Text-Book of Embryology 2-10 (1919)

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A personal message from Dr Mark Hill (May 2020)  
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I have decided to take early retirement in September 2020. During the many years online I have received wonderful feedback from many readers, researchers and students interested in human embryology. I especially thank my research collaborators and contributors to the site. The good news is Embryology will remain online and I will continue my association with UNSW Australia. I look forward to updating and including the many exciting new discoveries in Embryology!

Kerr JG. Text-Book of Embryology II (1919) MacMillan and Co., London.

Textbook Chapters: 1 Formation of the Germ Layers | 2 Skin and Derivatives | 3 Alimentary Canal | 4 Coelomic Organs | 5 Skeleton | 6 Vascular | 7 Internal Body Features | 8 Adaptation to Environmental Conditions | 9 General Considerations | 10 Common Fowl | 11 Lower Vertebrates | Appendix

- Currently only early Draft Version of Text -

Historic Disclaimer - information about historic embryology pages 
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Pages where the terms "Historic" (textbooks, papers, people, recommendations) appear on this site, and sections within pages where this disclaimer appears, indicate that the content and scientific understanding are specific to the time of publication. This means that while some scientific descriptions are still accurate, the terminology and interpretation of the developmental mechanisms reflect the understanding at the time of original publication and those of the preceding periods, these terms, interpretations and recommendations may not reflect our current scientific understanding.     (More? Embryology History | Historic Embryology Papers)

Chapter X The Practical Study of the Embryology Of The Common Fowl

FOR gaining practical experience in the study of embryology there is no type of material so convenient as that of the early stages in the development of the Common Fowl. Freshly laid eggs can be obtained practically anywhere and to obtain the various stages of development all that is necessary 1 is to keep the eggs at a suitable temperature (about 38° C.) either under a sitting hen, or in one of the incubators which can be purchased, or even in a simple water-jacketed even such as can be made by any tinsmith. If an incubator be purchased it will be provided with a proper heat regulator for use with electricity, gas or oil, while with the most primitive water-bath it is possible to arrange a lamp so as to give a temperature sufliciently constant as to carry the eggs through at least the first few days of incubation—the most important period for purposes of study. Bird embryos—apart from their use in learning practical embryology-— provide admirable material for giving practice in the ordinary methods of section-cutting which are in such constant use in Zoology, Anatomy, Physiology, and Pathology. This chapter will then be devoted to giving an account of the development of the Fowl with directions as to the technique involved in its practical study.


In the description which follows the developmental phenomena will be described in their natural sequence but on account of the practical difficulties involved in the extraction and preservation of blastoderms of the first day of incubation it will be found best, in actual laboratory work, after studying the new-laid egg and its envelopes, to proceed to the stage of about 42 hours’ incubation and gain some practice in the manipulation of it before attempting the earlier stages. In the following technical instructions the sequence is followed which has been found to be in practice most convenient for beginners.

Technical Directions

I. New-Laid Egg

Fil1 a glass vessel about 4% inches in diameter and 2 inches in depth With normal salt solution [Water

1 Provided the eggs have been fertilized. 9 The reader is assumed to have an elementary knowledge of the ordinary methods of cutting sections. See, however, the Appendix.

100 c.c., common salt '7 5 gramme] heated to a temperature of about 40° C. Submerge the egg upon its side in the salt solution and remove the side of the shell which is uppermost by cutting with a pair of strong scissors and then lifting off the isolated piece of shell with blunt forceps. Take care to keep the point of the scissors or forceps close to the inner surface of the °shell so as to avoid risk of injury to the true egg or “ yolk.”

II. Egg After 42 Hours’ Incubation

Open the egg as before. On removing the piece of shell the blastodcrm will be seen as a circular whitish area on the upper side of the yolk. Excise the blastodcrm by making a series of rapid cuts with the large scissors through the vitelline membrane a short distance external to the boundary of the blastodcrm. Should the yolk happen to be tilted round so that the blastodcrm is not uppermost but rather at one side make the first cut below the blastodcrm so that the elasticity of the vitelline membrane will tend to pull it upwards when the cut is made. Otherwise the blastodcrm may be lost by its being pulled downwards.


Having isolated the circle of vitelline membrane, with its adherent blastoderm, slide it off the yolk by pulling gently on one side with the forceps. Remove the remains of the egg from the dish so as to keep the salt solution clean. Take hold of the circle of vitelline membrane at one edge with the forceps and wave it backwards and forwards beneath the surface of the salt solution. The blastodcrm will gradually become detached. Should it not do so at once the separation should be started by freeing it from the vitelline membrane with a scalpel at one edge. Notice the difference in appearance between the vitelline membrane and the blastodcrm which has been detached from it. If the blastodcrm is yellow from adherent yolk this should be washed oil’ either by waving the blastodcrm backwards and forwards in the salt solution or by gently directing jets of salt solution 011 the yolky surface of the submerged blastodcrm by a wide-mouthed pipette.


The blastodcrm should. now be brought near the surface of the salt solution and a watch-glass slipped under it by which it may be lifted from the larger vessel. The blastodcrm is so delicate that it must be kept submerged in the fluid: no attempt must be made to lift it above the surface by forceps.


A microscope coverslip slightly larger than the blastodcrm should now be submerged in the watch-glass and the blastodcrm floated over it dorsal side above. The dorsal or upper side of the blastodcrm can easily be identified from the fact that the edges of the blastodcrm tend to curl upwards. Having floated the blastodcrm over the coverslip the latter should be gently raised to the surface of the fluid with a pair of large forceps. Take care to keep the coverslip absolutely horizontal and lift it out of the fluid very carefully so that the blastodcrm is stranded on its upper surface, the lower surface of the blastodcrm being in contact with the coverslip. The superfluous salt solution should be drawn away with blotting-paper so as to bring the blastoderm into close contact with the glass; take great care that the blotting-paper does not actually touch the blastoderm as in that event it will be apt to stick to it. Now take the coverslip between the finger and thumb and with the aid of a pipette place a very small drop of corrosive sublimate solution (or other fixing fluid) upon the centre of the blastoderm. This will cause the blastoderm to adhere to the coverslip. Now invert the eoverslip and drop it on to the surface of some fixing fluid in a watch-glass.


The blastoderm is then passed through the various operations of staining, dehydrating and clearing, preparatory to mounting whole oi‘ conversion into a series of sections as the case may be. The advantage of having the blastoderm adherent to a coverslip is that it makes it easier to handle and above all it keeps it from becoming wrinkled or folded. The blastoderm if fixed in corrosive sublimate can usually easily be detached from the coverslip at the stage of clearing if it has not already become free at some preceding stage. Should it adhere obstinately it should be placed i11 acidulated alcohol for an hobr or more.

The examination of the blastoderm should be carried out as follows:

1. Study the blastoderm and embryo as a whole under a, preferably binocular, dissecting microscope while it is submerged in the fixing fluid. As the fixing fluid penetrates the embryo the various details in its structure come into view. Continue the examination of the surface relief in the alcohol which is used for getting rid of the excess of the fixing agent. After examining from the dorsal side invert the blastoderm and examine from below.

2. Repeat the examination of the embryo as a whole as a transparent object after staining and clearing. If the individual embryo is to be cut into sections a careful drawing should be made at this stage, the outline being preferably drawn by means of the camera lucida.

3. Study serial sections cut transversely to the axis of the embryonic body.

Sagittal and horizontal sections will also be useful for study after the transverse ones.

III. Early Second-Day Blatoderm

The same method is used as for the 42-hour stage but special care must be taken on account of the more fragile character of the blastoderm. In all probability the blastoderm will remain adherent to the vitelline membrane in spite of repeated shaking and the process of detachment will have to be started by gently easing up the edge of the blastoderm on the side next the forceps in which the edge of the circle of vitelline membrane is held.

To get rid of adherent yolk the circle of vitelline membrane should be laid on the bottom of the dish of salt solution, blastoderm uppermost. A pipette with a wide mouth should be held vertically X TECHN ICAL DIRECTIONS 51]

a few millimetres above the blastoderm and the india-rubber bulb squeezed rhythmically so as to wash away the particles of yolk by very gentle currents of salt solution. When the blastoderm is lifted out of the solution stranded upon the coverslip it is very apt to become folded. When this happens, on account of the fragility of the blastoderm, no attempt should be made to stretch it out by the use of needles or forceps. The folds should rather be straightened out by a current of salt solution allowed to flow out from the orifice of a pipette held vertically just over the centre of the blastoderm.

IV. Early Blastoderms

Open the egg as before. Let the albumen run off until the vitelline membrane over the blastoderm is exposed. Raise the egg until the blastoderm touches the surface of the salt solution and then bring a wide-mouthed pipette of Flem1ning’s solution, held vertically, into such a position that its tip just touches the film of fluid over the blastoderm. Let the solution flow down slowly on to the vitelline membrane covering the blastoderm. If there is any albumen overlying the blastoderm this should be carefully stripped elf as it coagulatcs. A small piece of black bristle should be stuck into the vitelline membrane on each side to mark the line joining the chalazae so as to facilitate the orientation of the blastoderm for section-cutting. The fixing fluid should be allowed to act for several minutes and then a circle of vitelline membrane may be excised with the blastoderm adhering. to it. Floatv out the circle of vitelline membrane on a coverslip with the blastoderm above and submerge in a watch-glass of fixing fluid. If the circle of blastoderm adheres to the coverslip so much the better: it may be separated in the clearing agent.

Instead of a pipette as above indicated being used for the fixing fluid a small rim of cardboard, e.g. the rim of a small pill-box lid, may be placed on the surface of the yolk, raised up slightly out of the salt solution, so as to enclose the blastoderm and then the little tank so formed may be filled with Flemming’s solution which will gradually diffuse downwards. Minchin recommends a triangular instead of a circular rim for facilitating subsequent orientation.

For fine work it is preferable to embed the whole yolk in celloidin and then after the celloidin has been hardened to c11t out the portion in the region of the upper pole for sectioning. This method consumes however much more time than does the paraffin method.

V. Third-Day Egg

A. Open the egg as before.

B. Study the embryo and blastoderm while still alive and in situ. A large outline drawing should he made. The details of the body of the embryo will be seen better later but the arrangement of the blood-vessels can best be studied now while the circulation is still active. As a rule they can be seen distinctly through the vitelline membrane but if not the latter should be carefully stripped off. A Greenough binocular microscope with its lowest power objectives is a useful accessory for examining the blood-vessels.

C. Excise the embryo with the surrounding portion of blastoderm, float it on a slide, cover with coverslip supported by wax feet at the corners and examine as a transparent object, comparing the various features with those shown in Figs. 235 and 236.

D. Excise a second embryo with its surrounding blastoderm. Float it on to a coverslip, embryo above, and submerge it in a watchglass of fixing fluid. Watch it carefully under the lens or preferably low-power binocular as the tissues gradually become opaque. The amnion will be seen particularly clearly during this process. A drawing should be made of the embryo enclosed in its amnion as an opaque object.

E. Carefully strip off the amnion with a pair of needles 1 and study the configuration of the head end of the embryo.

F. Stain and mount the embryo.

G. Prepare series of sections (at) transverse to trunk region, (1)) horizontal through trunk region and therefore approximately sagittal in the region of the head which is lying over on its left side.

VI. THE FOURTH DAY.-On placing the egg in the salt solution the broad end will tilt up more decidedly than before owing to the increase in size of the air space. Care should therefore be taken to make the first perforation of the shell close to the broad end so as to allow the air to escape. Care must also be taken not to injure the vascular area as the whole blastoderm is now much closer to the shell than it was in earlier stages. As soon as the egg has been opened a careful drawing should be made while the embryo is still alive and in situ. The main features of the vascular system in particular should be carefully worked out at this stage. If the circulation becomes sluggish through cooling a little warm salt solution should be added but care must be taken not to bring about a great and sudden rise of temperature as in this case the greatly accelerated heart-beat is apt to cause rupture of a vessel.

The body of the embryo, allantois, ete., are covered over by the thin transparent serous membrane or false amnion as becomes apparent if the attempt is made to push a blunt needle down into the space round the allantois. This membrane should either be cut thrpugh with a pair of fine scissors, just outside the boundary of the allantois, or carefully stripped off with fine forceps. When this has been done it is possible to shift the body of the embryo into such a position that it with its blood-vessels can be observed in side view. Until this has been done it is impossible to get a proper view of the body of a well-developed embryo of this age owing to its dipping down out of sight into the yolk-sac.

