Book - Comparative Embryology of the Vertebrates 3-7

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Nelsen OE. Comparative embryology of the vertebrates (1953) Mcgraw-Hill Book Company, New York.

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Part III The Development of Primitive Embryonic Form

Part III - The Development of Primitive Embryonic Form: 6. Cleavage (Segmentation) and Blastulation | 7. The Chordate Blastula and Its Significance | 8. The Late Blastula in Relation to Certain Innate Physiological Conditions: Twinning | 9. Gastrulation | 10. Tubulation and Extension of the Major Organ-forming Areas: Development of Primitive Body Form | 11. Basic Features of Vertebrate Morphogenesis

A. Introduction

1. Blastulae without auxiliary tissue

2. Blastulae with auxiliary or trophoblast tissue

3. Comparison of the two main blastular types

B. History of the concept of specific, organ-forming areas

C. Theory of epigenesis and the germ-layer concept of development

D. Introduction of the words ectoderm, mesoderm, endoderm

E. Importance of the blastular stage in Haeckel’s theory of “The Biogenetic Law of Embryonic Recapitulation”

F. Importance of the blastular stage in embryonic development

G. Description of the various types of chordate blastulae with an outline of their organforming areas

1. Protochordate blastula

2. Amphibian blastula

3. Mature blastula in birds

4. Primary and secondary reptilian blastulae

5. Formation of the late mammalian blastocyst (blastula)

a. Prototherian mammal, Echidna

b. Metatherian mammal, Didelphys

c. Eutherian mammals

6. Blastulae of teleost and elasmobranch fishes

7. Blastulae of gymnophionan amphibia

A. Introduction

In the previous chapter it was observed thatftwo main types of blastulae are formed in the chordate group: ^

(1) those blastulae without accessory or trophoblast tissue, e.g., Amphioxus, frog, etc. and

(2) those possessing such auxiliary tissue, e.g., elasmobranch and teleost fishes, reptiles, birds, and mammals.

1. Blastulae Without Auxiliary Tissue

The blastulae which do not have the auxiliary tissues are rounded affairs composed of a layer. of blastomeres surrounding a blastocoelic cavity (figs. 140T; 143C). The layer of blastomeres forms the blastoderm. The latter may be one cell in thickness, as in Amphioxus (fig. MOT), or several cells in thickness, as in the frog (fig. M3C). This hollow type of blastula often is referred to as a coeloblastula or blastosphere. However, in the gymnophionan amphibia, the blastula departs from this vesicular condition and appears quite solid. The latter condition may be regarded as a stereoblastula, i.e., a solid blastula. A somewhat comparable condition is present in the bony ganoid fishes, Amia and Lepisosteus,

The main characteristic of the blastula which does not possess auxiliary tissue is that the entire blastula is composed of formative cells, i.e., all the cells enter directly into the formation of the embryo’s body.

2. Blastulae with Auxiliary or Trophoblast Tissue^

examination of those blastulae which possess auxiliary or trophoblast tissues shows a less simple condition than the round blastulae mentioned above. In the first plac^fi two types of cells are present, namely, formative cells which enter into the "exposition of the embryonic body and auxiliary cells concerned mainly with trophoblast, or nutritional, functions. In the second place, in the blastula which possesses auxiliary tissue, the latter often develops precociously, that is, in advance of the formative cells of the blastul^ As a result, the arrangement of the formative cells into a configuration comparable to that of those blastulae without trophoblast cells may be much retarded in certain instances. This condition is true particularly of the mammalian blastula (blastocyst).

Generally speak ingj(^the blastulae which possess auxiliary tissue consist in their earlier stages of a disc or a mass of formative cells at the peripheral margins of which are attached the non-formative, auxiliary cells (fig. 159, blastoderm-formative cells, periblast-non-formative; also figs. M5K, L; M7G, H). The blastocoelic space lies below this disc of cells. However, in mammals the auxiliary or nourishment-getting tissue tends to circumscribe the blastocoel, whereas the formative cells occupy a polar area (fig. MSG, H). Blastulae, composed of a disc-shaped mass of cells overlying a blastocoelic space, have been described in classical terms as discoblastulae.')

3. Comparison of the Two Main Blastular Types

If we compare these two types of blastulae in terms of structure, it is evident that a comparison is not logical unless the essential or formative cells and their arrangement are made the sole basis for the comparison, for only the formative cells are common to both types of blastulae. To make the foregoing statement more obvious, let us examine the essential structure of a typical coeloblastula, such as found in Amphioxus, as it is defined by the presentday embryologist.

The studies by Conklin, ’32 and ’33, demonstrated that the fertilized egg

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THE CHORDATE BLASTULA AND ITS SIGNIFICANCE

of Amphioxus possesses five major, presumptive, organ-forming areas (fig. 167A). These areas ultimately give origin to the ectodermal, mesodermal, entodermal, notochordal, and neural tissues. In the eight-cell stage of cleavage, the cytoplasmic substances concerned with these areas are distributed in such a way that the blastomeres have different substances and, consequently, differ qualitatively (fig. 167B). Specifically, the entoderm forms the ventral part of the four ventral blastomeres; the ectoderm forms the upper or dorsal portion of the four micromeres, while the mesodermal, notochordal, and neural substances lie in an intermediate zone between these two organ-forming areas, particularly so in the blastomeres shown at the left in figure 167B. In figure 167C and D is shown a later arrangement of the presumptive, organ-forming areas in the middle and late stages of blastular development. These figures represent sections of the blastulae. Consequently, the organ-forming areas are contained within cells which occupy definite regions of the blastula. In figure 167E-G are presented lateral, vegetal pole, and dorso-posterior pole views of the mature blastula (fig. 167D), representing the organ-forming areas as viewed from the outside of the blastula.

It is evident from this study by Conklin that the organization of the fertilized egg of Amphioxus passes gradually but directly through the cleavage stages into the organization of the mature blastula; also, that the latter, like the egg, is composed of five, major, presumptive, organ-forming areas. It is evident further that one of the important tasks of cleavage and blastulation is to develop and arrange these major, organ-forming areas into a particular pattern. (Note: Later the mesodermal area divides in two, forming a total of six, presumptive, organ-forming areas.)

If we analyze the arrangement of these presumptive, organ-forming areas, we see that the mature blastula is composed of a floor or hypoblast, made up of potential, entoderm-forming substance, and a roof of potential ectoderm with a zone of mesoderm and chordoneural cells which lie in the area between these two general regions. In fact, the mesodermal and chordoneural materials form the lower margins of the roof of the mature blastula (fig. 167D). Consequently, the mature blastula of Amphioxus may be pictured as a bilaminar affair composed essentially of a hypoblast or lower layer of presumptive entoderm, and an upper concave roof or epiblast containing presumptive ectoderm, neural plate, notochord, and mesodermal cells. It is to be observed further that the blastocoel is interposed between these two layers. This is the basic structure of a typical coeloblastula. Furthermore, this blastula is composed entirely of formative tissue made up of certain definite, potential, organforming areas which later enter into the formation of the body of the embryo; auxiliary or non-formative tissue has no part in its composition. All coeloblastulae conform to this general structure.

If we pass to the blastula of the early chick embryo, a striking similarity may be observed in reference to the presumptive, organ-forming areas (fig.

