Book - Comparative Embryology of the Vertebrates 1-3
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Nelsen OE. Comparative embryology of the vertebrates (1953) Mcgraw-Hill Book Company, New York.
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Part I The Period of Preparation
Part I - The Period of Preparation: 1. The Testis and Its Relation to Reproduction | 2. The Vertebrate Ovary and Its Relation to Reproduction | 3. The Development of the Gametes or Sex Cells
The Development of the Gametes or Sex Cells
A. General Considerations
In the two preceding chapters the conditions which prepare the male and female parents for their reproductive responsibilities are considered. This chapter is devoted to changes which the male and female germ cells must experience to enable them to take part in the processes involved in the reproduction of a new individual.
The gamete is a highly specialized sex cell or protoplasmic entity so differentiated that it is capable of union (fertilization; syngamy) with a sex cell 6T the opposite sex to form the zygote from which the new individual arises. The process of differentiation whereby the primitive germ cell is converted into the mature gamete is called the maturation of the germ cell.
The main events which culminate in the fully-developed germ cell are possible only after the primitive or undifferentiated germ cell has reached a certain condition known as the definitive state. When this stage is reached, the germ cell has acquired the requisite qualities which make it possible for it to differentiate into a mature gamete. Before the definitive state is reached, germ cells pass through an eventful history which involves:
- their so-called "origin" or first detectable appearance among the othercells of the developing body, and
- their migration to the site of the future ovary or testis.
After entering the developing substance of the sex gland, the primitive germ cells experience a period of multiplication. If the sex gland is that of the male, these undifferentiated sex cells are called spermatogonia; if female, they are known as oogonia.
B. Controversy Regarding Germ-cell Origin
The problems of germ-cell origin in the individual organism and of the continuity of the germ plasm from one generation to the next have long been matters of controversy. Great interest in these problems was aroused by the ideas set forth by Waldeyer, Nussbaum, and Weismann during the latter part of the nineteenth century. Waldeyer, 1870, as a result of his studies on the chick, presented the “germinal epithelium†hypothesis, which maintains that the germ cells arise from the coelomic epithelium covering the gonad. Nussbaum, 1880, championed the concept of the extra-gonadal origin of the germ cells. According to this view, derived from his studies on frog and trout development, the germ cells arise at an early period of embryonic development outside the germ-gland area and migrate to the site and into the substance of the germ gland.
At about this time the speculative writings of August Weismann aroused
great interest. In 1885 and 1892 Weismann rejected the popular Darwinian
theory of pangenesis, which held that representative heredity particles or
“gemmules†passed from the body cells (i.e., soma cells) to the germ cells
and were there stored in the germ cells to develop in the next generation
(Weismann, 1893). In contrast to this hypothesis he emphasized a complete
independence of the germ plasm from the somatoplasm. He further suggested
that the soma did not produce the germ plasm as implied in the pangenesis
theory, but, on the contrary, the soma resulted from a differentiation of the
germ plasm.
According to the Weismannian view, the germ plasm is localized in the
chromosomal material of the nucleus. During development this germ plasm
is segregated qualitatively during successive cell divisions with the result that
the cells of different organs possess different determiners. However, the nuclear
germ plasm (Keimplasma) is not so dispersed or segregated in those cells
which are to become the primitive sex cells; they receive the full complement
of the hereditary determiners for the various cells and organs characteristic
of the species. Thus, it did not matter whether the germ cells were segregated
early in development or later, so long as the nucleus containing all of the
determinants for the species was kept intact. In this manner the germ plasm,
an immortal substance, passed from one generation to the next via the nuclear
germ plasm of the sex or germ cells. This continuity of the nuclear germ
plasm from the egg to the adult individual and from thence through the
germ cells to the fertilized egg of the next generation, constituted the Weismann
“Keimbahn†or germ-track theory. The soma or body of any particular generation is thus the “trustee†for the germ plasm of future generations.
The Weismannian idea, relative to the qualitative segregation of the chromatin materials, is not tenable for experimental and cytological evidence suggests that all cells of the body contain the same chromosomal materials. However, it should be pointed out that Weismann was one of the first to suggest that the chromosome complex of the nucleus acts as a repository for all of the hereditary characteristics of the species. This suggestion relative to the role of the nucleus has proved to be one of the main contributions to biological theory in modern times.
Fig. 60. Representation of the concept of the early embryonic origin of the primordial germ cells and their migration into the site of the developing germ gland. (A-C are adapted from the work of Allen, Anat. Anz. 29, on germ cell origin in Chrysemys; D-F are diagrams based on the works of Dustin, Swift, and Dantschakoff, etc., referred to in the table of germ-cell origins included in the text.) (A~C) Germ cells arising within the primitive entoderm and migrating through the dorsal mesentery to the site of the primitive gonad, shown in (D), where they become associated in or near the germinal epithelium overlying the internal mesenchyme of the gonad. (E, F) Increase of the primitive gonia within the developing germ gland, with a subsequent migration into the substance of the germ gland of many germ cells during the differentiation of sex.
Fig. 61 Diagrammatic representation of the process of chromatin diminution in the nematode worm, Ascaris cquorum (A. megalocephala), and of the “Keimbahn†(in black, E). One daughter cell shown by the four black dots of each division of the germcell line (i.e., the stem-cell line) is destined to undergo chromatin diminution up to the 16-cell stage. At the 16-cell stage, the germ-cell line ceases to be a stem cell (e.g., P 4 ), and in the future gives origin only to sperm cells (E). (A-D, copied from King and Beams (’38); E, greatly modified from Durkin (’32).)
Animal pole of the cleaving egg (A) is toward the top of the page. (B) Metaphase conditions of the second cleavage. Observe the differences in the cleavage planes of the prosomatic cell, Si, and that of the stem cell, P,. (C) Anaphase of the second cleavage of Si. Observe that the ends of the chromosomes in this cleaving cell are left behind on the spindle. (D) It is to be noted that the ends of the chromosomes are not included in the reforming nuclei of the two daughter cells of S,. thus effecting a diminution of the chromatin substance. In P„ P., and E.M. ST. of (D), the chromosomes are intact. E.M. ST. = second prosomatic cell. MST = mesoderm-stomodaeal cell.