The embryo should now be excised by cutting round outside the boundary of the vascular area and floated into a watch-glass of clean warm salt solution. The embryo may now be studied as a transparent object on the stage of the dissecting microscope. It is better however in the first attempt to proceed at once to fix the embryo. An essential preliminary is to remove the true amnion which closely ensheaths the body of the embryo. In doing this it is best to commence at the region between the heart and the tip of the head where a couple of fine needles may be used to tear the amnion. Its anterior portion may then be seized with fine forceps and pulled backwards over the embryo’s head. The operation is simplified by carrying it out immediately after submerging the embryo in fixing fluid as the action of the fluid makes the amnion slightly opaque and therefore more easily visible. - If however corrosive sublimate be the fixing fluid fine splinters of coverslip should be used for dissecting off the amnion unless this is done prior to immersing in the fixing fluid. The embryo should again be carefully studied during the process of fixation, many details becoming particularly distinct before the creature becomes completely opaque. Finally the embryo should be studied, preferably with the binocular, as an opaque object, and then prepared either for section cutting or for mounting whole.


  • 1 Bearing in mind that steel needles must not be allowed to touch corrosive sublimate solutions. Picric acid solutions are convenient fixing agents to use for D and E.


VII. Six Days

Open freely into the air-space. Carefully tear away part of its inner wall so as to expose part of the vascular area, great care being taken 11ot to injure the latter. Notice the direction in which the vessels of the vascular area converge: this will indicate the direction in which the embryo is to be. found. Work towards the embryo, picking off the shell piece by piece, using blunt forceps. Frequently the escape of the air from the air-space allows the vascular area to sink down and leave a wide space between it and the shell membrane. In other cases however it remains in close contact with the shell membrane and in this event the greatest care must be taken not to injure the vascular area as by doing so the very fluid yolk is allowed to escape and the salt solution rendered so opaque that observation of the embryo in situ is made almost impossible.

Notice that the allantois has increased much in size, that it has become richly vascular and that it is spreading outwards in a mushroom-like manner underneath the serous membrane. It has already spread so far as to cover the embryo nearly completely.

It is best new to remove the shell entirely and to examine its contents as they lie submerged in the warm salt solution (as shown in Fig. 242).

With fine sharp scissors cut through the serous membrane just outside the limit of the allantois, commencing on the dorsal side of the embryo where the allantois is not yet closely applied to the yolksac. It is easy to do this owing to the coelomic cavity having spread outwards well beyond the limits of the allantois. The allantois being new no longer flattened out, by its continuity with the serous membrane all round, its vesicular character becomes apparent, as well as the difference in character of the vascular network on its proximal and distal walls. The relations of the vascular allantoic stalk to the vascular yolk-stalk should be noted: also the fact that the amnion is now widely separated from the embryonic body by secreted amniotic fluid. If the embryo is a well-advanced one towards the end of the sixth day the amnion, which is now muscular, may exhibit periods of muscular contraction during which the embryo is rocked to and fro in the amniotic fluid. These movements must be distinguished from the occasional contractions of the muscles of the embryonic body which also occur about this time though they are much less conspicuous.

After a careful study of the living embryo with the allantois and yolk-sac hanging from its ventral side it may be excised along with a circle of vascular area, floated into a watch-glass and examined alive with a lens or binocular, and then treated with fixing fluid such as Bouin’s solution. The latter brings out the surface modelling which should be carefully studied especially in the region of the gill clefts.

Dissect off the amnion, add more fixing fluid and after superficial fixation renew the llouin’s solution. It is a good plan to suspend the embryo by the yolk-sac so that the weight of the head causes the neck to become somewhat straightened. After the embryo is sufficiently fixed the neck may be cut through and the lower surface of the head studied for the relations of the olfactory rudiments and mouth.

Sagittal sections through the head are particularly instructive at this stage.

VIII. Segmentation

T0 obtain segmentation stages hens which are regular layers should be chosen. In such cases the egg is laid at a slightly later time on consecutive days. As a rule egg-laying is confined to the forenoon and early afternoon and when an egg is due after the end of this period it is retained within the oviduct and not laid until next day. The retention of an egg in this way inhibits the process of the ovulation so that a new egg is not shed from the ovary until the preceding one has been laid.

HISTORY or THE Eco UP TD run TIME or LAYING:--The egg arises as a single cell of the left ovary 1 which grows to a relatively enormous size as yolk is deposited in its cytoplasm. The yolk is of a characteristic yellow colour but in particular tracts the disintegration of its granules into finer particles gives it a white colour. Of this white yolk a mass occupying the centre of the egg is continuous through a narrow isthmus with a tract lying immediately beneath the germinal disc (“ Nucleus of Pander ”) and this latter is prolonged as a thin superficial layer over the surface of the egg. Between the superficial layer and the central mass are a number of thin concentric layers of white yolk.

‘ The right ovary and oviduct which are present in early stages undergo atrophy, never becoming functional. This is probably to be regarded as an ada tive arrangement which has been developed in Vertebrates with large eggs to avoi the dangers which would be involved in the synchronous passage of a pair of eggs of great size, more especially if contained in a rigid shell, into the narrow terminal portion of the passage to the exterior. ' x ‘EGG or COMMON FOWL 515

As the egg increases in size it bulges out beyond the surface of the ovary, becoming eventually dependent from the ovary by a thin stalk at the end of which it is enclosed within the distended follicle. The wall of this is richly vascular except on the side away from the stalk where an elongated patch—the “stigma ”--marks the position in which the follicle-wall will rupture to set the egg free.

When this process (ovulation) is about to take place the thin membranous lips of the oviducal funnel become active, apply themselves to the follicle containing the ripe egg and grip it tightly. The follicle then ruptures and the egg is as it were swallowed by the oviducal funnel. Within the funnel fertilization takes place provided that spermatozoa are present.‘

The egg proceeds now to travel slowly down the oviduct, propelled onwards by the peristaltic contraction of the oviducal wall, the entire passage occupying about 22 hours. As it does so the albumen is deposited on its surface by the secretory activity of the oviducal epithelium. The first to be deposited is rather denser than that formed subsequently. It forms a sheath immediately outside the vitelline membrane and extending in tapering spindle-like fashion for some distance up and down F'<=- 323


Unincubated egg of the Fowl. the oviducal cavity; the two a.s, air-space; alb, albumen; ch, clialaza; s.m,

- , 3 shell membrane. In the centre-—at the apical pole-prolongatlons a’r(" flu” chalazae is seen the germinal disc with the white “Nucleus of Puiulv-1"‘showingllwough it‘.

The envelope of (‘louse albumen enclosing the egg is not merely propelled onwards; it also undergoes a clockwise rotation about the axis along which it is travelling, caused probably by the cilia present on the oviducal epithelium.’ Owing to the prolongations of the albumen in front and in rear of the egg not undergoing this rotation the chalazae become twisted upon themselves in opposite directions.

Layer after layer of albumen (Fig. 223, alb) is deposited round the egg and chalazae until the full size is reached. The character of the secretion then changes and the shell membrane (Fig. 223, 3.-m) is formed. Finally in the dilatedhinder part of the oviduct (“uterus”) the secretion is in the form of a thick white fluid which, deposited on the surface of the shell membrane, gradually takes the form of the hard and rigid shell perpetuating the characteristically “oval” form impressed upon the egg envelopes during the passage down the oviduct. In composition the egg-shell consists of calcium salts infiltrating a slight organic basis of keratin-like material. Structurally the greater part of its thickness consists of calcareous trabeculae forming a fine sponge work. The inner surface of the shell is rough, projecting into minute conical papillae, while the outer surface is covered by a smooth apparently structureless layer perforated by numerous fine pores.

  • 1 The spermatozoa remain alive and active within the oviduct for a period of about three weeks.



Segmentation

If the egg has been fertilized it proceeds with its development as it slowly travels down the oviduct. The process of segmentation is accomplished during this period and consequently the obtaining segmentation stages involves the sacrifice of the parent hens. Owing to the difficulties in the way of obtaining a complete series our knowledge remained for long fragmentary but recently (1910) a number of stages have been described and figured by Patterson which give a fairly complete picture of the process (Fig. 224). From these data we may take it that the early phases of segmentation are based on the normal plan where a meridional furrow appears traversing, or passing close to, the centre of the germinal disc ’i.6. the apical pole of the egg, and is followed by a second meridional furrow perpendicular to the first. In the third phase there is occasionally a regular set of four vertical furrows but more usually the process now becomes irregular (Fig. 224, C). In the next phase also there may be a fairly regular development of latitudinal furrows demarcating a group of about eight cells round the apical pole but typically there is no such regularity. The initial furrows, which make their appearance as above indicated, gradually extend. They eat their way downwards into the thickness of the germinal disc,-never however cutting completely through it. They also extend outwards towards the edge of the disc which however again they never quite reach. The apparent segments into which the germinal disc is mapped out by the early furrows are therefore not really isolated from one another ——there being still continuity between the segments on the one hand peripherally and on the other on the lower side of the disc next the yolk.

Complete blastomeres are first marked off when, about the time the latitudinal furrows appear, division planes make their appearance parallel to the surface, cutting off the small segments in the centre from the underlying deep layer of the germinal disc.

The later stages of segmentation are quite irregular. Division planes make their appearance in all directions by which the germinal disc becomes completely divided up into small segments except on its lower surface and round its edge where there remains a syncytial mass in which the nuclei divide without their division being followed by any protoplasmic segmentation. It is to be noted that the process of segmentation throughout goes on more actively towards the centre of the disc, more slowly towards its margin, so that the blastoderm comes to be composed of smaller cells towards the centre and larger towards the periphery.

The result of the segmentation process is that the original germinal disc comes to be represented by a lenticular blastoderm lying at the apical pole of the egg and corresponding to the mass of micromeres of such a holoblastic egg as that of Lepidostren. The superficial layer of cells become fitted closely together and form a definite epithelium——which is destined to become the ectoderm. The cells of the lower layers on the other hand are rounded with chinks between them representing the segmentation cavity. The lowest of all have the appearance of being incompletely cut oil from what is ordinarily termed the white yolk lying below them but which is really a syncytial layer" full of fine granules of yolk and with scattered nuclei.


FIG. 2'24.———Views of the blastoderm of the F0\vl'.~: egg during segmentation. (Afher Pa.1..tei-son, 1910.)


A, 3 hours after fertilization ; IL 5}, In s. ; U, 4 In-:~'. ; J), 4-5 hrs. ; E, about .-3 hrs. ; l , .5111-s. ;

(L 7 }n'.~:. : H, 8 hrs.


Apparently a few accessory sperm nuclei are usually present in the fertilized eggs and faint traces of abortive segmentation may be visible round them (of. Elasmohranch, Fig. 8, B *, p. 14).

At the time of laying the blastoderm forms a small whitish disc covering the apical pole ot' the egg. Sections show it to consist of an upper layer of ectoderm and of a lower layer consisting of numerous rounded micromeres lying about in the fluid of the segmentation cavity. These micromeres become larger towards the lower face of the blastoderm and they are more crowded together round the periphery. _

It must not be supposed that all newly-laid eggs show exactly the same degree of development. As a matter of fact great variation occurs, one of the chief variable factors probably being the length of time occupied in the passage down the oviduct. Where this time is, longer, as e.g. towards the end of the laying season, the stage of development of the egg when laid is more advanced.


The First Day of Incubation

After the egg has been laid the lowering of the temperature leads to such a slowing of its vital processes that development appears to come to a standstill. If kept at a'low temperature it retains its vitality for‘ a considerable period but makes no appreciable advance in development. If the temperature be raised by incubation the developmental processes are at once accelerated and comparatively rapid changes come about. The blastoderm increases in size, its margin spreading outwards, and at the same time there comes about a distinct difference in appearance between its central and marginal parts-the central portion assuming a dark transparent appearance (pellucid area) which contrasts strongly with the whiter “opaque area” surrounding it. - The examination of sections at once explains this difference in appearance: the more opaque appearance peripherally is seen to be due to the lower layer cells being there closely crowded together.