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173). An upper, epiblast layer is present, composed of presumptive ectodermal, neural, notochordal, and mesodermal cells, while a hypoblast layer of entodermal potency lies below. Between these two layers the blastocoelic space is located. However, in the chick blastoderm, in addition to the formative cells, a peripheral area of auxiliary or trophoblast (periblast) tissue is present.

B. History of the Concept of Specific, Organ-forming Areas

The idea that the mature egg or the early developing embryo possesses certain definite areas having different qualities, each of which contributes to the formation of a particular organic structure or of several structures, finds its roots in the writings of Karl Ernst von Baer, 1828-1837. Von Baer’s comparative thinking and comprehensive insight into embryology and its processes established the foundation for many of the results and conclusions that have been achieved in this field during the past one hundred years.

Some forty years later, in 1874, Wilhelm His in his book, Unsere Korperform, definitely put forth the organ-forming concept relative to the germ layers of the chick, stating that “the germ-disc contains the organ-germs spread out in a flat plate,” and he called this the principle of the organ-forming germregions (Wilson, ’25, p. 1041). Ray Lankester, in 1877, advanced views supporting an early segregation from the fertilized egg of “already formed and individualized” substances, as did C. O. Whitman (1878) in his classical work on the leech, Clepsine. In this work. Whitman concludes that there is definite evidence in favor of the preformation of organ-forming stuffs within the egg. Other workers in embryology, such as Rabl, Van Beneden, etc., began to formulate similar views (Wilson, ’25, pp. 1041-1042).

The ideology embodied within the statement of Ray Lankester referred to above was the incentive for considerable research in that branch of embryological investigation known as “cell lineage.” To quote more fully from Lankester’s statement in this connection, p. 410:

Though the substance of a cell may appear homogeneous under the most powerful microscope, excepting for the fine granular matter suspended in it, it is quite possible, indeed certain, that it may contain, already formed and individualized, various kinds of physiological molecules. The visible process of segregation is only the sequel of a differentiation already established, and not visible.

The studies on cell lineage in many invertebrate forms, such as that of Whitman (1878) on Clepsine, of Wilson (1892) on Nereis, of Boveri (1892) and zur Strassen (1896; fig. 163B) on Ascaris, or the work of Horstadius (’28, ’37; fig. 163A) on the sea urchin, serve to emphasize more forcefully the implications of this statement. In these studies the developmental prospective fates of the various early cleavage blastomeres were carefully observed and followed.

Much of the earlier work on cell lineage was devoted to invertebrate forms. One of the first students to study the matter in the phylum Chordata was

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E. G. Conklin who published in 1905 a classical contribution to chordate embryology relative to cell lineage in the ascidian, Styela (Cynthia) partita. This monumental work extended the principle of organ-forming, germinal areas to the chordate embryo. However, the significance of the latter observations, relative to the chordate phylum as a whole, was not fully appreciated until many years later when it was brought into prominence by the German investigator, W. Vogt (’25, ’29).

Vogt began a series of studies which involved the staining of different parts of the amphibian blastula with vital dyes and published his results in 1925 and 1929. The method employed by Vogt is as follows:

Various parts of the late amphibian blastula are stained with such vital dyes as Nile-blue sulfate, Bismarck brown, or neutral red (fig. 168A). These stains color the cells but do not kill them. When a certain area of the blastula is stained in this manner, its behavior during later stages of development can be observed by the following procedure: After staining a particular area, the embryo is observed at various later periods, and the history of the stained area is noted. When the embryo reaches a condition in which body form is fully established, it is killed, fixed in suitable fluids, embedded in paraffin, and sectioned. Or, the embryo may be dissected after fixation in a suitable fluid. The cellular area of the embryo containing the stain thus may be detected and correlated with its original position in the blastula (cf. fig. 168A, B). This procedure then is repeated for other areas of the blastula (fig. 168C-E). Vogt thus was able to mark definite areas of the late blastula, to follow their migration during gastrulation, and observe their later contribution to the formation of the embryonic body. Definite maps of the amphibian blastula in relation to the future history of the respective blastular areas were in this way established (fig. 169C).

This method has been used by other investigators in the study of similar phenomena in other amphibian blastulae and in the blastulae and gastrulae of other chordate embryos. Consequently, the principle of presumptive, organforming areas of the blastula has been established for all of the major chordate groups other than the mammals. The latter group presents special technical difficulties. However, due to the similarity of early mammalian development with the development of other Chordata, it is quite safe to conclude that they also possess similar, organ-forming areas in the late blastular and early gastrular stages.

The major, presumptive, organ-forming areas of the late chordate blastula are as follows (figs. 167, 169, 1-73, 174, 179, 180, 181):

(1) There is an ectodermal area which forms normally the epidermal layer of the skin;

(2) also, there is an ectodermal region which contributes to the formation of the neural tube and nervous system;

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(3) a notochordal area is present which later gives origin to the primitive axis;

(4) the future mesodermal tissue is represented by two areas, one on either side of the notochordal area. In Amphioxus, however, this mesodermal area is present as a single area, the ventral crescent, which divides during gastrulation into two areas;

(5) the entodermal area, which gives origin to the future lining tissue of the gut, occupies a position in the blastula either at or toward the vegetative pole;

(6) there is a possibility that another potential area, containing germinal plasm, may be present and integrated with the presumptive entoderm or mesoderm. This eventually may give origin to the primitive germ cells;

(7) the pre-chordal plate region is associated with the notochordal area in all chordates in which it has been identified and lies at the caudal margin of the latter. In gastrulation it maintains this association. The pre-chordal plate material is an area which gives origin to some of the head mesoderm and possibly also to a portion of the roof of the foregut. It acts potently in the organization of the head region. Accordingly, it may be regarded as a complex of entomesodermal cells, at least in lower vertebrates.

C. Theory of Epigenesis and the Germ-layer Concept of Development

As the three classical germ layers take their origin from the blastular state (see Chap. 9), it is well to pause momentarily to survey briefly the germ-layer concept.

That the embryonic body is derived from definite tissue layers is an old concept in embryology. Casper Friedrich Wolff (1733-94) recognized that the early embryonic condition of the chick blastoderm possessed certain layers of tissue. This fact was set forth in his Theoria Generationis, published in 1759, and in De forniatione intestinorum praecipue, published in 1769, devoted to the description of the intestinal tract and other parts of the chick embryo. In these works Wolff presented the thesis that embryonic development of both plants and animals occurred by “a host of minute and always visible elements that assimilated food, grew and multiplied, and thus gradually in associated masses” produced the various structures which eventually become recognizable as “the heart, blood vessels, limbs, alimentary canal, kidneys, etc.” (The foregoing quotations are from Wheeler, 1898.) These statements contain the essence of Wolff’s theory of epigenesis. That is, that development is not a process of unfolding and growth in size of preformed structures; rather, it is an indirect one, in which certain elements increase in number and gradually become molded into the form of layers which later give rise to the organ structures of the organism.

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THE CHORDATE BLASTULA AND ITS SIGNIFICANCE

Two Other men contributed much to the layer theory of development, namely, Heinrich Christian Pander (1794-1865) and Karl Ernst von Baer (1792-1876). In 1817, Pander described the trilaminar or triploblast condition of the chick blastoderm, and von Baer, in his first volume (1828) and second volume (1837) on comparative embryology of animals, delineated four body layers. The four layers of von Baer’s scheme are derived from Pander’s three layers by dividing the middle layer into two separate layers of tissue. Von Baer is often referred to as the founder of comparative embryology for various reasons, one of which was that he recognized that the layer concept described by Pander held true for many types of embryos, vertebrate and invertebrate. The layer concept of development thus became an accepted embryological principle.