A second contributory concept to the germ-cell (germ-plasm) theory was
made by Nussbaum, 1880; Boveri, 1892, ’10, a and b, and others. These
investigators emphasized the possibility that a germinal cytoplasm also is
important in establishing the germ plasm of the individual. A considerable
body of observational and experimental evidence derived from embryological
studies substantiates this suggestion. Consequently, the modern view of the
germ cell (germ plasm) embodies the concept that the germ cell is composed of the nucleus as a carrier of the hereditary substances or genes and a peculiar,
specialized, germinal cytoplasm. The character of the cytoplasm of the germ
cell is the main factor distinguishing a germ cell from other soma cells.
The matter of a germinal cytoplasm suggests the necessity for a segregation of the germinal plasm in the form of specific germ cells during the early development of the new individual. As a result, great interest, as well as controversy, has accumulated concerning this aspect of the germ-cell problem: namely, is there a separate germinal plasm set apart in the early embryo which later gives origin to the primordial germ cells, and the latter, after migration (fig. 60), to the definitive gonia; or according to an alternative view, do some or all of the definitive germ cells arise from differentiated or relatively undifferentiated soma cells? The phrase primary primordial germ cells often is used to refer to those germ cells which possibly segregate early in the embryo, and the term secondary primordial germ cells is employed occasionally to designate those which may arise later in development.
The dispute regarding an early origin or segregation of the germinal plasm
in the vertebrates also occurs relative to their origin in certain invertebrate
groups, particularly in the Coelenterata and the Annelida (Berrill and Liu,
’48). In other fnvertebrata, such as the dipterous insects and in the ascarid
worms, the case for an early segregation is beyond argument. An actual demonstration of the continuity of the Keimbahn from generation to generation
is found in Ascaris megalocephala described by Boveri in 1887. (See Hegner,
’14, Chap. 6.) In this form the chromatin of the somatic cells of the body
undergoes a diminution and fragmentation, whereas the stem cells, from which
the germ cells are ultimately segregated at the 16-cell to 32-cell stage, retain
the full complement of chromatin material (fig. 61). Thus, one cell of the
16-cell stage retains the intact chromosomes and becomes the progenitor of
the germ cells. The other 15 cells will develop the somatic tissues of the body.
The diminution of the chromatin material in this particular species has been
shown to be dependent upon a certain cytoplasmic substance (King and
Beams, ’38).
In some insects the Keimbahn also can be demonstrated from the earliest stages of embryonic development. In these forms a peculiar polar plasm within the egg containing the so-called “Keimbahn determinants’’ (Hegner, ’14, Chap. 5) always passes into the primordial germ cells. That is, the ultimate formation and segregation of the primordial germ cells are the result of nuclear migration into this polar plasm and the later formation of definite cells from this plasm (fig. 62). The cells containing this polar plasm are destined thus to be germ cells, for they later migrate into the site of the developing germ glands and give origin to the definitive germ cells.
Many investigators of the problem of germ-cell origin in the vertebrate group of animals have, after careful histological observation, described the germ cells as taking their origin from among the early entodermal cells (see table, pp. 121-124). On the other hand, other students have described the origin of the germ cells from mesodermal tissue — some during the early period of embryonic development, while others suggest that the primordial germ cells arise from peritoneal (mesodermal) tissue at a much later time.
In more recent years much discussion has been aroused relative to the origin of the definitive germ cells in mammals, particularly in the female. According to one view the definitive germ cells which differentiate into the mature gametes of the ovary arise from the germinal epithelium (peritoneal covering) of the ovary during each estrous cycle (figs. 39A, 63, 64). For example, Evans and Swezy (’31) reached the conclusion that all germ cells in the ovaries of the cat and dog between the various reproductive periods degenerate excepting those which take part in the ovulatory phenomena. Accordingly, the new germ cells for each cycle arise from the germinal epithelium. A similar belief of a periodic proliferation of new germ cells by the germinal epithelium has been espoused by various authors. (See Moore and Wang, ’47; and Pincus, ’36, Chap. II.) More recent papers have presented views which are somewhat conflicting. Vincent and Dornfeld, ’48 (fig. 63) concluded that there is a proliferation of germ cells from the germinal epithelium of the young rat ovary, while Jones (’49), using carbon granules as a vital-marking technic, found no evidence of the production of ova from the germinal epithelium in rat ovaries from 23 days until puberty. In the adult rat, she concedes that a segregation of a moderate number of oogonia from the germinal epithelium is possible.
Fio. 62. Early development of the fly, Miflj/o/’. {A) Miastor metraloas. (B) Miastor americana. In (A) the division figures I and III (II not shown) are undergoing chromatin diminution, while nucleus IV divides as usual. In (B) one segregated germ is shown at the pole of the egg. This cell will give origin to the germ cells. Other division figures experiencing chromatin diminution.
Fig. 63. Cells proliferating inward from germinal epithelium of the ovary of a oneday-old rat. Observe cords of cells (Pfliiger’s cords) projecting into the ovarian substance. Within these cords of cells are young oogonia. (After Vincent and Dornfeld, ’48.)
Fig. 64. Cellular condition near the surface of the ovary of a young female opossum. This section of the ovary is near the hilar regions, i.e., near the mesovarium. Observe young oocytes and forming Graafian follicles. Primitive germ cells may be seen near the germinal epithelium.
Aside from the above studies of carefully-made, histological preparations relative to the time and place of origin of the primordial and definitive germ cells, many experimental attacks have been made upon the problem. Using an x-ray-sterilization approach, Parkes (’27); Brambell, Parkes and Fielding (’27, a and b), found that the oogonia and oocytes of x-rayed ovaries of the mouse were destroyed. In these cases new germ cells were not produced from the germinal epithelium. Brambell (’30) believed that the destruction of the primitive oogonia was responsible for the lack of oogenesis in these x-rayed ovaries. However, this evidence is not conclusive, for one does not know what injurious effects the x-rays may produce upon the ability of the various cells of the germinal epithelium to differentiate.