An important change soon comes over the lower layer cells, in as much as those next to the yolk, in the region underlying the pellucid area, lose their rounded shape, become somewhat flattened and adhere together edge to edge to form a continuous membrane-—the (secondary) endoderm. This appears first beneath the posterior poi tion of the pellucid area; it gradually extends x FOWL—--FIRST DAY’ 1 519

FIG. 225.-~111ustrating three stages of the blastoderm of the Fowl during the second half of the first day of incubation.

a..o, opaque area; mp, pellucid area; lap, head process; mes, boundary of sheet of mesoderin; m.f, medulla:-y fold ; p.g, primitive groove ; 12.3, primitive streak.

forwards and outwards, and eventually is continuous all round with the thickened marginal part of the b1astoderm.1

1 This thickening of the posterior edge of the blastoderm presents in sagittal section a. striking resemblance to a. gastrular lip growing back over the yolk and Patterson (1907) believes that an actual process of involution-—a reminiscence of gastrnlation by inva ination--takes place, It must not be forgotten that any explanation of such 0 scure developmental “phenomena. in Birds must, to be reliable,520 EMBRYOLOG-Y ()l<‘ '_l‘Hln‘ LUWEJ-t VEl~‘tTEBRATES an.

A gradual change takes plzme in the shape of the pellucid area which, up till now circular, a.ssuim,-s an oval or pear shape (Fig. 225, B)——the long axis perpendicular to the long axis of the eggshell, and the narrow end being next the observer when the broad end of the egg-shell is to the left. This narrow end may be called posterior from its relations to the rudiment. of the embryo which appears later. Together with the gradual change in the shape of the pellucid area there takes place the development of the primitive. streak. This makes its appearance usually during the first half of the first day of incubation, as a linear opacity stretching forwards along the long axi.s of the pellucid area in its posterior third. As the first day of incubation goes“ on the primitive streak becomes more and more distinct. A longitudinal groove develops along its middle———the primitive groove-—while on each side of this it forms a ridge, the primitive fold.

If a number of eggs be examined du.ring the first day of incubation it will be seen that the primitive streak, as is commonly the case with vestigial organs, shows extreme variability. More especially its hinder end is commonly bent to one side or the other, or even bifurcates into two branches. At its front end one or both halves of the primitive streak swell up into a slight knob while the primitive groove becomes somewhat deeper and wider.


FIG. 226.—-Transverse section through primitive streak of the Fowl. eat, vet-oderm ; vi-mi, 1-ndoderm; mes, mesoderm; 12.;/, primitive groove.


The primitive streak is shown by transverse section to originate from a linear tract of ectoderm along which the cells are undergoing rapid proliferation, as is indicated by the relatively numerous mitotic nuclei. The cells budded off by the ectoderm are aggregated together in a compact mass along the course of the primitive streak while on each side they become loosened out and wander away into the space between ectoderm and endoderm to take part in forming the sheet of mesoderm.


rest on a firm basis of knowledge of Reptilian development. At the present time however our knowledge of the exact relationship of these clevelopniental stages of Birds to the corresponding stages of Reptiles is not in the present writer’s opinion adequate to form a trustworthy basis for their interpretation.


For a short distance in- the region of its front end the mass of cells forming the primitive streak is continuous not only with the ectoderm but with the endoderm as well: the primitive streak of this region may be defined as a tract along which there is cellular continuity between the ectoderm and the endoderm.

During the latter half of the first day what is known as the “ Head process ” makes its appearance (Fig. 225, B, lap). In a view of the whole blastoderm this has the appear‘-aiice of being a somewhat less distinct prolongation forwards of the primitive streal<——-in front of the knob which marks its apparent front end. 'l‘he study of transverse sections shows that the so-called head process is exactly similar in structure to the primitive streak immediately behind it, except that it is separated fr cm the overlying ectoderm by a distinct split and that there are no primitive folds or primitive groove over it. On its lower side there is perfect continuity with the endoderm—-— as is the case with the anterior part of the obvious primitive streak into which it is continued.

During the same period of incubation there appears the first sign of the surface relief of the body of the embryo in the form of what is known as the head fold (Fig. 227, A, h. f). This is formed by the blastoderm bulging upwards and forwards, forming a projection bounded in front by a steep face crescentic in shape as seen from above, the two horns of the crescent directed backwards. The projection increases in prominence: its front edge soon comes to overhang, the blastoderm becoming tucked underneath it both in front and at the sides, the two horns of the crescent which the fold formed at its first appearance gradually extending farther and farther backwards. The projection is destined to give rise to the head end of the embryo and there are certain important details to be noticed about its structure which can be made out best by the study of sagittal sections.

The region of the blastoderm where the head fold develops is composed of the two primary layers, ectoderm and endoderm, the mesoderm not yet having spread into it. It follows that the head rudiment has a double wall, its outer sheath of ectoderm enclosing an inner wall, quite similar in shape, composed of endoderm. It will be understood that this inner wall of endoderm is continued at its hind end into the flattened layer of endoderm which lies on the surface of the yolk. In other words the endoderm within the head rudiment may be described as forming a very short wide tube, blind anteriorly but opening behind into the yolk. This endodermal tube is the rudiment of the front part of the endodermal lining of the alimentary canal of the adult and is termed the foregut.

Soon after the commencement of the formation of the head fold the ectoderm of the medullary plate becomes raised up into a longitudinal ridge (Fig. 227, A, m. f) upon each side of the median line. Between the two ridges is a groove——the medullary groove: the ridges themselves are the medullary folds: the two medullary folds F10. 227.—Fow1 embryos at about the end of the first day of incubation seen by reflectcd light.

5-6 segments. cr..p, pellucid area‘ aw, vascular area; f.g, foregut; h.j, head; m.f, medulla:-y fold; m..c, nnesoder-n1

A, 3 mesoderm segments; B, no segments yet demamcated; 0, segments ; p.a, proamniun ; p.g, primitive groove; 11.3, pximitive st.reaLk. CH. X FOWL--FIRST DAY 523

are continuous anteriorly. The two medullary folds gradually extend backwards and at the same time they become more prominent and arch over towards one another until at about the end of the first day they meet. It is to be _noticed (h‘ig.‘227, B) that the first meeting of the medullary folds is some little distance back from their anterior end, in about the position in which the division between mesencephalon and rhomhencephalon will develop later. Towards their anterior end the folds remain less prominent than they are farther back with the result that the meeting of the two folds is here con— siderably delayed.

During these later hours of the first day important advances are taking place in the development of the mesoderm. In the first place it is to be noted that the anterior limit of this layer is gradually extending forwards, encroaching more and more upon the proanmion ——the part of the blastoderm in front of the head fold which is still two layered. In the second place the mesoderm becomes considerably thickened and more compact in the region near the median 1ine——— adjacent to the head process or notochord. This thickened portion of the mesoderm becomes divided by transverse splits into a series of blocks——thc mesoderm segments——lying one behind the other (Fig. 227, A and C, m.s). The first pair of splits to make their appearance are placed obliquely, sloping outwards and backwards: they mark the hind boundary of the first or most anterior segment. A little later a pair of similar splits develop a little farther back forming the hinder limit of the second segment, and so on, segment after segment becoming separated oil’ from the still continuous mesoderm lying farther back.

While this portion of the mesoderm is becoming segmented it is at the same time becoming sharply marked off by its greater thickness from the lateral mesoderm lying farther out from the axis. Towards the end of the first day at further important development takes place in the mesoderm in as much as isolated splits appear in it parallel to its surface and these gradually spread and finally become continuous so as to divide the mesoderm into the outer somatic layer next the ectoderm and the inner splanchnic next the endoderm. The cavity which has made its appearance between somatic and splanchnic layers of mesoderm-is the coelome. The portion lying within the myotome, which soon becomes filled up by immigrant cells derived from its wall, is the myocoele (Fig. 228, me). The portion lying farther out, in the lateral mesoderm, is the splanchnocoele (splc). The two layers lying external to this cavity——the somatic mesoderm and the ectoderm —constitute the somatopleure or body-wall: the corresponding layers lying internal to the cavity-—the splanchnic mesoderm and the endoderm——-constitute the splanchnopleure or gut-wall.

While the changes above described have been taking place the blastoderm has constantly been increasing in area and by the end of the first day it forms a cap covering an extent of about 90° at the upper pole of the egg. In the opaque area—--the part of the blaste524 EMBRYOLOGY OF THE LOWER VERTEBRATES CH.

derm lying outside the boundary of the pellucid area---there are present the same layers of cells as in the pellucid area-—-the ectoderm, which extends. farthest peripherally, the endoderm which passes into «W thick ¥°1kSYI1cWe1layer vermherallv <2ermina1 wall. and the


Fig. 2‘28.—- Tansverse section through the bocl y of a Fowl embryo about the end of the tiist day of incubation. c. act, cctoderm; end, endodurm; me, myocoele; my, myotomu (lll6S0(lCI‘ll1 segment); N, notochord;

'n.'r, neural rudiment; som, somatopleure; spl, splanclnioplenre; splc, splzmclmocoele.

mesoderm the outer part of which is still unpenetrated by the coelomic split. The part of the opaque area where inesoderm is present assumes a very characteristic mottled appearance (Fig. 227, 0, (M2) caused by the rudiments of blood-vessels and blood: hence the name vascular area which is given to this part of the blaste derm. When the embryo has reached the stage with about seven

inesoderni segments the secretion of fluid (plasma) commences within the blood islands.

13313 Sacond DAY or INCUBATIQN:

opened during the second day of incubation is seen in Fig. 229. The blastoderm has increased considerably in size and now covers about 110". The pellucid area has assumed a somewhat fiddle-like shape.

On examining the excised blastederm about the commencement of this day it is seen that the formation of the head fold has progressed considerably and the head rudiment projects more conspicuously above the general level of the blastoderm. Within the head rudiment the foregut can be seen and it is noticeable that it stretches farther back than does the outer wall of the head rudiment. . In other words the head fold of the endoderm has spread farther back

FIG. 229.~—-Egg of the Fowl about the middle of the second day of incubation.

a..o, circular opaque area. the dark pellucid area, with the rudiment of the embryonic body lying along its axis.

In the centre is

-——The cg;-,ne1~a1 appearance of an seoegggi X F OWL-—SECON D DAY 525

than that of the ectoderm. This is brought out clearly by a sagittal section such as that shown in Fig. 230. Such a section a.lso brings out the fact that while the greater part of the portion of blastoderm tucked in beneath the head of the embryo is two-layered (proamnion), there being no mesoderm present, this does not apply to the farthest back part of the fold. Here, in the wide space between cctoderm and endoderm, mesoderm has penetrated which will give rise to the pericardiac wall and the heart. The medullary folds have met over a ' considerable extent but still remain separate at their extreme front ends as well as over the whole extent which will later form the spinal cord. Here they bound a deep neural groove. Towards their posterior ends the two medullary folds diverge to pass on either side of a lance—shaped area (rhomboidal sinus) which they enclose by converging towards one another behind it. Along the centre of the


Flu. 230. -Diagramattic sagittal section through anterior end of Fowl embryo with 15 segiiieiits.

am, rudiment of amnion; hr, brain ; er!-, ectmle-rm ; um.-, endocai-dinm; end, endoderm of yolk-sac; _/15;, l'm'v_~,r11t2 h._/‘, p0n'l'-i'I‘l')1' limit or lH.':ul rum of uctoderm; mo, myocardinm; N, notochord; pa, ect.-o«‘lm-In of pm.-nnninn ; -vile, split!|(7lI1I()(1()Ble.

floor of the rhomboidal sinus the primitive streak is still_ visible separated by a knob—li.l<e elevation from the part of the primitive streak which lies farther back.

The mottled appearance characteristic of the vascular area is now seen to be continued inwards, though much more faint, across the pellucid area to the body of the embryo.

An embryo with about ten segments is shown in Fig. 231. The pellucid area is still somewhat fiddle-shaped with the body of the embryo lying along its axis. Apart from the increase in number of the mesoderm segments the most conspicuous advances in development are in the central nervous system. The medullary folds have met and fused together to enclose the neural tube except towards their hind ends where they still bound the rhoinboidal sinus on each side. The forebrain region is greatly dilated, its projection on each side being the optic rudiment (am). It will be noticed that a slight notch in its wall in the mesial plane anteriorly indicates that at this point the two neural folds have even yet not completely fused. Posteriorly the neural folds seem to be continuous with the lips of the primitive groove. A faint continuation forwards of the primitive groove may be seen in the floor of the rhomboidal s1nus.