While Pander and von Baer, especially the latter, formulated the germlayer concept as a structural fact for vertebrate embryology, to Kowalewski (1846-1901) probably belongs the credit for setting forth the idea, in his paper devoted to the early development of Amphioxus (1867), that a primary, single-layered condition changes gradually into a double-layered condition. The concept of a single-layered condition transforming into a double-layered condition by an invaginative procedure soon became regarded as a fundamental embryological sequence of development.

Gradually a series of developmental steps eventually became crystallized from the fact and speculation present during the latter half of the nineteenth century as follows:

( 1 ) The blastula, typically a single-layered, hollow structure, becomes converted into

(2) the two-layered gastrula by a process of invagination of one wall or delamination of cells from one wall of the blastula; then,

(3) by an outpouching of a part of the inner layer of the gastrula, or by an ingression of cells from this layer, or from the outside ectoderm, a third layer of cells, the mesoderm, comes to lie between the entoderm and ectoderm; and finally,

(4) the inner layer of mesoderm eventually develops into a two-layered structure with a coelomic cavity between the layers.

This developmental progression became accepted as the basic procedure in the development of most Metazoa.

The original concept of the germ layers maintained that the layers were specific. That is, entodermal tissue came only from entoderm, ectodermal tissue from ectoderm, etc. However, experimental work on the early embryo in which cells are transplanted from one potential layer to another has overthrown this concept ( Oppenheimer, ’40). The work on cell lineage and the demonstration of the early presence of the presumptive, organ-forming areas

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also have done much to overthrow the concept concerning the rigid specificity of the three primary germ layers of entoderm, mesoderm, and ectoderm.

D. Introduction of the Words Ectoderm^ Mesoderm^ Endoderm

Various students of the Coelenterata, such as Huxley (1849), Haeckel (1866) and Kleinenberg (1872), early recognized that the coelenterate body was constructed of two layers, an outer and an inner layer. Soon the terms ectoderm (outside skin) and endoderm (inside skin) were applied to the outer and inner layers or membranes of the coelenterate body, and the word mesoderm (middle skin) was used to refer to the middle layer which appeared in those embryos having three body layers. The more dynamic embryological words epiblast, mesoblast, and hypoblast (entoblast) soon came to be used in England by Balfour, Lankester, and others for the words ectoderm, mesoderm, and endoderm, respectively. The word entoderm is used in this text in preference to endoderm.

E. Importance of the Blastular Stage in Haeckel’s Theory of ^^The Biogenetic Law of Embryonic Recapitulation”

In 1859, Charles Darwin (1809-82) published his work On the Origin of Species by Means of Natural Selection, This theory set the scientific world aflame with discussions for or against it.

In 1872 and 1874, E. Haeckel (1834-1919), an enthusiast of Darwin’s evolutionary concept, associated the findings of Kowalewski regarding the early, two-layered condition of invertebrate and vertebrate embryos together with the adult, two-layered structure of the Coelenterata and published the blastaea-gastraea theory and biogenetic principle of recapitulation. In these publications he applied the term gastrula to the two-layered condition of the embryo which Kowalewski has described as the next developmental step succeeding the blastula and put forward the idea that the gastrula was an embryonic form common to all metazoan animals.

In his reasoning (1874, translation, ’10, Chap. 8, Vol. I), Haeckel applied the word blastaea to a “long-extinct common stem form of substantially the same structure as the blastula.” This form, he concluded, resembled the “permanent blastospheres” of primitive multicellular animals, such as the colonial Protozoa. The body of the blastaea was a “simple hollow ball, filled with fluid or structureless jelly with a wall composed of a single stratum of homogeneous ciliated cells.”

The next phylogenetic stage, according to Haeckel, was the gastraea, a permanent, free-swimming form which resembled the embryonic, two-layered, gastrular stage described by Kowalewski. This was the simple stock form for all of the Metazoa above the Protozoa and other Protista. Moreover, he postulated that the gastrula represented an embryonic recapitulation of the adult stage of the gastraea or the progenitor of all Metazoa.

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THE CHORDATE BLASTULA AND ITS SIGNIFICANCE

The assumed importance of the blastula and gastrula thus became the foundation for Haeckel’s biogenetic principle of recapitulation. Starting with the postulation that the hypothetical blastaea and gastraea represented the adult phylogenetic stages comparable to the embryonic blastula and gastrula, respectively, Haeckel proceeded, step by step, to compress into the embryological stages of all higher forms the adult stages of the lower forms through which the higher forms supposedly passed in reaching their present state through evolutionary change. The two-chambered condition of the developing mammalian heart thus became a representation of the two-chambered, adult heart of the fish, while the three-chambered condition recapitulated the adult amphibian heart, etc. Again, the visceral arches of the embryonic pharyngeal regions of the mammal represented the gill-slit condition of the fish. Ontogeny thus recapitulates phytogeny y and phytogeny of a higher species is the result of the modification of the adult stages of lower species in the phylogenetic scale. The various steps in the embryological development of any particular species, according to this reasoning, were caused by the evolutionary history of the species; the conditions present in the adult stage of an earlier phylogenetic ancestor became at once the cause for its existence in the embryological development of all higher forms. Embryology in this way became chained to a repetition of phylogenetic links!

Many have been the supporters of the biogenetic law, and for a long time it was one of the most popular theories of biology. A surprising supporter of the recapitulation doctrine was Thomas Henry Huxley (1825-95). To quote from Oppenheimer (’40): “One wonders how the promulgator of such a distorted doctrine of cause and effect could have been championed by the same Huxley who wrote: Tact I know and Law I know; but what is this Necessity save an empty Shadow of my own mind’s throwing?’.”

The Haeckelian dogma that ontogeny recapitulates phytogeny fell into error because it was formulated upon three false premises due to the fragmentary knowledge of the period. These premises were:

( 1 ) That in evolution or phytogeny, recently acquired, hereditary characters were added to the hereditary characters already present in the species;

(2) that the hereditary traits revealed themselves during embryonic development in the same sequence in which they were acquired in phytogeny; and

(3) that Darwin’s concept of heredity, namely, pangenesis, essentially was correct.

The theory of pangenesis assumed that the germ cells with their hereditary factors were produced by the parental body or soma and that the contained hereditary factors within the germ cells were produced by gemmules which

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migrated from the various soma cells into the germ cells. This theory further postulated the inheritance of acquired characters.

If these three assumptions are granted, then it is easy to understand Haeckel’s contention that embryological development consists in the repetition of previous stages in phylogeny. For example, if we assume that the blastaea changed into the gastraea by the addition of the features pertaining to the primitive gut with its enteric lining, then the gastraea possessed the hereditary factors of the blastaea plus the new enteric factors. These enteric features could easily be added to the deric (outer-skin) factors of the blastaea, according to Darwin’s theory of pangenesis. Furthermore, according to assumption (2) above, in the embryonic development of the gastraea, the hereditary factors of the blastaea would reveal themselves during development first and would produce the blastaea form, to be followed by the appearance of the specific enteric features of the gastraea. And so it proceeded in the phylogeny and embryology of later forms. In this way the preceding stage in phylogeny became at once the cause of its appearance in the development of the next phylogenetic stage.