An experimental study of the early, developing, amphibian embryo relative
to the origin of the primordial germ cells also has been made by various investigators. Bounoure (’39) applied a vital-staining technic to certain anuran
embryos. The results indicate that the germinal plasm in these forms is associated with the early, entodermal, organ-forming area located at the vegetal
pole of the cleaving egg. This germinal plasm later becomes segregated into
definite cells which are associated with the primitive entoderm. At a later
period these cells migrate into the developing germ gland or gonad. On the
other hand, experimental studies of the urodele embryo indicate that the
early germinal plasm is associated with the mesoderm (Humphrey, ’25, ’27;
Nieuwkoop, ’49). Existence of an early germinal plasm associated with the
entoderm in the Anura and with the mesoderm in the Urodela thus appears
to be well established for the amphibia.
The evidence derived from amphibian studies together with the observations upon the fish group presented in the table (see pp. 121-124) strongly
suggests that an early segregation of a germinal plasm (germ cells) occurs in
these two major vertebrate groups. Also, in birds, the experimental evidence
presented by Benoit (’30), Goldsmith ('35), and Willier ('37) weighs the
balance toward the conclusion that there is an early segregation of germ cells
from the entoderm. Similar conditions presumably are present in reptiles. In
many vertebrates, therefore, an early segregation of primordial germ cells and
their ultimate migration by: (1) active ameboid movement, (2) by the
shifting of tissues, or (3) through the blood stream (see table, pp. 121-124)
to the site of the developing gonad appears to be well substantiated.
The question relative to the origin of the definitive ova in the mammalian ovary is still in a confused state as indicated by the evidence presented above and in the table on pp. 121-124. Much more evidence is needed before one can rule out the probability that the primordial germ cells are the progenitors of the definitive germ cells in the mammals. To admit the early origin of primordial germ cells on the one hand, and to maintain that they later disappear to be replaced by a secondary origin of primitive germ cells from the germinal epithelium has little merit unless one can disprove the following position, to wit: that, while some of the primordial germ cells undoubtedly do degenerate, others divide into smaller cells which become sequestered within or immediately below the germinal epithelium of the ovary and within the germinal epithelium of the seminiferous tubules of the testis, where they give origin by division to other gonial cells. Ultimately some of these primitive gonia pass on to become definitive germ cells.
However, aside from the controversy whether or not the primordial germ cells give origin to definitive germ cells, another aspect of the germ-cell problem emphasizing the importance of the primitive germ cells is posed by the following question: Will the gonad develop into a functional structure without the presence of the primordial germ cells? Experiments performed by Humphrey (’27) on Ambystoma, and the above-mentioned workers — Benoit (’30), Goldsmith (’35), and Willier (’37) — on the chick, suggest that only sterile gonads develop without the presence of the primordial germ cells.
Finally, another facet of the germ-cell problem is this; Are germ cells completely self differentiating? That is, do they have the capacity to develop by themselves; or, are the germ cells dependent upon surrounding gonadal tissues for the influences which bring about their differentiation? All of the data on sex reversal in animals, normal and experimental (Witschi, in Allen, Danforth, and Doisy, ’39), and of other experiments on the development of the early embryonic sex glands (Nieuwkoop, ’49) suggest that the germ cells are not self differentiating but are dependent upon the surrounding tissues for the specific influences which cause their development. Furthermore, the data on sex reversal shows plainly that the specific chromosome complex (i.e., male or female) within the germ cell does not determine the differentiation into the male gamete or the female gamete, but rather, that the influences of the cortex (in the female) and the medulla (in the male) determine the specific type of gametogenesis.
The table given on pp. 121-124 summarizes the conclusions which some authors have reached concerning germ-cell origin in many vertebrates. It is not complete; for more extensive reviews of the subject see Everett ('45), Heys ('31 ), and Nieuwkoop ('47, '49).
Species
Place of Origin, etc.
A uthor
Entospheniis wihieri (brook lamprey)
Germ cells segregate early in the embryo; definitive germ cells derived from "no other source"
Okkelberg. 1921.
J. Morphol. 35
Petromyzon marinus unicolor (lake lamprey)
Definitive germ cells derive from: a) early segregated cells, primordial germ cells, and b) later from coelomic epithelium. Suggests that primordial germ cells may induce germ-cell formation in peritoneal epithelium
Butcher. 1929.
Biol. Bull. 56
Squalus acanthias
Germ cells segregate from primitive entoderm; migrate via the mesoderm into site of the developing gonad
Woods. 1902.
Am. J. Anat. 1
Amia and
Lepidosteiis
Germ cells segregate early from entoderm; continue distinct and migrate into the developing gonad via the mesoderm (see fig. 60)
Allen. 1911.
J. Morphol. 22
Species
Place of Origin, etc.
Author
Lophius piscatorius
Germ cells segregate from primitive entoderm; migrate through mesoderm to site of gonad; migration part passive and part active
Dodds. 1910.
J. Morphol. 21
Fundulus
heteroclitus
Germ cells segregate from peripheral entoderm lateral to posterior half of body; migrate through entoderm and mesoderm to the site of the developing gonad
Richards and Thompson. 1921.
Biol. Bull. 40
Cottus bairdii
Primordial germ cells derive from giant cells, in the primitive entoderm; migrate through the lateral mesoderm into the site of the developing gonad
Hann. 1927.
J. Morphol. 43
Lebistes reticulatus (guppy)
Germ cells segregate early in development; first seen in the entodermmesoderm area; migrate into the sites of the developing ovary and testis, giving origin to the definitive germ cells
Goodrich, Dee, Flynn, and Mercer. 1939. Biol. Bull. 67
Rana tempo raria
Germ cells segregate from primitive entoderm; migrate into developing genital glands
Witschi. 1914.
Arch. f. mikr. Anat. 85
Rana temporaria
Primordial germ cells from entoderm discharged at first spawning. Later, the definitive germs cells of adults originate from peritoneal cells
Gatenby. 1916.
Quart. J. Micr. Sc. 61
Rana catesbiana
Primordial germ cells segregate from primitive entoderm; definitive germ cell derives from primordial cells according to author’s view but admits possibility of germinal epithelium origin
Swingle. 1921.
J. Exper. Zool. 32
Rana temporaria, Triton alpestris, Bufo vulgaris
Primordial germ cells segregate from entoderm
Bounoure. 1924. Compt. rend. Acad. d. Sc. 178, 179
Rana sylvatica
Primordial germ cells originate from entoderm and migrate into the developing gonads. They give origin to the definitive sex cells
Witschi. 1929.