Flu. 231.~-Bla.stml¢_-1'In with l"m\'l .wJl"1J]..‘.‘| will: :n.bm|t l() or ll 1m__e.

am, Vtl.s‘(:lll1ll':lI‘o-.1 ; _/'._:/, fur-e;;I11.: la, Imiul : m.‘/1, ma-«lnll:n'_\' rule! ; m..~:. rm-:-.mlerIn .\'I'_4_1_'lll6l|t-2 u./', upliv l'lllllllH_'lll.I /W. ]rmunminn.

Important mesoderrn features are to he noticed. The mottled appearance of the vascular area produced by the rudiments of bloodvessels developing in the splauchnic mesoderin is conspicuous. The formerly isolated vascular rudiments (white in the figure) are now becoming joined up to form a network and the network can be traced —less distinct and on a smaller scale—-across the pellucid area. At its anterior and inner corner the network is continuous with a short and wide vessel which slopes obliquely forwards and inwards and disappears beneath the hind end of the foregut (shown more clearly in Fig. 232, 72.22). This vessel is the rudiment of the vitelline vein, which drains the blood from the vascular area towards the heart. Another conspicuous vessel rudiment is the terminal sinus-a marginal vessel which bounds the vascular area externally. In front of the head of the embryo is a somewhat rectangular area of the blastoderm distinguished by its being very transparent (Fig. 232, pa). This is the proamnion—- -its transparency being due to the fact that the mesoderm has not yet spread into this region of the blastoderm. On each side of the head of the embryo the surface of the blastoderm bulges upwards into a dome-like swelling (Fig. 232, splc). This is due to a precocious splitting of the mesoderm in this region to form a large coelomic space. The bulging appearance is produced by the coelomic space being tensely filled with fluid. The raising up of this region of somatopleure is preliminary to the formation of the head fold of the amnion.


FIG. 232.~ llmul oi" l“u\vl «-mhr_\'0 of smm-. Hiilgif as that s'hu\\'n in Fig. 231, more highly inugnilieil and Ht-‘vll by tnuus-initlwl liglit.. f.g, foregut; 1!, heart: h.._/',himlerlimil ofhcml fold ()fvC.l'(_)¢ll'l‘lll; inf, infumlibnlnm; m..s, mesoderm segments; N, notochord; 0.7-, optic rmliuu-nt: ,.u, ]nn:unniuIl ; splc, patent portion of splanchnocoele containing coelomic fluid; -mi. vitellim-. \ win.


By turni.ng over the excised blastoderm and examining it from below or by staining and then examining it in dorsal view by transmitted light (Fig. 232) it will be seen that between the two coelomic spaces there lines a A-sha_pe<.l structure. The two diverging limbs of the A posteriorly are the vitelline veins already alluded to (cw), While the median portion (H)——a straight tube passing forwards beneath the foregut-—is the rudiment of the heart and ventral aorta. It will be noticed that the two vitelline veins when traced backwards from the heart are seen to fit round the tunnel-like opening of the foregut. In the forcbrain region is seen the downwardly projecting pocket of its floor—the infundibulum (Fig. 232, v}nf)———and extending -back from this in the middle line the notochord (IV). On each side of this posteriorly are seen the mesoderm segments (m.s).


In a slightly more advanced embryo with about fifteen mesoderm segments the tucking in of the blastoderm under the head has proceeded considerably further. The neural tube has become closed in entirely except for the slit-like remnant of the rhomboidal sinus posteriorly. The optic rudiments projecting prominently from the forebrain on each side and beginning to be narrowed slightly at their base give the brain a conspicuous T-shape. The wall of the brain in its posterior region shows a series of puckerings one behind the other marking it off into a series of what used to be called brain “ vesicles.” Of these the anterior one, the largest and most distinct, is destined to become the niesencephalon while those behind it enter into the formation of the rhombencephalon. The latter are often interpreted as vestiges of a once present segmentation of the brain, but are regarded by the author of this volume as being adequately accounted for by the active growth of the l.rain within its confined space, aided possibly by the varying consistency of the mesenchyme outside it (see p. 101).


On each side of the head region posteriorly, just in front of the first.obvious mesoderm segment, the rudiment of the otocyst has made its appearance as a cup-like depression of the ectoderm.


The heart, growing in length more rapidly than the neighbouring parts of the body, has been forced into its characteristic bulging outward on the right side. The first traces of haemoglobin are making their appearance in the posterior portion of the vitelline network.


An important new feature becomes visible about this stage in the form of a whitish line on the bulging roof of the splanchnocoele on each side. The lines in front curve in towards one another, meeting in front of the proamnion and sweeping back in a wide curve on each side. This line is the first rudiment of the amniotic fold. As the fold becomes more and more prominent it bends backwards and inwards, arching over the head region, and towards the end of the second day (Fig. 233) forming the anmiotic hood which ensheaths the head portion of the embryo.

Many of the important details in the structure of the second day blastoderm can only be made out by the study of series of transverse sections. In studying the stage new under consideration it is advisable to begin with a section taken from about the middle of the total length of the embryo such as that represented in Fig. 234A. The blastoderm some little distance away from the median line of the embryo is seen to consist of the usual two double 1ayers——-the somatop1eure(som) composed of ectoderm and somatic mesoderm and the splanchnopleure (spl) composed of splanchnic mesoderm and endo FIG. 233.-—Blastoderm and embryo Fowl with 18 mesoderm segments.

u,.e, hackgrowing edge of amniotic hood; asp, pellucid area; um, vascular area; 1!, heart; of, 0t0(‘,_yst,; sa, sero-amniotic connexion.

derm. In immediate contact with the lower surface of the endoderm in the complete egg there would be the yolk. I'n the splanchnic mesoderm overlying the endoderm are seen the blood-vessels of the vascular area. When traced inwards towards the mesial plane the two layers of mesoderm are seen to come together to formthe narrow protovertebral stalk or nephrotome which joins up the lateral mesoderm to the mesoderm segment. Immediately above the nephrotome, between it and the cctoderm, is seen the rudiment of the arehinephric du.ct-——a rod of cells which is gradually extending tailwards. i In the centre of the section is the neural tube (s.c) with its thick walls and the solid notochordal rudiment (N) -lying immediately


FIG. 234A.——Transverse section through the middle of El .*i('(‘.(')nIl~(l:l}-‘ Fowl embryo ' (15 segments).

4, p.-iiretl dorsal aorta; a..n.d, fll‘Clllllf‘])lll‘l('f duct; eat, t-(‘totlvrni : end, u-n«l0tl(‘,1‘ln ; my, myotome; N, notoeliord; s.c, spinal cord ; .~'u‘m, somatnpleure; spl, splam-hnopla.-m'e; .s-pin, splanclmoeoele.

below it. The blood-vessel (A) on each side between nephrotome

and endoderm is the dorsal aorta which is at this stage double. Working back towards the tail end of the embryo it is seen that

subsequent sections show less and less advancedstages of development


FIG. 2343.-Transverse section through a second-day Fowl embryo jllst lwliiml the binder limit of the l'(')1‘e;,5l1t.

A, dorsal aorta; and, endoderm; -‘my, myotome; N, not.oeho1.-d; .-.r_.-, spinal em-cl ; min, somatoploure; Sp], splanchnopleure; .~.-plc, splanehum-oele; r, \"('_5.‘€."~‘.(!l.\' ()f\'asc||I:1[‘;1rea,

in concordance with the fact that development proceeds from the head end tailwards. Thus the neural tube opens out by the slit—like rhomboidal sinus; the archinephric duct disappears; the notechord passes back into the undilferentiated tissue of the primitive streak. On the other hand the examination of sections farther forward towards the head region brings into view various important further developments. Such a section as that shown in Fig. 23413 illustrates clearly an early stage in the folding off of the foregut from the cavity of the yolk-sac-——a fold of splanchnopleure growing inwards on each side below what will become the foregut. The large vessels seen in the splanchnopleure external to the fold just mentioned are tributaries of the vitelline veins, and a few sections farther forwards they would be found to be united together to form the main vitelline vein on each side.


As the series of sections is traced forwards the two folds of the splanchnopleure are seen to approach one another and finally to meet and undergo fusion, so that there now exists a foregut cavity shut off (as seen in transverse ._section) from the yolk-sac, the walls of the two structures being still connected by a median vertical partition formed by the fusion of the endoderm from the two sides. A little farther forward this partition disappears from ‘the section and the foregut as seen in section (l*‘.ig'. 2340) is quite isolated from the endoderm of the yolk-sac wall. The vitelline veins have also fused to form the tubular heart. It is seen that the splanchnic inesoderni ensheaths the endothelial wall of the heart (em) on each side and that where it -does so it is somewhat thickened (me) as compared with the same layer in the region overlying the yolk“-sac. This localized thickening of the splanchnic inesoderm is destined to give rise to the entire thickness of the heart wall except the lining endothelium. It is seen to be continuous with the extracardiac portions of the splanchnic mesoderm by the dorsal (d.mc) and ventral Inesocardium (mnc).


FIG. 2340.~—Transverse S(‘.(‘.t-iml of a .~u-cowl-day Fowl embryo pa-1..<siiig through the I'udiment of the ll(‘iU.'l.

A, dorsal aorta : «l.nu°, 1l()l'H:ll Im-snmmliuiii 1 rm". l‘lIIl()l'L'l.l'|lllllll : crud, cnclorlc-rm : jig/, foregut; ma, myocardium; s.c, spimil cord; so.m, smiiatic nic.-‘ode’-rin; sp.m. spluncliiiiu ino.<o(lo'1'1ii; split, splanclb nocoele; v.m«_-, \'t'-ntl‘£ll mcsocardium.


Traced forwards through the series of sections the heart is seen to narrow in calibre as-it tapers off into the ventral aorta. Towards its front end the latter gives off a large branch on each side which 532 EMBRYOL()(_}YOF THE LOWER VER’1‘EBPA'l.‘ES (:11.

passes outwards and upwards round the foregut to become continuous with the dorsal aorta. These two hoop-like vessels which connect up ventral and dorsal aortae are the first pair of aortic arches.

Still further forward the region of the forebrain and optic rudiments is reached (Fig. 2341.)).

Owing to the folding oil‘ of the head rudiment the section of the head itself appears completely detached from the blastoderm and the latter is begiiining to form a depression which will later become more marked and in which the head will lie. In the blastoderm it

s will be noticed how away on each side it shows the normal four layers

of cells——-ectoderm, somatic mesoderm, splanchnic inesoderm, endoderm—While on the other hand in the region 1i11_de1-lying‘ the head of the embryo it is only two layered the mesoderm being here absent.

the Optic rudiments.

cot, ec.-tmlm-m : rm/, mulmlu-I-in ; /:H'S, nw.~:mu-liyiiie; n.r, optic. rudiment ; ,:u, pi-oanmion ; splr, n‘];l.'u|('l|ll0C0!..'lI'; Hull, roof ofthalamencephalon.

This two-layered region of blastoderm is the proamnion before alluded to.

The head itself is occupied almost entirely by the brain rudiinent ———the thalanieneephalon in the centre (tltal) continued outwards on each side as the optic rudiment (oxr). For the most part the external ectoderm is closely apposed to thesurface of the brain but dorsally the former is commencing to recede from the latter, the space between the two being occupied by mesenehy me (mes).

The Third Day of Lncubation

During the later hours of the second and earlier hours of the third day of incubation there take place a number of important changes which render this period perhaps the most i.nterest'ing of all to the morphologist. For the student who is training llllll.S(..‘.ll" practically in the technique of embryological observation tliem is no finer material than that afi"orded by liinl embryos of about this :.1.g<: for l_o:1.1‘ning one of the most important parts of that technique namely the interpretation

of serial sections. OWL--——SEC()N 1) ANI.) '.l.‘H’[R1) DAYb 533


It is mlvisable to make {L (;:.L1‘(3l"111 sbmly of the ana.t;omy of an embryo of about; the Htztgia shnxvn 111 I919‘. 235 01‘ Ii-.’.f_’.(-3,1

Flu. 255.3. 'l'hir«l—«l:ty l*'u\\'l t‘|lliH'_\'u with thv \'u..~"u:1l1:n|':u"u.

u..r, o.-«L-"v ul' :1nminII‘. P5. I-_\'I'*: .‘.’, ln~:H'l; wt, u1uc_\'.w-t; .-.u. >'I'l‘0-flllllliutiv t'Ullll\‘_\iUl|; .~'.I, sinus ternninali. ; I.‘/', Iziil-fnlulg ran. \'il».*llim- .-n'tc_*1-y; . ;u.n'l.iun u!’ splzuurhlmlalelu-v im-'0lnt.eil to form :1

rm-ass Inmul Llw he-:ul of the vInln'yn.