•These assumptions, relative to heredity and its mechanism of transference, were shown to be untenable by the birth of the Nageli-Roux-Weismann concept of the germ plasm (see Chaps. 3 and 5) and by the rebirth or rediscovery of Mendelism during the latter part of the nineteenth century. Studies in embryology since the days of Weismann have demonstrated in many animal species the essential correctness of Weismann’s assumption that the germ plasm produces the soma during development, as well as the future germ plasm, and thus have overthrown the pangenesis theory of Darwin. The assiduous study of Mendelian principles during the first twenty-five years of the twentieth century have demonstrated that a fixed relation does not exist between the original character and the appearance of a new character as implied in the Haeckelian law (Morgan, ’34, p. 148). Furthermore, that “in many cases, perhaps in most, a new end character simply replaces the original one. The embryo does not pass through the last stage of the original character and then develop the new one — although this may happen at times — but the new character takes the place of the original one” (Morgan, ’34, p. 148).

How then does one explain the resemblances of structure to be found among the embryos at various stages of development in a large group of animals such as the Chordata? Let us endeavor to seek an explanation.

In development, nature always proceeds from the general to the specific, both in embryological development and in the development of phylogeny or a variety of forms. The hereditary factors which determine these generalized states or structural conditions apparently are retained, and specialized factors come into play after the generalized pattern is established. Generalized or basic conditions, therefore, appear before the specialized ones. An example of this generalized type of development is shown in the formation of the

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blastula in chordate animals. Although many different specific types and shapes of blastulae are present in the group as a whole, all of them can be resolved into two basic groups. These groups, as mentioned in the beginning of this chapter, are:

( 1 ) blastulae without auxiliary, nutritive tissue and

(2) blastulae with auxiliary tissue.

Moreover, if the auxiliary tissue of those blastulae which possess this tissue is not considered, all mature chordate blastulae can be reduced to a fundamental condition which contains two basic layers, namely, hypoblast and epiblast layers. The epiblast possesses presumptive epidermal, neural, notochordal, and mesodermal, organ-forming areas, while the hypoblast cells form the presumptive entodermal area. The shapes and sizes of these blastulae will, of course, vary greatly. Moreover, the hypoblast cells may be present in various positions, such as a mass of cells at the caudal end of a disc-shaped epiblast (teleost and elasmobranch fishes), an enlarged, thickened area or pole of a hollow sphere (many Amphibia) y a single, relatively thin layer of cells, forming part of the wall of a hollow sphere (Arnphioxus), a rounded, disc-shaped mass of cells overlain by the thin, cup-shaped epiblast (Clavelina), a thickened mass attached to the underside of the caudal end of the disc-shaped epiblast (chick; certain reptiles), a thin layer of cells situated below the epiblast layer (mammals), or a solid mass of cells, lying below a covering of epiblast cells (gymnophionan Amphibia). Although many different morphological shapes are to be found in the blastulae of the chordate group, the essential, presumptive, organ-forming areas always are present, and all are organized around the presumptive notochordal area.

But the question arises: Why is a generalized blastular pattern developed instead of a series of separate, distinct patterns? For instance, why should the notochordal area appear to occupy the center of the presumptive, organforming areas of all the chordate blastulae when this area persists as a prominent morphological entity only in the adult condition of lower chordates? The answer appears to be this: The notochordal area at this particular stage of development is not alone a morphological area, but it is also a physiological instrument, an instrument which plays a part in a method or procedure of development. The point of importance, therefore, in the late blastular stage of development is not that the notochordal area is going to contribute to the skeletal axis in the adult of the shark, but rather that it forms an integral part of the biolgical mechanism which organizes the chordate embryo during the period immediately following the blastular stage. Thus, if the notochordal material can play an important role in the organization of the embryo and in the induction of the neural tube in the fish or in the frog, it also can fulfill a similar function in the developing chick or human embryo. Whatever it does later in development depends upon the requirements of the species. To use

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a naive analogy, nature does not build ten tracks to send ten trains with different destinies out of a station when she can use one track for all for at least part of the way. So it is in development. A simple tubular heart appears in all vertebrate embryos, followed by a simple, two-chambered* condition, not because the two-chambered heart represents the recapitulated, two-chambered, fish heart but rather because it, like the notochord, is a stage in a dynamic developmental procedure of heart development in all vertebrates. As far as the fish is concerned, when the common, two-chambered, rudimentary stage of the heart is reached, nature shunts it off on a special track which develops this simple, two-chambered condition into the highly muscular and efficient two-chambered, adult heart adapted to the fish level of existence in its watery environment. The three-chambered,* amphibian heart follows a similar pattern, and it specializes at the three-chambered level because it fits into the amphibian way of life. So it is with the embryonic pharyngeal area with its visceral and aortal arches which resemble one another throughout the vertebrate group during early embryonic development. The elaboration of a common, pharyngeal area with striking resemblances throughout the vertebrate group can be explained more easily and rationally on the assumption that it represents a common, physiologically important step in a developmental procedure.

This general view suggests the conclusion that ontogeny tends to use common developmental methods wherever and whenever these methods can be utilized in the development of a large group of animals. Development or ontogeny, therefore, recapitulates phylogenetic procedures and not adult morphological stages. One explanation for this conservation of effort may be that, physiologically speaking, the number of essential methods, whereby a specific end may be produced, probably is limited. Another explanation suggests that an efficient method never is discarded.

F. Importance of the Blastular Stage in Embryonic Development

Superficially in many forms, chordate and non-chordate, the blastula is a hollow, rounded structure containing the blastocoelic space within. It is tempting to visualize this form as the basic, essential form of the blastula. However, the so-called blastular stage in reality presents many forms throughout the animal kingdom, some solid, some round and hollow, and others in the form of a flattened disc or even an elongated band. Regardless of their shape, all blastulae have this in common/ they represent an association of presumptive organ-forming areas, areas which later move to new positions in the forming body, increase in cellular mass, and eventually become molded into definite structures. One of the main purposes of blastulation, therefore, may be stated as the elaboration (or establishment) of the major, presumptive organ-forming areas of the particular species and their arrangement in a particular pattern which permits their ready manipulation during the next

Exclusive of the sinus venosus.

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THE CHORDATE BLASTULA AND ITS SIGNIFICANCE

step of development or gastrulation. jThc particular shape of the blastula has its importance. However, this importance does not lie in the supposition that it conforms to a primitive spherical type but rather that the various, presumptive, organ-forming areas are so arranged and so poised that the cell movements so necessary to the next phase of development or gastrulation may be properly executed for the particular species. In most species, the formation of a blastocoelic space also is a necessary function of blastulation. In some species, however, this space actually is not formed until the next stage of development or gastrulation is in progress.

In summary, therefore, it may be stated that the importance of the blastula does not reside in the supposed fact that it is a one-layered structure or blastoderm having a particular shape. Rather, its importance emerges from the fact that the blastoderm has certain, well-defined areas segregated within it — areas which will give origin to future organ structures. Moreover, these areas foreshadow the future germ layers of the body. In diploblastic Metazoa, two germ layers are foreshadowed, while in triploblastic forms, three germ layers are outlined. As far as the Chordata are concerned, the hypoblast is the forerunner of the entoderm or the internal germ layer; whereas the epiblast is composed potentially of two germ layers, namely, the epidermal, neural plate areas which form the ectodermal layer and the chordamesodermal or marginal zone cells which give origin to the middle germ layer.