J. Exper. Zool. 52
H emidactylium scutatum
Primordial germ cells arise in mesoderm between somite and lateral plate; move to site of gonad by shifting of tissues
Humphrey. 1925.
J. Morphol. 41
Species
Place of Origin, etc.
Author
Ambystorna
maculatiirn
Most germ cells somatic in origin from germinal epithelium, although a few may come from primordial germ cells of entodermal origin
McCosh. 1930.
J. Morphol. 50
Triton, and Ambystorna mexicanum
Germ cells differentiate from lateral plate mesoderm
Nieuwkoop. 1946.
Arch. Neerl. de zool. 7
Chrysemys
marginata
(turtle)
Primordial germ cells from entoderm; most of definitive germ cells arise from peritoneal cells
Dustin. 1910.
Arch, biol., Paris. 25
Sternotherus
odoratus (turtle)
Primordial cells segregate early from entoderm; later definitive cells derive from these and from peritoneal epithelium
Risley. 1934.
J. Morphol. 56
Callus ( domesticus) gallus (chick)
Germ cells arise from primitive cells in entoderm of proamnion area and migrate by means of the blood vessels to the site of the developing gonad. Definitive germ cells of sex cords and later seminiferous tubules derive from primordial germ cells
Swift. 1914, 1916.
Am. J. Anat. 15, 20
Callus (domesticus) gallus (chick)
Primordial germ cells arise from enlodermal cells
Dantschakoff. 1931. Zeit. f. Zellforsch., mikr. Anat. 15
Chick and albino rat
Early primordial cells degenerate; definitive cells from peritoneal epithelium
Firket. 1920.
Anat. Rec. 18
Didelpliys
virginiana
(opossum)
Germ cells arise from germinal epithelium
Nelsen and Swain. 1942.
J. Morphol. 71
Mils musculiis (mouse)
Oogonia derived from primordial germ cells; spermatogonia from epithelial cells of testis cords
Kirkham. 1916.
Anat. Rec. 10
Mus musculus (mouse)
Primordial germ cells of ovary arise from germinal epithelium during development of the gonads. These presumably give origin to the definitive sex cells
Brambell. 1927.
Proc. Roy. Soc. London, s.B. 101
Felis dornestica (cat)
Primordial cells segregate early but do not give origin to definitive germ cells which derive from germinal epithelium
de Winiwarter and Sainmont. 1909. Arch, biol., Paris. 24
Species
Place of Origin, etc.
A uthor
Felis dome Stic a (cat)
Definitive ova derived from germinal epithelium of the ovary at an early stage of gonad development
Kingsbury. 1938.
Am. J. Anat. 15
Cavia porcellus (guinea pig)
Primordial germ cells from entodermal origin degenerate; the primordial germ cells derived from the germinal epithelium give rise to the definitive germ cells in the testis
Bookkout. 1937.
Zeit. f. Zellforsch, mikr. Anat. 25
Homo sapiens (man)
Primordial germ cells found in entoderm of yolk sac; migrate by ameboid movement into developing gonad
Witschi. 1948. Carnegie Inst., Washington Publ. 575. Contrib. to Embryol.' 32
C. Maturation (Differentiation) of the Gametes
1. General Considerations
Regardless of their exact origin definitive germ cells as primitive oogonia or very young oocytes are to be found in or near the germinal epithelium in the ovaries of all vertebrates in the functional condition (figs. 39B, 64). In the testis, the primitive spermatogonia are located within the seminiferous tubules as the germinal epithelium, in intimate association with the basement membrane of the tubule (figs. 65, 66).
The period of coming into maturity (maturation) of the gametes is a complicated affair. It involves profound transformations of the cytoplasm, as well as the nucleus. Moreover, a process of ripening or physiological maturing is necessary, as well as a morphological transformation. The phrase “maturation of the germ cells†has been used extensively to denote nuclear changes. However, as the entire gamete undergoes morphological and physiological change, the terms nuclear maturation, cytosomal maturation, and physiological maturation are used in the following pages to designate the various aspects of gametic development.
One of the most characteristic changes which the germ cell experiences during its maturation into a mature gamete is a reduction of chromatin material. Because of this, the germ cell which begins the maturing process is called a meiocyte. This word literally means a cell undergoing diminution and it is applied to the germ cell during meiosis or the period in which a reduction in the number of chromosomes occurs. The word haplosis is a technical name designating this reduction process.
Fig. 65. Semidiagrammatic representation of a part of the seminiferous tubule of the cat testis.
The word meiocyte thus is a general term applicable to both the developing male and female germ cells. On the other hand, the word spermatocyte is given to the developing male gamete during the period of chromosome diminution, whereas the word oocyte is applied to the female gamete in the same period. When, however, the period of chromosome diminution is completed and the chromosome number is reduced to the haploid condition, the developing male gamete is called a spermatid while the female gamete is referred to as an ootid or an egg. {Note: the word egg is applied often to the female gamete during the various stages of the oocyte condition as well as after the maturation divisions have been accomplished.)
The reduction of chromatin material is not the only effect which the meiotic process has upon the chromatin material, or possibly upon the developing cytosomal structures as well. This fact will become evident during the descriptions below concerning the meiotic procedures.
Another prominent feature of the gametes during the meiocyte period is their growth or increase in size. This growth occurs during the first part of the meiotic process when the nucleus is in the prophase condition and it involves both nucleus and cytoplasm. The growth phenomena are much more pronounced in the oocyte than in the spermatocyte. Due to this feature of growth, the oocyte and spermatocyte also are regarded as auxocytes, that is growing cells, a name introduced by Lee, 1897. The words meiocyte and auxocyte thus refer to two different aspects of the development of the oocyte and the spermatocyte.
2. Basic Structure of the Definitive Sex Cell as It Starts to Mature or Differentiate into the Male Meiocyte
(i.e., the Spermatocyte) or the Female Meiocyte (i.e., the Oocyte)
The definitive sex cells of both sexes have a similar cytological structure.
The component parts are (fig. 68):
- nucleus,
- investing cytoplasm,
- idiosome,
- Golgi substance, and
- chondriosomes.