‘ It is custonmr_y In mmml. lI':1)1.w'\':)1'. 0 sections with the posterior or tai1\':rd surface of the section next lilv sli.-In : nulls:-ql1t_‘lltiy the figures represvnt the sectinns as seen frmn in front and Lhv sidu nl' '..:u'.h Iig1u'v tmv.-n-«is H11‘ ri;;l11-lnnici side ml‘ the pzige c<2z't'e.~‘pn|u«ls tn the M't."hu.n«l side at" the crnlrryu. 534 EMBRYOLOGY OF THE LOWER VERTEBRATES CH.

On opening the egg it is at once seen that the blastoderm has increased considerably in size, the outer limit of the opaque area having spread downwards as far as about the equator of the egg. The vascular area has also increased considerably and is still bounded by the conspicuous terminal sinus which anteriorly turns inwards and passes back parallel to the corresponding part of the sinus of the other side to open into the vitelhne vein close to its inner end. Of these two veins which run parallel to the long axis of the embryo the right is reduced in size and eventually disappears.

The yolk has assumed a more fluid consistency; the proportion of white yolk has increased; the albumen has shrunk considerably in volume, and the air space has increased correspondingly.

The free edge of the amniotic hood (Fig. 235, a.e) has grown back so as to ensheath all the head and anterior trunk region of the embryo. It follows that when examined in sritw. the front part of the body is seen through two layers of somatopleure. Of these the outer—the serous n1en1brane———forms a kind of roof which passes outwards all round into the general blastoderm. The inner—the true aInnion——c1osely invests the head end of the embryo and is visible in profile as a sharp line immediately outside the outline of the head itself. Anteriorly the amnion very often seems to be prolonged into a sharp peak (Fig. 235, s.a): this is the sero-amniotic connexion.

The free edge of the amniotic fold, somewhat arch-like in outline, may die away posteriorly (Fig. 235) or it may be already continued into the lateral and caudal parts of the fold (Fig. 236)——but even if present these are still low and inconspicuous as compared with the headward part of the fold.

As regards the body of the embryo it is seen that the folding off of this from the yolk is proceeding rapidly. The head and anterior part of the trunk project freely and, correlated with this and with the ventral flexure of the head region, the latter has come to lie_ over on one side, usually the left, so that it is seen in profile when the blastoderm is looked down upon from above. At the extreme hind end the tail region is also seen to be in process of becoming marked off from the blastoderm by a tail fold (Fig. 235, tj) of the same nature as the head fold. Similarly the trunk region between the regions of head and tail fold is becoming demarcated from the blastoderm outside it by a lateral fold (Fig. 236).

The body of the embryo has increased considerably in length and this growth in length is particularly active towards the dorsal. side of the embryo where there is greater freedom from the clogging effect of the yolk. The result of this difference in rate of growth between dorsal and ventral sides is that those parts of the embryo which are detached from the general blastoderm assume a strong flexure towards the ventral side. This is particularly pronounced in the head region, the head being completely bent upon itself so that the front end of the brain is reversed in position, what was its ventral side having come to be dorsal.

The mesoderm segments have inc1'ea*‘ed in number there being now about 25-30 metotic segments and those towards the anterior end are showing a considerable amount of dorsiventral growth. , In some embryos (Fig. 236) the ‘series of definitive mesoderm segments is continued far into the head region by what appear to be the ghostly vestiges of formerly existing segments (see pp. 210,211)


FIG. 236.—~Third-day "Fowl embryo (N o. 47) viewed as a transparent object.

cw, edge of amnion; am, amnion; E, eye; H, in-._a.rt;; m.s, mesoderm segments; ut, otocyst; s, indications of preotic mesoderm segments ('9); u,a,, vltellino artery; me, visceral cleft II; *, portion of splanchnopleure bulging downwards into the yolk, forming a recess in which lies the head of the embryo.


The central nervous system has made important advances in development. The brain shows a relatively large increase in size as compared with the spinal cord : thalamencephalon, mesencephalon and rhombencephalon are marked off by definite constrictions-—tl1e mesencephalon being particularly prominent at the bend of the head. The greater part of the roof of the rhombencephalon is assuming its definitive thin membranous character. The three great organs of special sense have made their appearance. The eye (E) forms a large conspicuous cup-like structure lying at the side of the forebrain. Its rim is cleft ventrally by the choroid fissure (Fig. 236). Its mouth is partially blocked by the round lens rudiment. The otocyst (at) is also conspicuous—-—a pea_r-shaped sac, its narrow end dorsal, lying at the side of the hind brain. The olfactory organ is represented by a slight dimple of thickened eetoderm near the tip of the head.


The side walls of the foregut are perforated by visceral clefts. The series of these develop from before backwards and by this stage three have commonly appeared—c1efts I, II, and III of the series.


It is perhaps the vascular system which shows the most interesting features during the third day. The heart is still in the form of a simple tube, but its active growth in length has caused a great increase in the curvature which was already pronounced about the middle of the second day. Its y-like curvature is shown in Fig. 236. At its morphologically front end the heart is continued into the .ventral aorta and this at its end gives off a series of vessels, the aortic arches, which pass up round the sides of the foregut between adjacent gill-clefts and open dorsally into the aortic root which lies just dorsal to the clefts. Like the clefts themselves the aortic arches develop in sequence from before backwards and by this stage arches I, II, and III have made their appearance (Fig. 241, A).

At its front end the aortic root can be traced for some distance into the head as the dorsal carotid artery (Fig. 241, A, d.c). Posteriorly the two aortic roots become hidden from view by the myotomes but the study of sections shows that they have here united to form the unpaired dorsal aorta. Still farther back this vessel again becomes paired and a little behind the point of bifurcation each of the branches gives off a large vitclline artery (ea) which passes outwards at right angles to the axis of the body to supply the vascular area.

Of the venous system the most conspicuous components are the great vitelline veins (Fig. 241, A, 72.7)) which, receiving numerous branches from the vascular area, pass forwards converging towards one another to form by their fusion the hind end of the heart. Examination of the vascular area shows that the branches of the vitelline arteries and of the veins accompany one another in their ramifications. In the living condition, in which all these arrangements of the vascular system should be studied, the arteries are seen to be more deeply coloured and more conspicuous than the veins. The two vitelline veins by their -fusion form the hind end of the tubular heart and on tracing this forwards a. somewhat Y-shaped vessel is seen opening into it laterally. The stalk of the Y which is very short, though showing considerable variability within its limits, is the right duct of Cuvicr (Fig. 2-11, A, d.C). The branches of the Y are the cardinal veins. Of these the posterior (p.c.'v), coming from the region of the kidneys, is only visible for a short distance, being soon hidden as it is traced backwards beneath the myotomes. The anterior cardinal vein (a.c.v_) on the other hand can be traced forwards for a long distance into the head from which it drains the blood back towards the heart. It will be noted that here in the embryonic Bird we find exactly the same arrangement of main veins—-—duct of Cuvier, anterior cardinal and posterior cardina1— as is characteristic of-the adult condition of lowly organized fish-like Vertebrates.



F In. 237A.——'l‘ransverse sections through thircl-«lay Fowl (:lnln'_\'t_). (Partly based on figures by Duval.) A, '|‘hrough the hinder part. of the trunk region.

A, dorsal aortne; um, annniotic folds ; «mi, «-ndoderm ; 'm._c/_. xnyotume ; s.«', spinal cord; sum, sonmtoplenre ; spl, s]alams-lannplenre; splc, splanchnocoelt-r.


For the study of such details of structure as cannot be made out in the whole embryo the most useful sections are series cut transversely to the long axis of the trunk region. These should be supplemented by series parallel to the sagittal plane in the head region.

It is well to commence the study of the transverse sections with one through the binder trunk region, about the level of the vitelline arteries. Such a section is depicted in Fig. 237A.

In comparing this section with a corresponding section through the second-day chick (Fig. 23-1A) the same general features will be recognized--—the differences being mainly differences in detail. The most conspicuous of these is caused by the development of the amniotic fold of the somatopleure which rises up on each side, arching towards ‘the median plane over the dorsal side of the embryo (am). Traced forwards through the series the amniotic folds of the two sides are seen to meet and undergo fusion so as to give rise 538 EMBRYOLOGY OF THE LOWER VERTEBRATES CH. rte/the inner true amnion and the outer false amnion or serous inembrane: the T6fif1“e"r"'"'contin11o11s at its inner '"ed‘g‘é"'\vitl1 the somatopleure of the embryo’s body, the latter at its outer edge with that of the blastoderm. It will be readily seen that the space between true and false amnion is morphologically part of the splanchnocoele. It will also be realized that both true and false amnion being somatopleural in nature are composed of ectoderm and somatic mesoderm but that the relative position of these two layers is reversed in the amnion compared with the false amnion. Important changes have taken place in the mesoderm. The mesoderrn segment is no longer connected with the lateral mesoderm the nephrotoine having become converted into renal structures———the archinephric duct and mesonephric tubules. The relations of these will be understood by referring back to the general description of renal


FIG. 23713.—--'l‘r:msvcrs:- section just behinzl the point of union of the two vitelline veins.

A, dorsal aorta ; um, amnion ; r~, umllls .'.lrt,m"i0suS ; err, m-1.m.h-run; emf, enteron ; ,/Lu, false amnion or scrolls membrane; my/, inyotome; N, ]I(_)l.-U(:l1UI‘Il; .s-.e, spinal coral: sn, sc-re-amniotic isthmus; sum, somatopletire; cpl, splanchnoplenru; .s-pic, splmuzlinocoole; I’, \'«-nlrivh-; 42.:-, vitelline wins; y, yolk.

organs in Chapter IV. (p. 254). The inner wall of the segment has lost its epithelial character and broken up into a mass of actively proliferating mesenchyme cells. Many of these cells will wander away in amoeboid fashion and settle down round notochord and spinal cord to form the protective sheath in which eventually develops the vertebral column. Collectively these arnoeboid cells constitute the sclerotome which is therefore much more diffuse in its origin than in the lower vertebrates illustrated on p. 285.

Certain blood-vessels are visible in the section. In the splanchnic mesoderm of the yolk-sac numerous vessels of the vitelline network

are visible : over the mesonephros may usually be seen the posterior.

cardinal vein, while on each side of the mesial plane ventral to the notochord are the two dorsal aortae.

e As the series of sections is traced towards the head the most conspicuous change is the incri:-asing asymmetry due to the body of the embryo coming to lie over niore and more upon its left side. Fig. 237]} represents a section just behind the‘ posterior limit of the foregut.


The body of the embryo lying over on its left side is closely invested by the amnimi (run) while over this lies the thin roof (_/lam) constituting the serous mcnibrane. At set the two membranes are united by the sero-amniotic connexion. «In the mesoderm of the two folds of splanchnopleure which are approaching one another to floor in the alimentary canal (ant) are Hl‘l_'ll the two large vitelline veins (72.22). The ventricle and the cones are seen out longitudinally in the wide coelomic space lying to the right of the body of the embryo.