In the following pages, the chordate blastula is described as a two-layered structure composed of various, potential, organ-forming areas. This twolayered configuration, composed of a lower hypoblast and an upper epiblast, is used to describe the chordate blastula for the dual purpose of comparison and analysis of the essential structure of the various blastulae. The bilaminar picture, it is believed, will enable the student to understand better the changes which the embryo experiences during the gastrulative period.

G. Description of the Various Types of Chordate Blastulae with an Outline of Their Organ-forming Areas

1. Protochordate Blastula

The following description pertains particularly to Amphioxus. With slight modification it may be applied to other protochordates, such as Clavelina, Ascidiella, Styela, etc.

As noted in the introduction to this chapter, the potential entodermal cells of Amphioxus lie at the vegetal pole and form most of the floor or hypoblast of the blastula (fig. 167D). The upper or animal pole cells form a roof of presumptive epidermal, notochordal, mesodermal, and neural cells arched above and around the entoderm. The latter complex of organ-forming cells forms the epiblast. The blastocoelic cavity is large and insinuated between the hypoblast and epiblast. The presumptive notochordal and mesodermal

Fig. 167. Presumptive organ-forming areas in the uncleaved egg and during cleavage and blastulation in Amphioxus. (Original diagram based upon data obtained from Conklin, ’32, ’33.) (A) Uncleaved egg. (B) Eight-cell stage. (C) Early blastula in

section. (D) Late blastula in section. (E) Late blastula, external view from side. (F) Late blastula, external, vegetal pole view. (G) Late blastula, external, dorsoposterior view. The localization of cytoplasmic materials in Styela partita is similar to that of Amphioxus. Observe that the pointed end of the arrow defines the future cephalic end of the embryo. The position of the polar body denotes the antero-ventral area, while the position of the notochordal and neural plate material represents the antero-dorsal region. The “tail end’’ of the arrow is the postero-ventral area of the embryo.

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354 THE CHORDATE BLASTULA AND ITS SIGNIFICANCE

areas lie at the margins of the entodermal layer and surround it. As such, some of the cells of these two, organ-forming areas may form part of the floor of the bias tula. The presumptive, notochordal and neural plate cells lie at the future dorsal lip of the blastopore and form the dorsal crescent, while the mesodermal area occupies the ventral-lip region as the ventral crescent (fig. 167F). In Amphioxus, the mature blastula is pear shaped, with the body

Fig. 168. Ultimate destiny within the developing body of presumptive organ-forming areas of the late amphibian blastula, stained by means of vital dyes. (After Pastecls: J. Exper. Zool., 89.) (A) Area of blastula, stained. (B) Destiny of cellular area, stained

in (A). (D, E) Ultimate destiny shown by broken lines of cellular areas, stained in

late blastula shown in (C). (E) Anterior trunk segment. (D) Posterior trunk segment.

Fig. 169. Presumptive organ-forming areas in the amphibian late blastula and beginning gastrula. (A, B) General epiblast and hypoblast areas of the early and late blastular conditions, respectively. The hypoblast is composed mainly of entodermal or gut-lining structures, whereas the epiblast is a composite of ectodermal (i.e., epidermal and neural), mesodermal, and notochordal presumptive areas. Observe that the epiblast gradually grows downward over the hypoblast as the late blastula is formed. (C) Beginning gastrula of the urodele, Triton. (Presumptive areas shown according to Vogt, ’29.) (D) Same as above, from vegetative pole. (Slightly modified from Vogt, ’29.)

(E) Lateral view of beginning gastrula of anuran amphibia. (F) Dorsal view of the same. (E, F derived from description by Vogt, ’29, relative to Rami fusca and Bombinator; also Pasteels: J. Exper. Zool., 89, relative to Discogkmus.) Observe that an antero-posterior progression of somites is indicated in C and D.

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THE CHORDATE BLASTULA AND ITS SIGNIFICANCE

of the mesodermal crescent comprising much of the neck portion of the “pear” (fig. 167E).

The blastula of Amphioxus thus may be regarded essentially as a bilaminar structure (i.e., two-layered structure) in which the hypoblast forms the lower layer while the epiblast forms the upper composite layer.

.^ 1 , Amphibian Blastula

In the amphibian type of blastula, a spherical condition exists similar to that in Amphioxus (fig. 169). The future entoderm is located at the vegetative (vegetal) pole, smaller in amount in the frog, Rana pipiens, and larger in such forms as Necturus maculosus (fig. 169 A, B). The presumptive notochordal material occupies an area just anterior to and above the future dorsal lip of the blastopore. The dorsal lip of the gastrula, when it develops, arises within the entodermal area (fig. 169C-F). Extending laterally on either side of the presumptive notochordal region is an area of presumptive mesoderm (fig. 169C-F). Each of these two mesodermal areas tapers to a smaller dimension as it extends outward from the notochordal region. The presumptive notochordal and mesodermal areas thus form a composite area or circular marginal zone which surrounds the upper rim of the entodermal material.

Above the chordamesodermal zone are two areas. The presumptive neural area is a crescent-hke region lying above or anterior to the presumptive notochord-mesoderm complex. Anterior to the neural crescent and occupying the remainder of the blastular surface, is the presumptive epidermal crescent (fig. 169C-F).

In the various kinds of blastulae of this group, the yolk-laden, vegetal pole cells actually form a mass which projects upward into the blastocoelic space (fig. 169A, B). The irregularly rounded, presumptive entodermal, organforming area, therefore, is encapsulated partially by the other potential germinal areas, particularly by the chordamesodermal zone (fig. 169B). In a sense, this is true also of the protochordate group (fig. 167D).

The amphibian type of blastula includes those of the petromyzontoid Cyclostomes, the ganoid fishes with the exception of bony ganoids, the dipnoan fishes, and the Amphibia with the exception of the Gymnophiona, where a kind of solid blastula is present.

It is to be observed that the amphibian and protochordate blastulae differ in several details. In the first place, there is a greater quantity of yolk material in the blastula of the Amphibia; hence the presumptive entodermal area or hypoblast projects considerably into and encroaches upon the blastocoel. Also, in Amphioxus, the presumptive notochordal area forms a distinct dorsal crescent apart from the presumptive mesodermal or ventral crescent (fig. 167F), whereas, in the Amphibia, the notochordal material is sandwiched in between the two wings of mesoderm, so that these two areas form one composite marginal zone crescent (fig. 169D, E).

TYPES OF CHORDATE BLASTULAE

357

As in Amphioxus, the amphibian blastula may be resolved into a twolayered structure composed of a presumptive entodermal or hypoblast layer and an upper, epiblast layer of presumptive epidermal, notochordal, mesodermal, and neural tissues. Each of these layers, unlike that of Amphioxus, is several cells in thickness.

^3. Mature Blastula in Birds

Development of the hen’s egg proceeds rapidly in the oviduct (fig. 157B-G), and at the time that the egg is laid, the blastodisc (blastula) presents the following cellular conditions:

( 1 ) a central, cellular blastoderm above the primary blastocoel and

(2) a more peripheral portion, associated with the yolk material forming the germ-wall tissue (fig. 156G).