The nucleus is vesicular and enlarged, and the nuclear network of chromatin may appear reticulated. A large nucleolus also may be visible. The
investing cytoplasm is clearer and less condensed in appearance than that
of ordinary cells. The idiosome (idiozome) is a rounded body of cytoplasm
which, in many animal species, takes the cytoplasmic stain more intensely
than the surrounding cytoplasm. Within the idiosome it is possible to demonstrate the centrioles as paired granules in some species. Surrounding the
idiosome are various elements of the Golgi substance, and near both the
idiosome and Golgi elements, is a mass of chondriosomes (mitochondria) of
various sizes and shapes. The idiosome and its relationship with the Golgi
material, the mitochondria, and the centrioles varies considerably in different
species of animals.
Much discussion has occurred concerning the exact nature of the idiosome.
Some investigators have been inclined to regard the surrounding Golgi substance as a part of the idiosome, although the central mass of cytoplasm containing the centrioles is the “idiosome proper†of many authors (Bowen, ’22).
Again, when the maturation divisions of the spermatocyte occur, the idiosome
and surrounding Golgi elements are broken up into small fragments. However, in the spermatids the Golgi pieces (dictyosomes) are brought together
once more to form a new idiosome-like structure, with the difference that the
latter “seems never to contain the centrioles†(Bowen, ’22). It is, therefore,
advisable to regard the idiosome as being separated into its various components during the maturation divisions of the spermatocyte and to view the
reassemblage of Golgi (dictyosomal) material in the spermatid as a different
structure entirely. This new structure of the spermatid is called the acroblast
(Bowen, ’22; Leuchtenberger and Schrader, ’50). (See fig. 68B.) A similar
breaking up of the idiosome occurs in oogenesis (fig. 68F, G). However, all
meiocytes do not possess a typical idiosome. This fact is demonstrated in
insect spermatocytes, where the idiosomal material is present as scattered
masses to each of which some Golgi substance is attached.
The various features which enter into the structure of the definitive germ cell do not behave in the same way in each sex during gametic differentiation. While the behaviors of the chromatin material in the male and female germ cells closely parallel each other (fig. 67), the other cytosomal features follow widely divergent pathways, resulting in two enormously different gametic entities (fig. 68A-H).
Fig. 66. Section of part of a seminiferous tubule of human testis. (Redrawn from Gatenby and Beams, ’35.)
Fig. 67. Diagrammatic representation of the nuclear changes occurring during meiosis
in spermatocyte and oocyte. Six chromosomes, representing three homologous pairs, are
used. Observe the effects of the crossing over of parts of chromatids. The diplotene condition of oocyte depicted by arrows and the enlarged nucleus. The haploid condition is
shown in each of the spermatids or in the egg and its three polocytes.
Fig. 68. Possible fate of the primitive meiocyte and its cytoplasmic inclusions when exposed to testicular or ovarian influences. Particular attention is given to the idiosome. Under male-forming influences the idiosome components are dispersed during the maturation divisions and are reassembled into three separate component structures, namely, (1) Acroblast of Golgi substance, (2) centriolar bodies, and (3) mitochondrial bodies (see B). Each of these structures, together with the post-nuclear granules of uncertain origin, play roles in spermatogenesis as shown. Under ovarian influences the idiosome is dispersed before the maturation divisions. The Golgi substance and mitochondria play (according to theory, see text) their roles in the formation of the deutoplasm.
3. Nuclear Maturation of the Gametes
Most of our information concerning the maturation of the nucleus pertains to certain aspects of chromosome behavior involved in meiosis, particularly the reduction of the chromosome number together with some activities of “crossing over†of materials from one chromosome to another. But our information is vague relative to other aspects of nuclear development. For example, we know little about the meaning of growth and enlargement of the nucleus as a whole during meiosis, an activity most pronounced in the oocyte. Nor do we know the significance of nuclear contraction or condensation in the male gamete after meiosis is completed. Therefore, when one considers the nuclear maturation of the gametes, it is necessary at this stage of our knowledge to be content mainly with observations of chromosomal behavior.
a. General Description of the Chromatin Behavior During Somatic and Meiotic Mitosis
As the maturation behavior of the chromatin components in the spermatocyte and oocyte are similar, a general description of these activities is given in the following paragraphs. Before considering the general features and details of the actions of the chromosomes during meiosis, it is best to recall some of the activities which these structures exhibit during ordinary somatic and gonial mitoses.
Cytological studies have shown that the chromosomes, in most instances, are present in the nucleus in pairs, each member of a pair being the homologue or mate of the other. Homologous chromosomes, therefore, are chromosomal pairs or mates. During the prophase condition in ordinary somatic and gonial mitoses, the various chromosomal mates do not show an attraction for each other. A second feature of the prophase stage of ordinary cell division is that each chromosome appears as two chromosomes. That is, each chromosome is divided longitudinally and equationally into two chromosomes. At the time when the metaphase condition is reached and the chromosomes become arranged upon the metaphase plate, the two halves or daughter chromosomes of each original chromosome are still loosely attached to each other. However, during anaphase, the two daughter chromosomes of each pair are separated and each of the two daughter nuclei receives one of the daughter chromosomes. Reproduction of the chromatin material and equational distribution of this material into the two daughter cells during anaphase is a fundamental feature of the ordinary type of somatic and gonial mitoses. The two daughter nuclei are thus equivalent to each other and to the parent nucleus. In this way, chromosomal equivalence is passed on ad infinitum through successive cell generations.
On the other hand, a different kind of chromosomal behavior is found during meiosis, which essentially is a specialized type of mitosis, known as a meiotic mitosis. In one sense it is two mitoses or mitotic divisions with only one prophase; that is, two metaphase-anaphase separations of chromosomes preceded by a single, peculiar prophase. The peculiarities of this meiotic prophase may be described as follows: As the prophase condition of the nucleus is initiated, an odd type of behavior of the chromosomes becomes evident — a behavior which is entirely absent from ordinary somatic mitosis: namely, the homologous pairs or mates begin to show an attraction for each other and they approach and form an intimate association. This association is called synapsis (figs. 67, 69, zygotene stage). As a result, the two homologous chromosomes appear as one structure. As the homologous chromosomes are now paired together and superficially appear as one chromosome, the number of “chromosomes†visible at this time is reduced to one-half of the ordinary somatic or diploid number. However, each “chromosome†is in reality two chromosomes and, therefore, is called a bivalent or twin chromosome.