A section a little farther forward in the series has the appearance shown in Fig. 2370. The definitive gut (ent) is completely separated at this level frorn the yolk-sac, and corresponding with this the two vitelline veins, which in sections farther back lay one on each

FIG. 237C.—-T1‘£tl1S\'el‘.<U .\f(‘(_‘llHll :1. little in front of the hind «_-ml of the lic:n't..

mu, mnnion: .-l,«l0rs.-il aor-tn ; d..r, «incl us \-mu)sii.s' : «uni, ulinu-m.-u'y 4-.-uml : _1'.vun, i'.'ilso~ mnnion ; /-i.l, 8-Ilte!‘i0l' 1i\'t-1‘ 1'luli1IwI1t ; li.;’, }no.~'.l'erioi' «lit't.o ; N. nolnvlionl : I-.r.r, ])().\'lI'l'lU[' (‘:ll'lllll:ll win ; sum, somatoph-nrv; :=_n/, spl.-in<:lmople-uri: ; splc, splanclinocoele ; I-'_, \'l'lIll‘l(‘lt.‘.

side of the yoll<—sta1l<, are now completely fused into a large median vessel, the ductus venosus (dxv), which is simply the backward prolongation of the heart. The posterior liver rudiment, a blindly ending pocket of the ‘gut-wall projecting forwards ventral to the ductus venosus, is seen in the section figured (Z222), although its communication with the gut-wall is no longer visible, lying as it does several sections farther back. At this level however a second pocketlike outgrowth of the gut-wall has made its appearance (l7§.1). This is the anterior liver rudiment. It will be noticed that it lies dorsal to the ductus venosus. In the coelomic space ventral to the ductus venosus and liver rudiments, and quite isolated, is the rounded section through the ventricular region of the heart (V).

In the sections studied so far the body-wall of the embryo is widely Open on its Ventral side—the opciiiiig l)elllf_,"lml.1I.l(l€(l by the recurved edge along which the soniatopleure ol’ the body is continuous with the non-embryonic region of the somatopleure forming the amnion. As however the _foli_iing oil‘ of the embryo progresses the edge alluded to grows inwards and the opening bounded by it becomes reduced in size. It will be gathered readily from Fig. 237D that through the opening in question the splanchnocoele, included within the definitive body of the embryo, is continuous with that part of the coelome which lies outside (extra-embryonic coclome). In the section figured the heart is seen to be cut through in two places. Reference to the figure of the whole embryo (p. 535) will show that the piece of heart which lies towards the leit side of the embryo (at) is the atrium, while that on the e1nbryo’s right (0') is the ventricle or conus. In the section iigiiiwl a large blood-vessel (d.0) is seen out longitudinally in the sornatopleure. By tracing this vessel through neighbouring sections it will be found to open at its ventral end into the atrial part of the heart while dorsally it splits into the two cardinal veins-—anterior and posterior. These relations show the vessel in question to be the duct of Cuvier. The only other point calling for special mention in the section figured is that the ventral part of the pharyngeal cavity projects outwards upon either side: this dilated ventral part of the pharynx forms the rudiment of the lung.



Flu. 2371),——'l‘r:i.nsvm-so section a short distance behind the front end of the heart.

.1, dorsal aorta ; am, amnion ; at, atrium ; l..', eonus : «(.!.', duct oi‘ (iuvier: f.«.¢.m., false amnion ; N, notm-hord; 1'-h, pharynx; .-om, sonuttopieure; .-pl, spinnohnopleure; .5-pl:-, splanchnocoele.


In the region in front of the heart the dorsiventral depth of the body of the embryo becomes comparatively suddenly reduced and in the Vacant space within the amnion so provided there appears a_ new structure quite detached from the rest of the section. The structure in question is a section through the recurved tip of the head (see figure of whole embryo). In Fig. 237E this shows the thi.c.k-wallmel forobrain ( with its wide ventricular cavity while upon each side and ventrally’ there is seen a localized thickening (olf) of the ectoderm: this is the dimple-like 1 udi o 4'

of the section there is seen in its centre the wide pharyngeal space and on the embryo’s left side the pharyngeal wall projects out to the ectoderm as an endodermal pocket--~the rudiment of the second visceral cleft (12.0.11). Immediately ventral to the pharynx is the ventral aorta (o.A). On the left side of the embryo the aortic root (am) is seen immediately dorsal to the pharynx, while on the right side--—the section not being accurately transverse—~ ya hoop-like aortic arch (a:.a.III) is seen passing dorsalwards round the side of the pharynx from ventral aorta to aortic root. The large vessel lying dorsaland slightly external to the aortic root (a.c.v) is the anterior cardinal vein. Traced tailwards it is found. to open into the dorsal end of the duct of Cuvier. The neural tube (lab) is seen to have a thin roof and widely expanded lumen indicating that it is now passing into the region of the hind brain.



FIG. 237E.-—-—Transverse section passing through the second visceral cleft and the ' olfactory rudiment.

a.a.IIl, third aortic arch: a.r:.2v, anterior cardinal vein; am, amnion; a.r, aortic root; f.am, false amnion; f.b, forebrain; h.h, hind brain; olf, olfactory rudiment; 11/l, pharynx; ‘I'.A, ventral aorta; no.1], second visceral cleft.


In tracing the series -of sections further forwards it will be realized that the front part of the head region is, owing to its reflexed position, actually being traced in a morphologically tailward direction. In the section figured (Fig. 237F) the reflexed portion of the head is cut at the level of the eye rudiments (opt) which are seen to be in the optic cup stage with the inner or retinal layer


y organ. To return to the am part“ .



‘ It will be realized from an inspection of the figure of the entire embryo that the recnrved part of the head is reversed in position. Its ventral side lies therefore in the figure towards the right. 542 EMBRYOLOGY or THE Lowna VEI-LTEBRATES en.

distinctly thickened as compared with the outer or pigment layer, and with a narrow optic stalk passing to the thalamencephalon near its floor. In the mouth

of the optic cup is the lens but this is seen better a few sections farther on in the series.

Turning to the other Pf half of the section it is

seen that it is no longer connected with the extraembryonic soinatopleure:

in other words the series
   e     of sections has now passed

FIG. 2371-‘.—-—.Transverse section passing through the ' the binder limit -of the rmlunents of the eye and otocyst. hemlfold of the SOmatO_ I (l.('.I‘:{|.llt8['ll-ll"(‘fll‘(illHll‘\'|.‘lll; l:.h, hind brain; .."\'t|1nl«-t.-.lu.n-«I; pleura The pharynx opt, optic cup. of. nt.u('_\ sl , 1:14, ph.'-u'_\nx, (no.1, lust \'1.~'(-er:c1 cleft; aum, ventralearotid. Passes out as 3' Pocket

on each side towards the ectoderm-—4the rudiments of the first pair of visceral elefts (11.0.1). The neural tuhe has become greatly increased in size forming the hind brain with its widely expanded cavity——-the -fourth ventricle. On each side is a large thick-walled sac—-—the otocyst. Examination of


Pin. ..'.‘37(:.- -—'|‘rn.nsv(-rse section passing thrnugln the (*._\'v and just in front of the otocyst.

1I_.I'. 4', il.lli;(‘]'i()!‘ (‘:ll‘(llll:ll Vein : I(..:‘, :u_n'ti(_'. run! ; 41,:-«(,«lm'.~4:nl c-:n-utitl :u't«'I'y; _I[tHI(l. g:u|;;:lin|| of I-i_L,-'l1H'1 H-,-mi,-.1 m-rm: /uh, hind hr.-Lin; lm. pit.-niI_.-n-_\' hotly; .\'_, lllrl_.H(:ll()l‘ll; pin hlmr_\'m.'; pin, ]:ine:ul nr:,r:m; Hull, 1h:'1l:uumn--‘plmlnn: I‘.v'.l. i'l]‘.~}i \i.~:---ml rl--ft.

neighbouring sections shows that it is still connected with the outer skin by a narrow neck. ,In the spongy connective tissue which forms packing between the various organs are seen a number of blood-vessels such as ventral and dorsal carotids and anterior cardinal veins.


As will be gathered by sliding a straight-edge forward over the figure of the whole embryo, its edge parallel to the plane of the sections, there comes a point in the series where the sections through the reflexed part of the head and the rest become continuous. This happens as soon as the deep niche in the bend of the head is passed. Such a section is represented in Fig. 2370.. Comparison of this figure with the preceding one will make clear the fact that the extreme ends of the section are both “of them morphologically dorsal. The brain is cut through twice—on the right of the figure is the hind brain while on the left is the thalamencephalon distinguished by the pocket-like rudiment of the pineal organ (pin). The thin optic stalk lies outside the section, but the structure of the optic cup otherwise is well seen. The lens is in the form of a closed vesicle which has by this stage become completely nipped off from the external ectoderm. Immediately ventral to the thalamencephalon is the pituitary involution cut transversely- The section passes through the ganglia of the auditory nerve (gang) and on the embryo’s right through the nerve root connecting the ganglion with the medulla oblongata. Various blood-vessels are cut through: their names and relations with one another are most easily determined by sliding a straight-edge along the drawing of the embryo as a whole.




FIG. 238.—-Diagrammatic sagittal section through third-day Fowl embryo. '.l‘he notochord and dorsal aorta are omitted Ectoderm and endoderm are indicated by cotltinllous lines, mcsoderm (except endocardium) by dots.

a, position of anus, not yet perforate; ull.,al1antois; am, zumiiong at, atrium; a.e, amniotic edge; f.a,, serous mcinbrane ; fl g, foregut; l, lung rudiment; nws, mesencephalon ; pa.g, postnnal gut; pt, pituitary involution; rh, rhombencephaion; .s~pl, splanchnopleure of yolk-sac; t, thalamencephalon; th, thyroid; V, ventricle; v.A, ventral aorta; 'v.m., remains of velar membrane; ,1/.s, ‘cavity of yolk. sac; 3:/..s-t, cavity of yolk-stalk.




The study of this stage should be completed by examining series of sections parallel to the sagittal plane in the ‘head region and interpreting them by what has been made out from the whole embryo and the series of transverse sections. The most instructive sections are those in or close to the sagittal plane. Fig. 238 shows diagrammatically a sagittal section through the whole length of the embryo, but it will of course be understood that, owing to the head of the embryo having come to lie over on its left side while the trunk region retains its original position, a section which is sagittal in the head region will, in actual fact, be practically horizontal in the trunk.


The feature that dominates the section is the cerebral flexure-— the strongly marked curvature of the head region towards the ventral side. The brain is of relatively enormous size: a distinct dip in its roof marks the boundary between the thin-roofed rhombencephalon which lies behind it and the region in front of it-—the cerebrum--whichwill give rise to mesencephalon, thalamencephalon and hemispheres.


The next instructive feature brought out by such a section is the general relation of gut to yolk-sac. The rounded head-fold of the splanchnépleure has extended far back so as to floor in the foregut (jig). The velar membrane (am) has just ruptured so that the foregut communicates in front with what will become the stomodaeum into which also opens the pituitary involution of the ectoderm (pt). The floor of the foregut dips downwards to form the rudiments of the thyroid (th) and lung (1). In a slightly more advanced embryo the two liver rudiments would be seen also as pocket-like outgrowths of the enteric floor in the neighbourhood of the atrial end of the cardiac tube.

The posterior end of the definitive alimentary canal is also becoming folded off from the yolk-sac though the cavity of the yolkstalk—-—the communication between the definitive alimentary canal and the cavity of the yolk-sac—-—is still very wide. The position of the future anal opening is indicated by a thick septum (Ct) composed of fused ectoderm and endoderm. Dorsal and posterior to this the enteron extends back as a blindly ending pocket—-the remains of the postanal gut (pay), while anterior to the anus the enteric floor dips downwards as the rudiment of the allantois (all). The latter is covered with a thick layer of mesoderm and bulges into a dilated portion of the splanchnocoele. Towards the front end of the embryo a still more widely dilated portion of the splanchnocoele accommodates the cardiac tube. At its anterior (o..A) and posterior ends (at) this, is ensheathed in the thick mesoderm on the ventral side of the foregut, while its middle portion (V) hangs free in the cavity.


Finally the amniotic fold of the soinatopleure is seen to extend almost completely over the body of the embryo, the amniotic edge (cue) bounding a comparatively small opening near the tail end.


Having studied in some detail the features- characteristic of an individual third-day embryo it will be convenient now to give a general sketch of the chief advances in development which take place during this day.


At the commencement of the day the body of the embryo lay flat along the surface of the yolk: only at its head end was it clearly demarcated from the surrounding blastoderm and this head region owing to the commencing ventral curvature was beginning to lean over on to its left side. During the course of the third day the tucking in of the blastoderm under the definitive body proceeds apace so that the body becomes more and more completely demarcated from the part of the blastoderm forming the yolk-sac wall, and the yolk~stalk becomes correspondingly narrowed. The preponderance of growth activity on the dorsal side which leads to the ventral curvature is during the early hours of the day especially marked in the region of the mesencephalon but as the day goes on becomes very pronounced about the level of the heart and still later in the tail region. Thus the axis of the body develops strong ventral curvature especially marked at three different levels——mesencephalic, cardiac and caudal. Along with this increasing curvature the whole body of the embryo comes to lie over on its left side so that the observer looking down upon the egg from above sees the body of the embryo in profile from its right side.