The central blastoderm is free from the yolk substance and is known as the area pellucida, whereas the germ-wall area with its adhering yolk material forms the area opaca (fig. 170). Around its peripheral margin the area pellucida is somewhat thicker, particularly so in that region which will form the posterior end of the future embryo. In the latter area, the pellucid margin may consist of a layer of three or even four cells in thickness (fig. 172A). This thickened posterior portion of the early pellucid area forms the embryonic shield (fig. 170). Anterior to the embryonic shield, the pellucid area is one or two cells in thickness (figs. 171A; 172B).

Eventually the pellucid area becomes converted into a two-layered structure with an upper or overlying layer, the primitive ectoderm or epiblast and a lower underlying sheet of cells, the primitive entoderm or hypoblast (figs. 171 A; 172A). The space between these two layers forms the true or secondary blastocoel. The cavity below the hypoblast is the primitive archenteric space. At the caudal and lateral edges of the pellucid area, cells from the inner zone of the germ wall appear to contribute to both hypoblast and epiblast.

The two-layered condition of the avian blastula shown in figure 171 A may be regarded as a secondary or late blastula. At about the time that the secondary blastula is formed (or almost completely formed), the hen’s egg is laid, and further development depends upon proper incubational conditions outside the body of the hen. Shortly after the latter incubation period is initiated, the primitive streak begins to make its appearance in the midcaudal region of the blastoderm, as described in Chapter 9.

Much controversy has prevailed concerning the method of formation of the entoderm and the two-layered condition in the avian blastoderm. Greatest attention has been given to the origin of the entoderm in the eggs of the pigeon, hen, and duck. The second layer is formed in the pigeon’s egg as it passes down the oviduct, in the hen’s egg at about the time of laying, and in the duck’s egg during the first hours of the external incubation period. The

Fig. 171. Origin of the hypoblast (entoderm) in the avian blastoderm. (A) Median, antero-posterior section of chick blastoderm. Entoderm arises by delamination from upper or epiblast layer; possibly also by cells that grow anteriad from thickened posterior area. (Based upon data supplied by Peter, ’34, ’38, and Jacobson, ’38.) (B-D) For mation of the hypoblast (entoderm) from epiblast by a process of delamination in the duck embryo. (Based upon data supplied by Pasteels, ’45.)

unincubated chick blastoderm is about 3 mm. in diameter, that of the duck, about 2 to 3 mm.

The most recent observations, relative to the formation of the second or hypoblast layer, have been made upon the duck’s egg (Pasteels, ’45). In this egg, Pasteels found that, at about nine hours after incubation is initiated, a two-layered condition is definitely formed and that “the primary entoblast of the duck is the result of a progressive delamination of the segmenting blastodisc

TYPES OF CHORDATE BLASTULAE

359

separating the superficial cells from the deeper ones” (fig. 171B-D). He further suggests that “the bilaminar embryo of birds is to be homologized with the blastula of the Amphibia, the cleft separating the two layers being equivalent to the blastocoele” (p. 13). The formation of the hypoblast (primary entoderm ) by a process of delamination from the upper layer or epiblast agrees with the observations by Peter (’38) on the developing chick and pigeon blastoderm (fig. 172) and of Spratt (’46) on the chick. It also agrees with some of the oldest observations, concerning the matter of entoderm formation, going back to Ollacher in 1869, Kionka, 1894, and Assheton, 1896. Others, such as Duval (1884, 1888) in the chick, and Patterson (’09) in the pigeon, have ascribed the formation of the primary entoderm to a process of invagination and involution at the caudal margin of the blastoderm, while Jacobson (’38) came to the conclusion that the entoderm of the pellucid area arose in chick and sparrow embryos through a process of outgrowth of cells from the primitive plate and from an archenteric canal produced by an inward bending of the epiblast and primitive plate tissue. The latter author believed that the entoderm of the area opaca arose by delamination.

The hypoblast of the chick gives origin to most of the tissue which lines the future gut, and, therefore, may be regarded as the potential entodermal area. As in the amphibia and Amphioxus, the epiblast is composed of several, presumptive organ-forming areas (fig. 173A). (See Pasteels, ’36c; Spratt, ’42, ’46.) At the caudal part of the epiblast is an extensive region of presumptive mesoderm bisected by the midplane of the future embryonic axis. Just anterior to this region and in the midplane is the relatively small, presumptive notochordal area. Between the latter and the mesodermal area is located the presumptive prechordal plate of mesodermal cells. Immediately in front of the notochordal region lies the presumptive neural area in the form of a crescent with its crescentic arms extending in a lateral direc

Fig. 172. Delamination of hypoblast (entoderm) cells from upper or epiblast layer in the chick blastoderm. (A) Posterior end of blastoderm (cf. fig. 17 lA). (B) Anterior end of blastoderm.

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THE CHORDATE BLASTULA AND ITS SIGNIFICANCE

Fig. 173. Presumptive organ-forming areas in the chick blastoderm. (A) Slightly modified from Spratt, ’46. (B) Schematic section of early chick blastoderm passing

through antero-posterior median axis.

tion from the midline of the future embryonic axis. Anterior to the neural crescent is the presumptive epidermal crescent. Within the area opaca is found potential blood-vessel and blood-cell-forming tissue, as well as the extensive extra-embryonic-tissue materials.

The above description of the presumptive organ-forming areas pertains to the avian blastula just previous to the inward migrations of the notochordal, pre-chordal plate, and mesodermal areas; that is, just previous to the appearance of the primitive streak and the gastrulative process.

4. Primary and Secondary Reptilian Blastulae

The primary blastula of turtle, snake, and lizard embryos is akin in essential features to that of birds. It consists of a central blastoderm or area pellucida, overlying a primary blastocoelic cavity, and a more distally situated opaque blastoderm, together with an indefinite periblast syncytium. A localized region of the central blastoderm, situated along the midline of the future embryonic axis and eccentrically placed toward the caudal end, is known as the embryonic shield.

A specialized, posterior portion of the embryonic shield, in which the upper layer (epiblast) is not separated from the underlying cells (hypoblast), is known as the primitive plate (fig. 174A-D). (Consult also Will, 1892, for

Fig. 174. Formation of hypoblast (entoderm) layer in certain reptiles; major presumptive organ-forming areas of reptilian blastoderm. (A) Section through blastoderm of the turtle, Clemmys leprosa. This section passes through the primitive plate in the region where the entoderm cells are rapidly budded off (invaginated?) from the surface layer. It presumably passes through (E) in the area marked entoblast. It is difficult to determine whether the entoderm cells are actually invaginated, according to the view of Pasteels, or whether this area represents a region where cells are delaminated or budded off in a rapid fashion from the overlying cells. (B) Similar to (A), diagrammatized to show hypoblast cells in black. (C) Section through early blastoderm of the gecko, Platydactylus. Epiblast cells are shown above, primitive entoderm cells below. (D) A later stage showing primitive plate area with the appearance of a delamination or proliferation of entoderm (hypoblast) cells from the upper layer of cells. (E) Presumptive, organ-forming areas of the turtle, Clemmys leprosa, before gastrulation. (F) Presumptive, organ-forming areas of the epiblast of turtle and other reptiles if the hypoblast is budded off or separated from the underside of the epiblast without invagination. It is to be observed that B and D represent modifications by the author.