Fig. 69. Steps in spermatogenesis in the grasshopper. In the center of the chart is represented a longitudinal section of one of the follicles of a grasshoper testis with its various regions of spermatogenic activity. In the upper right of the chart the apical-cell complex is depicted with its central apical cell, spermatogonia, and surrounding epithelial cells. The primary spermatogonia lie enmeshed between the extensions of the apical cell and the associations of these extensions with the surrounding epithelial elements of the complex. (Also see Wenrich, 1916, Bull. Mus. Comp. Zool. Harvard College, 60.)
While the homologous chromosomes are intimately associated, each mate
reproduces itself longitudinally just as it would during an ordinary mitosis
(fig. 67, pachytene stage). (The possibility remains that this reproduction of
chromatin material may have occurred even before the synaptic union.)
Hence, each bivalent chromosome becomes transformed into four potential
chromosomes, each one of which is called a chromatid. This group of chromatids is, collectively speaking, a tetrad chromosome. (As described below,
interchange of material or crossing over from one chromatid to another may
take place at this time.) As a result of these changes, the nucleus now contains the haploid number of chromosomes, (i.e., half of the normal, diploid
number) in the form of tetrads (fig. 67, pachytene stage). However, as each
tetrad represents four chromosomes, actually there is at this time twice the
normal number of chromosomes present in the nucleus (fig. 67; compare
leptotene, pachytene, diplotene and diakinesis).
The next step in meiosis brings about the separation of the tetrad chromosome into its respective chromatids and it involves two divisions of the cell.
These divisions are known as meiotic divisions. As the first of these two
divisions begins, the tetrad chromosomes become arranged in the mid- or
metaphase plane of the spindle. After this initial step, the first division of
the cell occurs, and half of each tetrad (i.e., a dyad) passes to each pole of
the mitotic spindle (fig. 67, first meiotic division). Each daughter cell (i.e.,
secondary spermatocyte or oocyte) resulting from the first maturation
(meiotic) division thus contains the haploid or reduced number of chromosomes in the dyad condition, each dyad being composed of two chromatids.
A resting or interphase nuclear condition occurs in most spermatocytes, following the first maturation division, but in the oocyte it usually does not
occur (fig. 69, interkinesis).
As the second maturation division is initiated, the dyads become arranged on the metaphase plate of the mitotic spindle. As division of the cell proceeds, half of each dyad (i.e., a monad) passes to the respective poles of the spindle (fig. 67, second meiotic division; fig. 69). As a result of these two divisions, each daughter cell thus contains the haploid or reduced number of chromosomes in the monad (monoploid) condition (fig. 67, spermatid or egg). Meiosis or chromatin diminution is now an accomplished fact.
It is to be observed, therefore, that the meiotic phenomena differ from those of ordinary mitosis by two fundamental features:
(1) In meiosis there is a conjugation (synapsis) of homologous chromosomes during the prophase stage, and while synapsed together each of the homologues divides equationally; and
(2) following this single prophase of peculiar character, two divisions follow each other, separating the associated chromatin threads.
While the meiotic prophase is described above as a single prophase preceding two metaphase-anaphase chromosome separations, it is essentially a double prophase in which the process of synapsis acts to suppress one of the equational divisions normally present in a mitotic division; a synapsed or double chromosome, therefore, is substituted for one of the longitudinal, equational divisions which normally appears during a somatic prophase. It is this substitution which forms the basis for the reduction process, for two mitotic divisions follow one after the other, preceded by but one equational splitting, whereas in ordinary mitosis, one equational splitting of the chromosomes always precedes each mitotic division.
b. Reductional and Equational Meiotic Divisions and the Phenomenon of Crossing Over
In the first meiotic division (i.e., the first maturation division), if the two chromatids which are derived from one homologous mate of the tetrad are separated from the two chromatids derived from the other homologous mate the division is spoken of as reductional or disjunctional. In this case the two associated chromatids of each dyad represent the original chromosome which synapsed at the beginning of meiotic prophase (fig. 67, tetrads B and C, first meiotic division). If, however, the separation occurs not in the synaptic plane but in the equational plane, then the two associated chromatids of each dyad come, one from one synaptic mate and one from the other; such a division is spoken of as an equational division (fig. 67, tetrad A, first meiotic division). There appears to be no fixity of procedure relative to the separation of the tetrads, and great variability occurs. However this may be, one of the two meiotic divisions as far as any particular tetrad is concerned is disjunctional (reductional) and the other is equational, at least in the region of the kinetochore (see p. 135 and fig. 70). If the first division is reductional, the second is equational and vice versa. Disjunction in the first maturation division is often referred to as pre-reduction, while that in the second maturation division is called post-reduction.
Fig. 70. Some of the various possibilities which may occur as a result of the exchanges
of parts of chromatids during the crossing-over phenomena associated with meiosis. Two
chiasmata (singular, chiasma) are shown in (A), (C), (E). Observe that homologous
chromosome A has split equationally into chromatids A and A', while homologous
chromosome B has divided equationally into B and B'. The resulting interchanges between respective chromatids of the original homologous chromosomes are shown in (B),
(D), (F). The kinetochore (place of spindle-fiber attachment) is indicated by the oval
or circular area to the left of the chromatids. (Modified from White: Animal Cytology
and Evolution, London, Cambridge University Press, 1943.)
