 128, 252; exerted by spicules of Pluteus, 137. 275; directive, in post-generation, 162.

Stolen, see Hydroids.

Stomodaeum, 9; formed in Aneuteria, 106; in Exogastrulae, 137; absent without potassium, 146; formed from isolated ectoderm, ‘-95; of Ctenophora, 210, 211.

Stratiﬁcation of organ-forming substances, 242, 280.

Strongylocentrotus lividus, segmentation of, 23 ; egg stretched, 39; decline of growth-rate, 67; growth-rate of arms of Pluteus, 70; decline of variability in, 71 ; eggs exposed to heat, 107; to action of alkaloids, 126 sqq. ; normal development, 183-185; mieromeres suppressed, 186; hybridization experiments, 261, 262; purpuratus, heterogeneous cross-fertilization, 262.

Strontium, 156; bromide, 182, 133.

Structure, increase of, 2, 279 ; of egg, see Egg-axis, Symmetry of egg.

Strychnine, eggs of Bufo, 120; effect on fertilization, 128-130; on Chick, 132.

Substances, organ-forming in cytoplasm, I9, 207, 208, 217, 224, 225, 228, 233, 236, 240, 241, 246, 280, 286; speciﬁc action of substances present in sea-water, 152.

Succession, in the formation of parts, 13.

Suckers, 239.

Sulphates, 123; of quinine, 128-130 ; of magnesium, 135; of sodium, 135; necessity of, 143, 144.

Sulph-ion, see Sulphate.

Sulphuric acid, 145.

Superﬁcial cell-aggregates, see Cell.

Surface, area of, in partial larvae, 198, 269; area of nuclei and number of chromosomes, 268.

Surface-tension, 41 sqq., 55, 56.

Surfaces of contact between blastemeres, in radial segmentation, 22; in spiral segmentation, 45-47; between isolated cells, 54; between soap-bubbles, 43; between oildrops, 49 sqq.

Suspension of development, at low temperatures, 97, 99, 102, 104.

Swelling after shrinkage in hypertonic solutions, 124.

SUBJECTS 339

Sycandra, 22.

Symmetry, bilateral, of egg cytoplasm, 33, 80, 163, 232, 247, 280, 281; radial, of egg, 79, 163, 204, 222, 232, 280; bilateral, and sagittal plane, 80, 229, 235, 244 ; produced by gravity, 82, 84, 87; radial, of Plutei without sulphate, 144; hilateral and first furrow, 167, 229, 235.

System, equipotential, 176, 277 ; inequipotential, 176, 277.

Systems of lamellae, see Lamellae; of drops, see Drops.

Tadpole, tail of, phagocytosis in, 5; absorption of water, 58. 59; rate of growth in, 67; inﬂuence of light on, 94; effect of pure oxygen, 113, 114; effect of increased atmospheric pressure on, 114; brain and eyes removed, 175, 275 ; formation of lens and cornea, 275-277.

Tail of Tadpole, phagocytosis in, 5; doubled at high temperatures, 99.

Taxis, Tactic, see Stimuli and Responscs.

Teleology, of Kant, 286-289 ; dynamic, 291, 296; of Aristotle, 297.

Teleostei, nervous system of, 6, 138; segmentation, 26, 35; injuries to germ-ring, 178; effect of fertilization, 251.

Tellurium, 144.

Teloblast, continued division in same direction, 29; continued unequal division, 30 ; unequal asters in, 31.

Telolecithal eggs, 24, 30.

Temperature, as affecting growth, 2, 59; cardinal points, for Frog’s egg_ 97; for Hen’s egg, 102; for Plutei, 107 ; alteration of variability with, 107; high, see Heat; low, see Cold.

Tendon, 5.

Tension, see Surface-tension, Decomposition-tension.

Tentacles, branching of, 6.

Teratology, 119, 155, 156.

Teredo, 26.

Tetrahedral arrangement of cells, 24, 25, 26, 28, 29, 47; of soapbubbles, 43.

Tetrasters, 107; produced by alkaloids, 128.

Thalamencephalon, roof of, 7.

Thelykaryotic, 267..

Theory, ‘Mosaic’, or self-differentian tion, 17, 19, 158 ; analytical, of Driescli, - 280-286. 340

Thickenings of cell-aggregates, see Cells, movements of.

Thigh, rate of growth, 70.

Thigmotropism, 272, 275-277.

Thinnings of cell—aggregates, see Cells, movements of.

Thio-sulphate, 144.

Tlﬁmus, concentric corpuscles of,

Toad, 82, 102.

Totipotent, parts, 76; see also Whole nuclei, 266, 283, 284, 285.

Toxicity, 122, 135, 136.

Toxines, bacterial. 132.

Totcopneusfes, 185, 250.

Transportation, by cells, see Cells, movements of.

Trematoda, segmentation, 28.

Triasters, 107 ; produced by alkaloids, 128.

Triclads, blastomeres of, 5, 54, 273; segmentation, 28, 55.

Tripolar mitosis, 265.

Trochoblasts, 213, 220.

Trochophore, 224, 225.

Trochus, 26, 30, 45-47.

Trophoblast of Mammalia, 7; fusion of, with embryonic plate, 9; {used with somatopleure, 9; coenocytic and syncytial, 55.

Trophochromatin, 258.

Tropism, Tropic, see Stimuli Responses.