361

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THE CHORDATE BLASTULA AND ITS SIGNIFICANCE

accurate diagrams of the reptilian blastoderm.) Surrounding the primitive plate, the central blastoderm is thinner and is but one (occasionally two cells) cell in thickness (see margins of figs. 174A, C). As development proceeds, a layer of cells appears to be delaminated or proliferated off from the undersurface of the primitive plate area (fig. 174C, D). This delamination gives origin to a second layer of cells, the entoderm or hypoblast (Peter, ’34). Some of these entodermal cells may arise by delamination from more peripheral areas of the central blastoderm outside the primitive plate area. In the case of the turtle, Clemmys leprosa, Pasteels (’37a) believes that there is an actual invagination of entodermal cells (fig. 174A-B). More study is needed to substantiate this view.

Eventually, therefore, a secondary blastula arises which is composed of a floor of entodermal cells, the hypoblast, closely associated with the yolk, and an overlying layer or epiblast. The epiblast layer is formed of presumptive epidermal, mesodermal, neural, and notochordal, organ-forming areas. The essential arrangement of the presumptive organ-forming areas in the reptiles is very similar to that described for the secondary avian blastula. The space between the epiblast and hypoblast layers is the secondary blastocoelic space.

Fig. 175. Early blastoderms of the prototherian mammal, Echidna. (A) Early blastoderm showing central mass of cells: with peripherally placed vitellocytes. (B) Later blastoderm. Central cells are expanding and the blastoderm is thinning out. Smaller cells (in black) are migrating into surface layer. Vitellocytes have fused to form a peripheral syncytial tissue. (C) Later blastoderm composed of a single layer of cells of two kinds. The smaller cells in black represent potential entoderm cells. (D) Increase of hypoblast cells and their migration into the archenteric space below to form a second or hypoblast layer.

TYPES OF CHORDATE BLASTULAE

363

Fig. 176. Early development of blastoderm of the opossum. (Modified from Hartman,

16.) (A) Blastocyst wall composed of one layer of cells from which entoderm cells are migrating inward, (B-D) Later development of the formative portion of the blastoderm. Two layers of cells are present in the formative area, viz., an upper epiblast layer and a lower hypoblast. Trophoblast cells are shown at the margins of the epiblast and hypoblast layers.

Both hypoblast and epiblast are connected peripherally with the periblast tissue.

5. Formation of the Late Mammalian Blastocyst (Blastula) a. Prototherian Mammal, Echidna

In Echidna, according to Flynn and Hill (’39, ’42), a blastoderm somewhat comparable to that of reptiles and birds is produced. An early primary blastular condition is first established, consisting of a mass of central cells with specialized vitellocytes at its margin (fig. 175A). A little later, an extension of this blastoderm occurs, and a definite primary blastocoelic space is formed below the blastoderm (fig. 175B). During this transformation, small, deeper lying cells (shown in black, fig. 175B) move up to the surface and become associated with the thinning blastoderm which essentially becomes a single layer of cells (fig. 175C). The marginal vitellocytes in the meantime fuse to form a germ-wall syncytium. This state of development may be regarded as the fully developed primary blastula. A little later, this primary condition becomes converted into a two-layered, secondary blastula, as shown in figure 175D by the secondary multiplication and migration inward of the small cells to form a lower layer or hypoblast. The latter process may be

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THE CHORDATE BLASTULA AND ITS SIGNIFICANCE

regarded as a kind of polyinvagination. In this manner the secondary blastula is formed. It is composed of two layers of cells, the epiblast above and the hypoblast below with the secondary blastocoelic space insinuated between these two layers.

b. Metatherian Mammal, Didelphys

The opossum, Didelphys virginiana, possesses a hollow blastocyst akin to the eutherian variety. (See Hartman, T6, T9; McCrady, ’38.) As observed in the previous chapter, it is produced by a peculiar method. The early blastomeres do not adhere together to form a typical morula as in most other forms; rather, they move outward and adhere to the zona pellucida and come to line the inner aspect of this membrane. As cleavage continues, they eventually form a primary blastula with an enlarged blastocoel.

Following this primary phase of development, one pole of the blastocyst begins to show increased mitotic activity, and this polar area gradually thickens (fig. 176A). At this time certain cells detach themselves from the thickened polar area of the blastocyst and move inward into the blastocoel (fig. 176A, B) .

Fig. 177. Schematic drawings of early pig development. (A) Early developing blastocyst. (B) Later blastocyst, showing two kinds of cells in the inner cell mass. (C) Later blastocyst, showing disappearance of trophoblast cells overlying the inner cell mass. (D) Later blastocyst. Two layers of formative cells are present as indicated with trophoblast tissue attached at the margins.

TYPES OF CHORDATE BLASTULAE

365

Fig. 178. Schematic drawings of the developing blastocyst of the monkey. (After Hcuser and Streeter: Carnegie Inst,, Washington, Publ. 538. Contrib. to Embryol. No. 181.) (A, B) Early blastocysts showing formative and non-formative cells in the inner

cell mass. (C-E) Later arrangement of the formative cells into an upper epiblast and lower hypoblast layer.

These cells form the mother entoderm cells, and by mitotic activity they give origin to an entodermal layer which adheres to the underside of the thickened polar area (fig. 176B, C). The polar area then thins out to form the expansive condition shown in figure 176D. A bilaminar, disc-shaped area thus is formed in this immediate region of the blastocyst, and it represents the area occupied

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THE CHORDATE BLASTULA AND ITS SIGNIFICANCE

by the formative cells of the blastula. The edge of this disc of formative cells is attached to the trophoblast or auxiliary cells (fig. 176D). Only the formative cells give origin to the future embryonic body.

c. Eutherian Mammals

The eutherian mammals as a whole present a slightly different picture of blastocyst development from that described above for marsupial species. These differences may be outlined as follows:

( 1 ) During the earliest phases of blastocyst development in most eutherian mammals, a distinct, inner cell mass is elaborated at the formative or animal pole (fig. 177 A, B). This characteristic is marked in some species (pig, rabbit, man, and monkey) and weaker in others (mink and armadillo). It may be entirely absent in the early blastula of the Madagascan insectivore, Hemicentetes semispinosus; however, in the latter, a thickening corresponding to the inner cell mass later

Fig. 179. Presumptive organ-forming areas in the blastoderm of the shark embryo. (A) Median section of the blastoderm of Torpedo ocellata. Hypoblast cells are shown in black. Caudal portion of the blastoderm is shown at the right. Cf. (B). (This figure partly modified from Ziegler, ’02 — see Chap. 6 for complete reference.) (B) Map of the presumptive organ-forming areas of the blastoderm of the shark, Scyllium canicula.

TYPES OF CHORDATE BLASTULAE

367

E P I BLAST

N TO DERM OR PRIMARY ' YPOBLAST

NEURAL ECTOOE RM

NOTOCHORD

ENTO DE RM DORSAL BLASTOPORAL LIP

Fig. 180. Presumptive organ-forming areas of the teleost fish blastoderm. (A) Median section through the late blastoderm of Fundulus heteroclitus just previous to gastrulation. Somewhat schematized from the author’s sections. Presumptive entoderm or hypoblast is shown exposed to the surface at the caudal end of the blastoderm and, therefore, follows the conditions shown in (B). (B) Presumptive organ-forming areas

of the blastoderm of Fundulus heteroclitus. Arrows show the direction of cell movements during gastrulation. (Modified from diagram by Oppenheimer, ’36.)

appears. Within the inner cell mass, two types of cells are present, namely, formative and trophoblast (figs. 177B; 178A).