The foregoing statement regarding disjunctional and equational divisions
should be considered in the light of the phenomenon of crossing over. In
the latter process, a gene or groups of genes may pass from one chromatid
to the other and vice versa during their association at the four strand stage
(fig. 70). In the region of the centromere or kinetochore (i.e., the point)
of the achromatic, spindle-fiber attachment) and nearby regions, cross overs
are thought not to occur (fig. 70, kinetochore). Consequently, in the regions
of the kinetochore, the statements above regarding disjunctional and equational divisions of the chromosomes appear to be correct. However, the terms
disjunctional and equational may mean little in other regions of the chromosomes of a tetrad during the meiotic divisions. For example, let us assume as
in fig. 70 (see also fig. 67), that we have chromatids A and A', B and B',
A and B representing the original homologues or synaptic chromosomes which
have divided into these chromatids respectively. Then during the tetrad stage
of association or slightly before, let us assume that there has been a crossing
over of genes from chromatid A to chromatid B and from chromatid B to
chromatid A in a particular area (fig. 70A). (It is to be observed that chromatids A' and B' are not involved in this particular instance.) Further, let us
assume that AA' and BB' as a whole are separated at the first maturation
division, the kinetochore and immediate regions would represent a disjunctional division, but for the particular area where crossing over is accomplished,
the division would be equational (fig. 70A, B; central portions of chromatids
A and B in fig. 70B). Thus, it would be for other regions where cross overs
may have occurred. Other cross-over possibilities are shown in fig. 70C-F.
c. Stages of Chromatin Behavior During the Meiotic Prophase in Greater Detail
The following five stages of chromatin behavior within the prophase nucleus during meiosis are now in common usage. They are based on the stages originally described by H. von Winiwarter, ’00. The substantive form is presented in parentheses.
1) Leptotene (Leptonema) Stage. The leptotene stage (figs. 69, 71) represents the initial stage of the meiotic process and is seen especially well in the spermatocyte. At this time the nucleus of the differentiating germ cell begins to enlarge, and the diploid number of very long, slender chromatin threads make their appearance. (Compare “resting†and leptotene nuclei in figs. 69, 71.) The chromatin threads may lie at random in the nucleus or they may be directed toward one side, forming the so-called “bouquet†condition (fig. 69, leptotene stage) . The nucleolus is evident at this time (fig. 7 IB) .
2) Zygotene or Synaptene (Zygonema) Stage. The zygotene stage (figs. 69, 71, 85) is characterized by a synapsis of the chromatin threads. This synapsis or conjugation occurs between the homologous chromosomes, that is, the chromosomes which have a similar genic constitution. Synapsis appears to begin most often at the ends of the threads and progresses toward the middle (fig. 67, zygotene). At this stage the chromatin threads may show a strong tendency to collapse and shrink into a mass toward one end of the nucleus (fig. 85C, D). This collapsed condition, when present, is called synizesis. The zygotene stage gradually passes into the pachytene condition.
Fig. 71. Certain aspects of the oocyte nucleus during the meiotic prophase. (A-G)
Chromatin and nuclear changes in the oocyte of the cat up to the diplotene condition
when the germinal vesicle is fully developed. (After de Winiwarter and Sainmont, Arch,
biol., Paris, 24.) (H, I) Germinal vesicle in the dogfish, Scy Ilium canicula, and in Amphioxus. (After Marechal, La Cellule, 24.) Observe the typical “lamp-brush†chromosome conditions in the germinal vesicle of the shark oocyte. These lamp-brush chromosomes are developed during the diplotene stage of meiosis by great attenuation of the
chromosomes and the formation of lateral extensions or loops from the sides of the
chromosomes.
3) Pachytene (Pachynema) Stage. Gradually, the synapsis of the homologous chromosomes becomes more complete, and the threads appear shorter
and thicker. The contracted threads in this condition are referred to as
pachynema (figs. 69, 71, 85E). The nucleus in this manner comes to contain
a number of bivalent chromosomes, each of which is made up of two homologous mates arranged side by side in synaptic union, known technically as
parasynapsis. (Telosynapsis probably is not a normal condition.) Consequently, the number of chromosomes now appears to be haploid. Each pachytene chromosome (i.e., each of the pair of homologous chromosomes) gradually divides equationally into two daughter thread-like structures, generally
referred to as chromatids. The exact time at which division occurs during
meiosis is questionable. The entire group of four chromatids which arise from
the splitting of the synapsed homologues is called a tetrad.
4) Diplotene (Diplonema) Stage. In the diplotene stage (figs. 67, 69, 71, 85F, G), two of the chromatids tend to separate from the other two. (See fig. 70A, C, E.) The four chromatids in each tetrad may now be observed more readily, at least in some species, because the various chromatids of each tetrad show a repulsion for one another, and the chromatids move apart in certain areas along their length. This condition is shown in both the male and female meiocyte, but in the latter, the repulsion or moving apart is carried to a considerable degree and is associated with a great lengthening and attenuation of the chromatids. (See fig. 67.) In the female meiocyte at this stage, the chromosomes become very diffuse and are scattered throughout the nucleus, somewhat resembling the non-mitotic condition (figs. 71F-T, 72B-E). The peculiar behavior of the chromosomes and nucleus of the oocyte in the diplotene stage of meiosis is described more in detail on p. 141.
Although there is a tendency for the chromatids to widen out or separate from each other at this time, they do remain associated in one or more regions. In these regions of contact, the paired chromatids appear to exchange partners. This point of contact is called a chiasma (plural, chiasmata). Hence, a chiasma is the general region where the chromatids appear to have exchanged partners when the tetrad threads move apart in the diplotene state. (See fig. 70, chiasmata.)
5) Diakinesis. The diplotene stage gradually transforms into the diakinesis state (figs. 67, 69, 72F, 85H) by a process of marked chromosomal contraction. There also may be an opening up of the tetrads due to a separation of the homologous mates in the more central portions of the tetrad, with the result that only the terminal parts of the chromatids remain in contact. This latter process is called “terminalization.†Coincident with this partial separation, a further contraction of the tetrads may occur. As a result, at the end of diakinesis the tetrads may assume such curious shapes as loops, crosses, rings, etc., scattered within the nucleus of the female and male meiocyte (fig. 69, diakinesis). The nuclear membrane eventually undergoes dissolution, and
Fig. 72. Growth of the nucleus during meiosis in the amphibian egg, showing the enlarged germinal vesicle and diplotene lamp-brush chromosomes with lateral loops. (A) Early diplotene nucleus of the frog. (B, C, E) Different phases of the diplotene nucleus in this form. These figures are based upon data provided by Duryee (’50) and sections of the frog ovary. (D) Drawing of the unfixed germinal vesicle of Triturus. Some aspects of the attenuate chromatin threads with lateral loops are shown. The nucleoli are numerous and occupy the peripheral region of the germinal vesicle. (F) Semidiagrammatic drawing of the later phases of the developing frog egg. It shows the germinal vesicle assuming a polar condition, with the initial appearance of germinal vesicle shrinkage before the final dissolution of the nuclear membrane. Observe that the chromosomes are contracting and now occupy the center of the germinal vesicle.