Trout eggs placed between poles of a magnet, 91 ; between electrodes, 91 ; in light of various colours, 95.

Truncus arteriosus of Mammals, splitting of, 6.

Tubercle toxine, 132.

Tubularia, 114, 144.

Tunicata, follicle cells of, 5.

Turbellaria, yolk-glands, 5.

'1‘urgor_of plant-cells, 246.

and

Umbrella, segmentation, 26, 30; ﬂattening of blastomeres, 45 ; parallel polar furrows, 46. ,

Understanding, 301.

Unequal division, see Cell-division ; of oil-drops, 62; numbers of chromosomes, see Chromosomes.

Unio, segmentation, 25, 80, 47; unequal asters, 30 ; ﬂattening of blastomeres, 45; crossed polar furrows, 46.

Unipolar, sec Immigration.

Units, representative of inheritable qualities, 19, 159, 246, 246, 279, 280.

INDEX OF SUBJECTS

Urea, Frog's eg, 121; notochordal tissue, 184; nuclei, 135. Urodela, notochord in, 8, 183.

Vacuolation of cells of spaerechinue, 146, 261.

Valency, 136.

Variability, decline of, during growth, 71-74 ; increased at time of puberty, 78; causes of decline, 74 ; increased by an adverse change of conditions, 74, 141, 156; in period, 74.

Variation, see Variability.

Vasa eiferentia, union of, with mesonephric tubules, 6.

Vegetative cells, see Cells.

Vegetative pole, blastopore at, 244.

Veliger, 217.

Ventricosity, of shell of Littorina, 71.

Vertebral column, rate of growth, 70.

Vertebrates, spleen of, 5; bone and tendon, 5; vitreous body, 5; vasa etferentia, 6; nerve-crest, 11; segmental duct, 5; lens, 7; auditory vesicle, 6; limb-buds, 11 ; medulla, 7 ; artiﬁcial parthenogenesis, 124; thalamencephalon, 7; segmentation, 22, 24, 47.

Viole de Parme, 94. Violet light, 94-96. Vital constant, 291. Vitalism, 291 sqq.

Vitellophags, 4, 273 ; vitelline membrane, see Membrane.

Vitreous body of Vertebrate eye, 6.

Volume of partial larvae, 199 ; of cells, and number of chromosomes, 26:; of nucleus, and cell-surface, 26 .

Waist, rate of growth, 70.

Water, absorption of, in growth, 2, 58, 59, 115, 116; in liypotonic solutions, 118; decrease of, in later stages, 58 ; evaporation of, from albumen, 115; not absorbed by egg of Frog, 123.

- withdrawal of, 119; in fertilization, 128 ; and artiﬁcial parthenogenesis, 128, 124; aggregated about apical body and middle-piece of spermatozoon, 123.

-— fresh, 117, 136; distilled, 136.

Weight, increase of, as a measure of growth, 58 sqq.; correlations with other magnitudes, 75; loss of, in Hen's egg, 115.

plate, in inverted Frog's eggs, INDEX OF SUBJECTS 341

Whole larvae from blastomeres or 29, 48; speciﬁc gravity of, 78; egg-fragments, in the Newt, 178; descent of, in inverted eggs, 84; 1.1 Teleostei, 178; in Amphioxus, in centrifugalized eggs, 88; in ISO: in Coelenterates, 181 ; in jured by heat, 97; by cold, 98; by Echinoderms, 198, 197 ; in Nemer- salts, 119, 185; elfect of removal

tines, 204, 207. in Teleostei, 178; of Cynthia, 280 ; Withdrawal of water, see Water. . -cells of Triclads ‘and Salps, 6 ; Woman, see Human being. -gland of Turbellaria, 5 ; -1ol_>e, see Worcester, Mass, school-children, Lobe, polar; -nuclei in centrifugagrowth of, 63. lized Fi-og’s egg, 30, 88; in eggs grown in salt, 119; -plug persistent

xxlol’ 54' 313 8 result ofheat, ; Of cold, ;

ln.B0l11tl0nS of salts, &c., 119 sqq. ;

-sac ruptured in eggs that are not Yellow light, 94-96. turned, 89; of Fundulus, pigment Yolk, 2; inﬂuence of, in cleavage, 24, of, 94, 111, 114, 274.

ADDENDA ET CORRIGENDA

P. 5, 5 lines from bottom, for unicellular read multicellular. P. 28, line 10, after irregular, insert and in Triclads.

P. 57. To Literature acid J. Sacns. Die Anordnung den-Zellen in jiingsten Pﬂanzentheilen, Arb. Bot. Inst. Wurzburg, ii, 1882. _

P. 114. To Literature add G. BUNGE. Weitere Untersuchungen iiber die Athmung der Wiirmer, Zeitsc-hr. physiol. Chem. xiv, 1890.

P. 140, line 22, for prospective potentialities read prospective signiﬁcanoes.

P. 225, 2 lines from bottom, for is now placed in road has now moved into.

P. 271. To Literature add W. S. Surrox. On the morphology of the chromosome group in Brachyslola magna, Biol. Bull. iv, 1902.

P. 278. To Literature add J. W. Jnxxmsox. On the effect of certain solutions upon the development of the Frog's egg, Arch. Ent. Mech. xxi, 1906.

Book - Experimental Embryology (1909)

Contents

Experimental Embryology

Preface

Contents

Chapter I