During the day the embryo becomes ensheathed in the amnion in the manner already described. The vitelline network of bloodvessels attains to its highest development, forming as it does the organ for respiration as well as for absorption of the food and its transport into the body of the embryo. Correlated with the lying of the embryonic body over on its left side the paired venous channels which convey the blood from the vitelline network into the heart gradually lose their symmetry, those of the right side dwindling in size while their fellows show a corresponding increase.


In the brain the main regions become established: the roof of the thalamencephalon and medulla oblongata. assume their thin membranous character while the hemispheres bulge out in front of the thalamencephalon. The central canal of the spinal cord becomes reduced to a vertical slit by the thickening of the side walls. The olfactory rudiment makes its appearance: the auditory rudiment becomes converted into the closed pear-shaped otocyst, still however connected with the ectoderm by a solid strand of cells. In the eye the lens thickening has become involuted and converted into a closed vesicle with its inner wall markedly thickened. The optic cup has been completely formed and the retinal layer differentiated from the thin and degenerate pigment layer. In the latter the first deposition of pigment takes place during the later hours of the day.


The definitive alimentary canal is still open towards the yolk-sac over about half its extent but in addition to the foregut there becomes folded off during the course of the third day a considerable extent of hind-gut, the ventral wall of which commences to bulge out to form the rudiment of the allantois towards the close of the day. The hind-gut is still closed posteriorly but the foregut late in the third or during the fourth day becomes thrown into communication with the stomodaeum by rupture of the velar membrane. The pituitary rudiment makes its appearance. The four gill-pouches are formed and reach the ectoderm, the fourth in the closing hours of the day, and the first or it may be the first two become perforate. The thyroid rudiment makes its appearance and during the latter half of the day becomes closed. The pulmonary rudiment develops and becomes constricted off from the pharynx except at its front end. About the beginning of the day the two liver rudiments appear and during its course the process of anastomosis begins between the branches which sprout out from them. During the latter half of the day the pancreatic rudiments make their appearance——first the dorsal, then the left ventral, then the right ventral.


During the course of the day the mesoderm segments increase from about 20 to 25 up to about 40. Early in the day the Wolfiian duct becomes tubular and in the latter half of the day it completes its backward growth and reaches the cloaca. The germinal epithelium becomes recognizable.


The skeleton remains throughout the day purely notochordal.

The heart retains its S—shape and during the latter half of the day the atrial septum begins to develop. The two dorsal acrtae begin about the commencement of the third day to undergo their fusion to form the definitive unpaired aorta. In addition to the first one or two aortic arches which are already present the third makes its appearance (Fig. 241, A, III, p. 550), then the fourth, and during the latter half of the day the sixth, while the first becomes obliterated. As regards the venous system the most important feature is the assumption of the same general plan of the main trunks as is characteristic of Fishes.

Finally it should be noted that during this day the body of the embryo-becomes enclosed within the amnion.

It will be realized even from the bare summary that has been given that the third day of incubation of the Fowl’s egg'is morphologically the most important of all and the student will be well advised to devote a good deal of time to making a detailed study of embryos of this period. _

The Fourth Day of Incubation

By the end of the fourth day of incubation the blastoderm has spread about half-way round the yolk. The vessels of the vascular area are conspicuous, though it is to be noticed that the terminal sinus is becoming relatively less so than it was during the third day. The folding off of the body of the embryo has progressed greatly. By the extension backwards of the head fold the region of the heart has become floored in on its ventral side. Posteriorly the tail fold is deepening in a similar fashion. Between head fold and tail fold the somatopleure of the embryonic body is prolonged ventralwards into a very short and wide tube--the somatic stalk-——the wall of which is reflected dorsalwards as the true amnion. The latter is now complete and closely invests the body of the embryo. Lying loosely within the somatic stalk and of much smaller diameter is the splanchnic or yolk stalk--the continuation of the splanchnopleure in a ventral direction as it passes out into the wall of the yolk-sac. The body of the embryo has undergone a great increase in size. The growth of its tissues has been particularly active in its dorsal region and this has led to a continuation of the flexure towards the ventral side which was already well marked in the third day embryo.


An important new feature in the fourth day embryo is provided b y the two pairs of limb rudiments each in the form of a dorsiventrally flattened ridge with rounded edge and broad base of attachment to the body. The head of the embryo at once attracts attention by its relatively enormous size. This is due to the relatively immense size of the brain and eyes. We have here to do apparently with a case of the precocious growth in size of organs which in the fully developed condition possess extreme complexity of minute structure. The main regions of the brain can be seen very distinctly: the relatively large mesencephalon with its bulging dome-like roof, the thalamencephalon with the pineal rudiment, the rapidly growing rudiments of the hemispheres, and the hind-brain with its relatively thin and membranous roof. The three main special sense organs are all conspicuous—-the olfactory organ, the eye with its choroid fissure and lens, the pyriform otocyst. Arranged in a row ventral to the otocysts are the pharyngeal clefts—three or four in number. In the case of cleft I the ventral part of the cleft is becoming much narrowed by the approach of its anterior and posterior walls. The dorsal end of the cleft on the other hand remains dilated: it corresponds to the spiracle of fish-like forms.


The heart, which forms a large structure lying between the tip of the head and the region of the fore limbs, is still in the form of a coiled tube but the appearance of localized bulgings of its wall foreshadows its division into the various chambers characteristic of the adult. Thus the curve of the tube lying posteriorly and on the right is becoming dilated to form ‘the ventricle: the part morphologically in front of this leading towards the ventral aorta is slightly dilated to form the conus arteriosus, while the curve lying anteriorly and on the left side shows a slight bulging on each side foreshadowing the two auricles. Slight constrictions separate these various bulgings-—an atrioventricular constriction narrowing the cavity to form the auricular canal, and a less conspicuous one between ventricle and conus. '


The general arrangement of the peripheral vessels is intermediate between that of the third day (Fig. 241, A) and that of the fifth day (Fig. 241, B) and need not be described in detail. Aortic arches I and II undergo in turn a gradual process of obliteration while arches IV and VI make their appearance farther back if they have not already done so. It is also during this day that arch V makes its brief appearance.


The allantoic veins, which at first are merely veins of the body-wall, during the fourth day establish their connexion with the allantois, and in the course of the day the right vein disappears.

The allantois itself forms a conspicuous new feature for towards the end of the day it begins to project distinctly from the ventral side of the embryo about the level of the hind limb.

Owing to the increasing size and complexity of the embryo the elementary student will not as a rule prepare complete series of sections later than the third day. He will however find it profitable to have transverse sections through the developingsense organs, sagittal sections through the head, and transverse sections through the posterior trunk region.

From the study of sections the following advances in development during the fourth day may be made out.

In the brain the rudiment of the paraphysis makes its appearance Fm. 239.--Fowl's egg opened at the end and the pineal outgrowth begins to of the fifth day. The embryo enclosed sprout; out into diverticula, about; in its amnion is sunk down in the - 1 centre of the vascular area, the allan~


the end of the day‘ rlhe Olfactory

tois projecting upwards towards the» rudiment b3C0m3S Conneclied With serous membrane — a transparent mem-

the buccal gavjty by a, slight,

brane through which the embryo and ' allantois are seen. The increasing groove‘ The rudunents of lagena

fluidity of the yolk is shown by the and recess make their appear-.

outward bulging of the yolk-saewall ance as bulging of the ototytt watt The cavity of the

Iiowlies completelyunderneath the yolk 13113 b(.3C01T1€3 Obliterated by th9

so as ‘to be invisible in a view from grgwth of its inner wall; pigment, becomes conspicuous in the outer wall of the optic cup: the layer of nerve fibres in the retina becomes recognizable: mesenchyme begins to invade the cavity of the optic cup and about the end of the day also intrudes between the lens and the ectoderm.

The post-anal gut becomes reduced to a solid strand of cells and finally disintegrates. The yo1k—stalk becomes narrowed to a fine tubular channel. The gall-bladder begins to dilate towards the'close of the day :. the dorsal pancreas begins to develop outgrowths: and the rudiments of the caeca make their appearance.

The mesoderm segments increase in number to about 50. Early in the day, if it has not done so already, the Wolffian duct opens into the cloaca. The mesonephric glomeruli begin to appear and the tubules become elongated and coiled. In the posterior region of the mesonephros secondary tubules make their appearance while in the anterior region a process of degeneration becomes apparent. During the second half of the day the ureter begins to sprout out from the Wolffian duct and about the end of the day the rudiments of Milllerian ducts and of the metanephric units may become recognizable.

In the heart the atrial septum becomes completed about the end of the fourth day and the endothelial cushions begin to develop.

FIG. 240.—-—Chick extracted from the egg at about the middle of the fifth day of incubation.

all, allantois; C.H, cerebral hemisphere; Is‘, eye; Hy, opereulum; M, mandibular arch; pin, pineal rudiment faintly visible as slight elevation on root’ of thalzuuencephalon; Rh, thin roof of rhombencephalon; som, edge of somatopleure cut through where it becomes reflected back over the body of the embryo to form the amnion; Lu, roof of tnesencephalon (optic lobe); V, ventricle; 47.0,

visceral clefts Ill and IV ; y.s, yolk-sac.

FIFTH DAY.—--The progress in development during the course of the fifth day is illustrated by Figs. 239-241. The albumen has so shrunk in volume as to be no longer visible in a view of the opened egg from above: the yolk has become extremely fluid: the vascular area has increased considerably in size. The allantois is now a conspicuous object and the mesoderm covering its surface is beginning to develop blood-vessels. The ‘head of the embryo is, as before, of relatively very large size: the flexure in the region of the mesencephalon is still more pronounced. The operculum (Fig. 240, Hg) is conspicuous, growing back from the hyoid arch over the posterior visceral clefts. The limb rudiments now project freely though their form is that of simple flippers without any of the peculiarities of the leg or wing of the Bird. The body of the embryo is floored in onits ventral side completely but for the rounded opening (som) along whose lips the somatopleure is continued into the amnion and through which emerge the narrowing yolk-stalk and the stalk of the allantois. The study of the living embryo in situ shows the general plan of the blood system to be as is shown in Fig. 241, B. The heart still betrays its tubular origin though the chambers are clearly recognizable as dilatations. Three aortic arches (III, IV and VI) are distinctly visible and occasionally the fleeting vestige of the penultimate arch as in the specimen represented in the diagram. In front of the aortic arches the ventral aorta is seen extending forwards as the ventral carotid (ac) :. the pulmonary artery (p.a) passes back from the sixth arch. Dorsally the aortic root extends forwards into the head as the dorsal carotid artery (d.o). A little distance behind the liver the vitelline artery (ua) leaves the dorsal aorta and farther back the allantoic artery (cm) a branch of which, the iliac artery, passes to the hind limb.



FIG. 241.—Diagrarn showing the main parts of the vascular system as seen in a Fowl embryo during the third day (A) and the fifth day (B).

a..a, allantoic artery; a.c.w,-, anterior cardinal vein; at, atrium; (1.17, allantoic vein; d.C, duct of

Cuvier; d.c, dorsal carotid; il.a, iliac artery; p.a, pulmonary artery; p.c.v, posterior cardinal vein; p.v.c, posterior vena cava ; v.A, ventral aorta; 1:.a, vitelline artery ; v.c, ventral carotid ; v.z~, vitelline vein ; I-VI, aortic arches. '


In the venous system the duct of Cuvier is seen, continuous at its dorsal ‘end with the anterior and posterior cardinal veins. ' The

‘former (a..o.q2) branches through‘ the head: the latter (19.0.72) can be X FOWL-—FIFTH AND SIXTH DAYS 551

traced dimly back into the region of the kidney. The main blood-'

stream to the heart comes from the vitelline vein (ac) and is joined

within the substance of the liver by the blood from the left allantoic

vein (am) and the posteriorvena cava (p/ac).