(2) Unlike that of the marsupial mammal, an overlying layer of trophoblast cells, covering the layer of formative cells, always is present (fig. 177B). In some cases (rabbit, pig, and cat) they degenerate (the cells of Rauber, fig. 177C), while in others (man, rat, and monkey) the overlying cells remain and increase in number (fig. 178A-E).

(3) The entodermal cells arise by a separation (delamination) of cells from the lower aspect of the inner cell mass (figs. 177C; 178A), with the exception of the armadillo where their origin is similar to that of marsupials. With these differences, the same essential goal arrived at in the marsupial mammals is achieved, namely, a bilaminar, formative area, the embryonic disc, composed of epiblast and hypoblast layers (figs. 177D; 178D, E), which ultimately gives origin to the embryonic body. A bilaminar, extra-embryonic, trophoblast area, consisting of extra-embryonic entoderm and ectoderm, also is formed (figs. 177D; 178D, E). The secondary blastocoel originates between the epiblast and hypoblast of the embryonic disc, while below the hypoblast layer is the archenteric space (fig. 178E).

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THE CHORDATE BLASTULA AND ITS SIGNIFICANCE

6. Blastulae of Teleost and Elasmobranch Fishes

In the teleost and elasmobranch fishes, the primary blastula is a flattened, disc-shaped structure constructed during its earlier stages of an upper blastoderm layer of cells, the formative or strictly embryonic tissue, and a peripheral and lower layer of trophoblast or periblast tissues; the latter is closely associated with the yolk substance (figs. 179A; 180A; 181 A). The primary blastocoelic space lies between the blastoderm and the periblast tissue.

That margin of the formative portion of the blastoderm which lies at the future caudal end of the embryo is thickened considerably, and presumptive entodermal material or primary hypoblast is associated with this area. Its relationship is variable, however. In some teleost fishes, such as the trout, the entodermal cells are not exposed to the surface at the caudal portion of the blastodisc (fig. 181 A; Pasteels, ’36a). In other teleosts, a considerable portion of the entodermal cells may lie at the surface along the caudal margin of the blastoderm (fig. 180A; Oppenheimer, ’36). In the elasmobranch fishes the disposition of the entodermal material is not clear. A portion undoubtedly lies exposed to the surface at the caudal margin of the disc (fig. 179 A, B; Vandebroek, ’36), but some entodermal cells lie in the deeper regions of the blastoderm (fig. 179A).

Turning now to a consideration of the other presumptive organ-forming areas of the fish blastoderm, we find that the presumptive pre-chordal plate material lies exposed on the surface in the median plane of the future embryo immediately in front of the entoderm and near the caudal edge of the blastoderm. (It is to be observed that, in comparison, the pre-chordal plate lies well forward within the area pellucida of the bird blastoderm.) This condition is found in the shark, Scyllium, in Fundulus, and in the trout, Salrno (figs. 179B; 180B). However, in the trout it lies a little more posteriorly at the caudal margin of the disc (fig. 181B). Anterior to the pre-chordal plate is the presumptive notochordal material, and anterior to the latter is a rather expansive region of presumptive neural cells. These three areas thus lie along the future median plane of the embryo, but they exhibit a considerable variation in size and in the extent of area covered in Scyllium, Fundulus, and Salrno (figs. 179, 180, 181).

Extending on either side of these presumptive organ-forming areas, is an indefinite region of potential mesoderm. In Salrno, presumptive mesodermal cells lie along the lateral and anterior portions of the blastoderm edge (fig. 18 IB). However, in Scyllium and in Fundulus, it is not as extensive (figs. 179B; 180B). In front of the presumptive neural organ-forming area is a circular region, the presumptive epidermal area.

In their development thus far the three blastulae described above represent a primary blastuiar condition, and the cavity between the blastodisc and the underlying trophoblast or periblast tissue forms a primary blastocoel. This condition presents certain resemblances to the early blastocyst in the higher

PERI BLA ST ^

ENTODERM OR PRIMARY . . . HYPOBLAST

— MESODERM

1 . •• ■>;.'.\ rr E P I OE R M AL

ECTODERM

-NEURAL ECTO DERM

-NOTO CHORD

— -—P RE-CHORDAL PLATE

DORSAL BLASTOPORAL LIP

Fig. 181. Presumptive organ-forming areas of the blastoderm of the trout, Salma irideus. (A) Schematized section through blastoderm just previous to gastrulation. Presumptive entoderm (hypoblast) shown in black at caudal end of the blastoderm. Observe that entoderm is not exposed to surface. Cf. (B). (B) Surface view of presumptive

organ-forming areas of the blastoderm just before gastrulation.

Fig. 182. Late blastoderms of Gymnophiona. (Modified from Brauer, 1897.) (A)

Late blastoderm of Hypogeophis alternans. Entoderm cells in black lie below. (B) Beginning gastrula of H. rostratus. Observe blastocoelic spaces in white between the entoderm cells.

369

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THE CHORDATE BLASTULA AND ITS SIGNIFICANCE

mammals and the late blastula of birds. In both groups the trophoblast tissue is attached to the edges of the formative tissue and extends below in such a way that the formative cells and trophoblast tissue tend to form a hollow vesicle. In both, the formative portion of the blastula is present as a disc or mass of cells composed of presumptive, organ-forming cells closely associated at its lateral margins with the trophoblast or food-getting tissue. A marked distinction between the two groups, however, is present in that the future entodermal cells in fishes are localized at the caudal margin of the disc, whereas in mammals and birds they may be extensively spread along the under margin of the disc. In reptiles the condition appears to be somewhat similar to that in birds and mammals, with the exception possibly of the turtles, where the future entoderm appears more localized and possibly may be superficially exposed. Therefore, while great differences in particular features exist between the fishes and the higher vertebrates, the essential fundamental conditions of the early blastulae in teleost and in elasmobranch fishes show striking resemblances to the early blastulae of reptiles, birds, and mammals.

The blastulae of teleost fishes remain in this generalized condition until about the time when the gastrulative processes begin. At that time the notochordal and mesodermal, cellular areas begin their migrations over the caudal edge of the blastodisc to the blastococlic space below, where they ultimately come to lie beneath the epidermal and neural areas. Associated with the migration of notochordal and mesodermal cells, an entodermal floor or secondary hypoblast is established below the notochordal and mesodermal cells by the active migration of primary hypoblast cells in an antero-lateral direction. In the elasmobranch fishes there is a similar cell movement from the caudal disc margin, as found in teleost fishes, but, in addition, a delamination of entodermal (and possibly mesodermal cells) occurs from the deeper lying parts of the blastodisc.

7. Blastulae of Gymnophionan Amphibia

In the Gymnophiona, nature has consummated a blastular condition different from that in other Amphibia. It represents an intermediate condition between the blastula of the frog and the blastodiscs of the teleost and elasmobranch fishes and of higher vertebrates (fig. 182). In harmony with the frog blastula, for example, a specialized periblast or food-getting group of cells is absent. On the other hand, the presumptive entoderm and the presumptive notochordal, mesodermal, neural, and epidermal cells form a compact mass at one pole of the egg, as in teleosts, the ohick, and mammal. Similar to the condition in the chick and mammal, the entodermal cells delaminate (see Chap. 9) from the under surface of the blastodisc (Brauer, 1897).

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