Ignoring the vitelline and allantoic vessels which are clearly adaptations to the peculiar conditions of the developing embryo the main plan of the blood system is seen to be clearly the same as is characteristic of Fishes.

By cutting off the head after fixing and viewing it from below (Fig. 245, A) the modelling of the face can be studied. The frontenasal process ( f .72) is bounded on each side by the shallow oro-nasal groove connecting it with the buccal cavity. The ridge forming the outer boundary of the olfactory organ is demarcated from the maxillary process by a faint transverse groove passing outwards towards the eye-—the lachrymal groove. Posteriorly the stomodaeal opening is bounded by the mandibular ridge with a distinct break in the middle line between the two mandibular arches.

Of other developmental features of the fifth day we may note the following. The first indications of turbinals appear on the mesial wall of the olfactory organ, and of semicircular canals in the otocyst. The optic stalk becomes solid: the rudiments of the ocular muscles become recognizable. The pituitary body begins to form outgrowths. The rudiments of thymus and bursa fabricii make their appearance: the bronchi begin to develop branches. The formation of new mesonephric tubule rudiments comes to an end and the mesonephros begins to show signs of functional activity. The atrial septum develops secondary perforations. The fourth aortic arch on the left side, and the portions of aortic root immediately behind the third arch undergo reduction. The horizontal septum of the ventral aorta begins to extend back into the conus and the anterior portions of the posterior cardinal veins begin to undergo atrophy.

SIXTH DAY.-—During the sixth day of incubation the body of the embryo increases rapidly in size and in correlation with this it dips down into the very fluid yolk, pushing the splanchnopleure of the yolk-sac wall in front of it, so that it is almost hidden from view when the egg is first opened. The amnion is, now raised up from the body of the embryo by a marked accumulation of amniotic fluid (Fig. 242). The allantois has increased greatly in size and in the natural condition is flattened rnflushroomwise againetnthfi iIme1‘..Su1:fa0B

\.—__-—-on-n.n-—. -—a of the serous membrane. In the embryo excised as directed on p. 513

‘it will be seen that the somatopleure of the embryonic body is

completely closed in ventrally except for a small circular space round which it is reflected outwards in a funnel-like fashion and continued into the thin membranous amnion. Through the funnel-like opening a slender probe can be passed from the extra-embryonic coelomic space beneath the serous membrane into the portion of coelome enclosed within the body of the embryo which will become the definitive splanchnopleure or body-cavity. Through the opening i552 EMBRYOLOGY OF THE LOWER VERTEBRATES CH.

there pass out the stalks of the yolk-sac and the allantois (Fig. 246, B) each conspicuous owing to its large blood-vessels. The peripheral distribution of the vitelline and allantoic vessels shows a characteristic difference (Fig. 242)———the vitelline network (vascular area) terminating, in the now greatly reduced terminal sinus at a considerable distance from the distal pole of the yolk-sac while on the other hand the allan


mic networkis most richly developed on the distal side of the allantois (p. 474)

The body of the embryo now for the first time’ begins to show indications of bird - like form, and faint traces of digits and of feather-rudiments may become apparent about the end of the day.

In the eye the rudiment of the pecten, which first became recognizable during the fourth day, is now conspicuous as an ingrowth of . mesenchynie through the choroidal fissure, bounded on each face by the inflected lips

, _ of the fissure. FIG. 242.-Coiiinioii Fowl. View of contents of the

, . egg-shell extracted at the end of the sixth day of he tongue beglns 1,30 _ incubation. The serous meinbraiie has been removed Pr0.]eCt and the thyrold

- so as {to gllow the allantois to be f(iltfi1pl&1C(3(l1 slightllly becolneg constrictgd off in or( er 0 give a c carer view 0 e )O(y o t e embryo contained within its ainnion. from the pharynx‘ The oesophagus towards the a. ii, edge or vascular area; all), reimilns of albumin; all’, . outer wall of allaiitols; all”, inner wall of allantois; am, end of the day loses lbs amnion; *, portion of vascular area lying, in the natural cavity; the dilatation of

ppiiaigilcgn. beneath the head of the eiiibryo and free from blood- the gizzard becomes evi_

. dent; the intestine begins to grow actively in length (Fig. 246, B). The three pancreatic rudiments become continuous with one another. ,

The muscles of the body begin to exhibit contractility, the trunk occasionally showing twitches of ventral flexure. The ureter develops outgrowths to form the primary collecting tubes of the metanephros about the beginning.of the sixth or the end of the fifth day and the terminal part of the duct of the opisthonephros may become incorporated in the cloaca so. as to give the ureter its independent opening. About this time the first indications of sexual differentiation become recognizable, the genital strands beginning to show signs of degeiiera-T tion in the female.


The main portions of the skeleton become laid down in prochondral tissue and, towards the end of the day, in cartilage.

The heart begins to assume its definitive external form; the ventricular septum develops and the conus septum begins to do so. The fourth aortic arch becomes obliterated on the left side.

SEVENTH DAY (Figs. 243 and 244).-——The mushroolm -shaped allantois is spreadin r activel all round beneath t 1e serous membrane. The amnijon is begilining to show waves of contraction passing along its wall. The brain and eyes and consequently the head as a whole are of relatively enormous size. In sections the roof of the fourth ventricle is found to be developing irregular folds in which the vessels of the choroid plexus will appear. All three turbinal rudiments are present in the nose. The crop is beginning to expand. The visceral clefts are all closed. The glands of the stomach are beginning to make their appearance as rudiments. The cavity of the enteron disappears for some distance forwards from the point of Omugln the a'11antmS' The Flu. 243.——Fowl's egg opmuul during the seventh day. Mllllerlan ducts 1113)’ Show The body of the chick is semi dimly through the

incipient asymmetry, The, highly vascular allantois. 'l‘ho \(.'.\‘.\'(.‘lf\‘ of the

. ° ° ' allantois can he tlistitlgllisllc-<l lmm lhn.~'(* of the notochorq 18 beglnnlng to vascular area by their turning back at the edge of be consbrlcted by the Var 179'‘ the allantois while those of the vascular area pass

brace, The first traces of onwards uninterruptedly. The highly fluid charQssification are Inaking their {‘l.Ct(‘.I: of the yolk‘ is .>‘hm\'l1 the _\'w«.»ll<-sac. wall

. . iulgmg outwards oi oi the broken shell at. the appearance, especially in the point m,.,k.,.1 «_ skeleton of the limbs.


The septum of the conus arteriosus is complete and the muscular coat extends into it from each side: the pocket-valves are becoming excavated. The fourth aortic arch on the left side has disappeared while the portion of aortic root between arches III and IV on the right side, and behind arch III on the left side, are becoming ulvlitica-1':1.tf.ed.

EIGHTH DAY.— The inoveinents of the amnion now reach their highest degree of activity. The fronto-nasal process (Fig. 245, C) is growing out to form the. pointed beak while the lower jaw is taking a siniilar pointed form, the two mandilmlar arches being new con tinua-.<l into um-. :uml<her ventrall,\' \\'il;.hout a l>1'e.ak. 'l‘he rudiments‘

of l'eathers are beginning to inake themselves apparent. In the brain the cerebellum is becoming folded on itself so as to bulge outwards, The oro-nasal grooves are covered in to form the tubular communication between nose’ and mouth. The lachrymal groove is no longer visible: the lachrymal glands are developing as solid ingrowths of ectoderm. The pituitary body now forms a rounded mass of branched glandular‘ tubes lying between the trabeculae and communicating with the buccal cavity by a narrow tubul_ar duct opening immediately over the glottis. The air-sac rudiments make their appearance on the surface of the lung (Fig. 246, C, a.s).


The mesonephric tubules have been growing actively up till now: the metanephric units are making their appearance: theMiillerian duct reaches the cloaca if it has not already done so although no actual communication is established until about six months after hatching.


Ossification becomes conspicuous in the limb-bones and the investing bones of the head. The keel of the sternum forms an ossification distinct from the two lateral rudiments of the body of the sternum.

The terminal sinus of the vascular area has disappeared. The septum of the conus is now completely traversed by muscle so that both aortic and pulmonary cavi. ties are completely ensheathed by muscle. The splitting apart of the two vessels is inaugurated by the appearance of a longitudinal incision along the line of attachment of the septum.


FIG. 244.—-—Chick extracted from egg during seventh day showing operculum (op).

As regards the further progress of development the following approximate times maybe mentioned.

About the ninth day the oesophagus gradually becomes patent again. On the tenth day the arterial arches have practically assumed the definitive condition and the metapodial skeleton is ossified.


Up to about the eleventh day. the contractions of the amnion remain very active, but thereafter they gradually become more gentle until during the closing days of incubation they stop. The mesonephros also attains to its maximum activity and there commences the process of degeneration which will continue till the time of hatching: tubules have developed throughout the length -of the metanephros. ‘

By the twelfth day the duct of the pituitary body has become reduced to a solid cellular strand: the exact time at which this happens is very variable; it may be as early as the sixth or seventh day. The lachrymal duct, which originated as a _solid ingrowth of ectoderm along the line of the lachrymal groove, now-becomes tubular. About the twelfth or thirteenth day the cavity reappears over the greater part of the rectum except just at the hinder limit of the occluded portion immediately in front of the allantois. Here the cavity remains blocked till nearly the time of hatching.

Flo. 2515.-—-View of head of Fowl embryo as seen from below. (After l‘)uval, 1889.)

A, five days; B, six days; C, eight days. fin, fronto-nasal process; mac, maxillary process; olf, olfac-V tory opening; o.'n., oro-nasal groove ; sp,hyomandihu1ar cleft; V, ventricle; I, II, visceral arches.

About_the thirteenth day the cartilaginous skeleton is complete and the rudiments of claws begin to develop.

About \the fifteenth day the Eustachian valve develops in the heart.

By the sixteenth day the albumen has all gone -and the yolk-sac wall becomes completed ventrally.

About the nineteenth day the yolk-sac becomes enclosed within the body-walland the partition between mesenteron and proctodaeum breaks down so that the alimentary canal communicates with the exterior. a .

About the twentieth day the umbilicus closes. The violent struggles of the young bird cause its beak to penetrate the air-space: its lungs are filled with air: its further struggles cause its beak to break the shell and it emerges, leaving behind the broken shell lined with the cast-off allantois and serous membrane.

Correlated with the, process of hatching important changes take place in the circulation? the gap in the atrial septum (foramen ovale) becomes closed so thatthe blood arriving in the right auricle can only reach the left auricle by the circuitous route through the

Fro. 246.-—Dissections from the right side showing the general arrangement of the viscera of 9. Fowl embryo at the end of the fifth (A), sixth (B), and eighth (0) days of incuba,tion_ (After Dnval, 1889.)

a.s, abdominal air-sac; all, allantois; c.a; conus arteriosus; wee, caecum; gi, gizzard; li, liver; mp, mesonephros; rr.a, right auricle; r.l, right lung; V, ventricle; y.d, yolk-stalk; y.s, yolk-sac.

right ventricle and pulmonary circulation, and the allantoic vein, duct of Botallus, and ductus venosus in the liver become obliterated.

Literature

Duval. Atlas d'Embryologie. Paris, 1889.

Poster and Balfour. The Elements of Embryology. Second Edition, edited by A. Sedgwick and W. Heape. London, 1883.

Koibol und Abraham. Keibels Normentafeln zur Entwicklungsgeschichte der Wirbeltiere, II. Jena, 1900.

Lillie. The Development of the Chick. New York, 1908.

Marshall. Vertebrate Embryology. London, 1893.

Patterson. Biol. Bulletin, xiii, 1907.

Patterson. Journ. Morpli.. xxi, 1910.

The most complete account of the development of the Fowl is that by Lillie. It, and Duval’s Atlas if a copy can be obtained, for it is unfortunately out of print, should form part of the equipment of every embryological laboratory.



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- Currently only Draft Version of Text -

Textbook Chapters: 1 Formation of the Germ Layers | 2 Skin and Derivatives | 3 Alimentary Canal | 4 Coelomic Organs | 5 Skeleton | 6 Vascular | 7 Internal Body Features | 8 Adaptation to Environmental Conditions | 9 General Considerations | 10 Common Fowl | 11 Lower Vertebrates | Appendix

Reference

Kerr JG. Text-Book of Embryology II (1919) MacMillan and Co., London.



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