Fig. 73. Various aspects of Sertoli-cell conditions in the fowl. (Redrawn from Zlotnick,
Quart. J. Micr. Sc., 88.) (A) Resting Sertoli cell, showing mitochondria. (B) Sertoli element at the beginning of cytoplasmic elongation. (C) Sertoli cell with associated
late spermatids.
Fig. 74. Types of chordate sperm. All the chordate sperm belong to the flagellate
variety. (A) Amphioxus (protochordate). (B) Salmo (teleost). (C) Perea (teleost).
(D) Petromyzon (cyclostome). (E) Raja (elasmobranch). (F) Bufo (anuran). (G)
Rana (anuran). (H) Salamandra (urodele). (I) Anguis (lizard). (J) Crex (bird).
(K) Fringilla (bird). (L) Turdus (bird). (M) Echidna (monotrematous mammal).
(N) Mus (eutherian mammal). (O, P) Man (full view and side view, respectively).
the tetrads become arranged on the metaphase plate of the first maturation division. (See figs. 69, first maturation division; 72F, 119A, B.) This division is described on pp. 132 and 133.
d. Peculiarities of Nuclear Behavior in the Oocyte During Meiosis; the Germinal Vesicle
Although the movements of the chromosomes during meiosis in the developing male and female gamete appear to follow the same general behavior
Fig. 75. Non-flagellate sperm. (A-C) Ameboid sperm of Polyphemus. (After Zacharias.) (D) Lobster, Homarus. (After Herrick.) (E) Decapod Crustacea, Galathea (Anomura). (After Koltzoff.) (F) Nematode woim, Ascaris.
Fig. 76. Conjugate sperm of grasshopper associated temporarily to form the “sperm boat.â€
pattern (fig. 67), some differences do occur. For example, in the female when the diplotene stage is reached, the repulsion of the tetrad threads is greater (figs. 67, $ and 9 \ 12). Furthermore, the chromatids elongate and become very attenuate although they appear to retain their contacts or chiasmata (fig. 72). Side loops and extensions from the chromatids also may occur, especially in those vertebrates with large-yolked eggs (e.g., amphibia, fishes, etc.). (See figs. 71H, 72B-D.) When these lateral extensions are present, the chromosomes appear diffuse and fuzzy, taking on the characteristics which suggest their description as “lamp-brush†chromosomes. Another difference of chromatic behavior is manifested by the fact that the chromosomes in the developing female gamete during the diplotene stage are not easily stained by the ordinary nuclear stains, whereas the chromosomes in the spermatocyte stain readily.
Fig. 77. Spatula-type sperm of various mammals. (Compiled from Bowen; Gatenby
and Beams; Gatenby and Woodger; see references in bibliography.) Observe the vacuole
inside the head of the sperm. Gatenby and Beams found that this vacuole, in some
instances, stains similar to a nucleolus, but suggest it may be a hydrostatic organ, or
respiratory structure. (P. 20, Quart. J. Micr. Sc., 78.)
Aside from the differences in chromosomal behavior, great discrepancies
in the amount of growth of the nucleus occur in the two gametes during meiosis.
The nucleus of the oocyte greatly increases in size and a large quantity of
nuclear fluid or sap comes to surround the chromosomes (figs. 7 IF, G; 72C,
^ F). Correlated with this increase in nuclear size, the egg grows rapidly.
and deutoplasmic substance is deposited in the cytoplasm (fig. 68F-H). As differentiation of the oocyte advances, the enlarged nucleus or germinal vesicle assumes a polar position in the egg (figs. 68H, 70F). When the oocyte has finished its growth and approaches the end of its differentiation, the
Fig. 78. Different shapes and positions of the acrosome. (A) Type of acrosome
found in Mollusca, Echinodermata, and Annelida. (B) Reptilia, Aves, and Amphibia.
(C) Lepidoptera. (D) Mammalia. (E) Many Hemiptera and Coleoptera. (After
Bowen, Anat. Rec., 28.) (F) Sperm of certain birds, i.e., finches. (After Retzius, Biol.
Untersuchungen, New Series 17, Stockholm, Jena.) Observe the well-developed acrosome in the form of a perforatorium. The spiral twist of the acrosome shown in this drawing is characteristic of passerine birds.
Fig. 79. Sperm of urodele amphibia. (After Meves, 1897, Arch. f. mikr. Anat. u.
Entwichlingsgesch., 50; McGregor, 1899, J. Morphol., 15. (A~E) Stages in the morphogenesis of the sperm of Salamandra. (F) Diagram of head, middle piece, etc. of the
sperm of the urodele.
chromosomes within the germinal vesicle condense once again, decrease in length (fig. 72F), and assume conditions more typical of the diakinesis stage (figs. 67; 1 19A). The tetrad chromosomes now become visible. Following the latter chromosomal changes, the nuclear membrane breaks down (fig. 119A), and the chromatin elements pass onto the spindle of the first maturation division (fig. 119B). The nuclear sap, membrane, nucleolus, and general framework pass into the surrounding cytoplasmic substance (figs. 119A; 132A-C). This nuclear contribution to the cytoplasm appears to play an important part in fertilization and development, at least in some species (fig. 132C; the clear protoplasm is derived from the nuclear plasm).
e. Character of the Meiotic (Maturation) Divisions in the Spermatocyte Compared with Those of the Oocyte
1) Dependent Nature of the Maturation Divisions in the Female Meiocyte.
The maturation divisions in the developing male gamete occur spontaneously and in sequence in all known forms. But in most oocytes, either one or both of the maturation divisions are dependent upon sperm entrance. For example, in Ascaris, a nematode worm (fig. 133), and in Nereis, a marine annelid worm (fig. 130), both maturation divisions occur after the sperm has entered and are dependent upon factors associated with sperm entrance. A similar condition is found in the dog (van der Stricht, ’23; fig. 115) and in the fox (Asdell, ’46). In the urochordate, Styela, the germinal vesicle breaks down, the nuclear sap and nucleolus move into the surrounding protoplasm, and the first maturation spindle is formed as the egg is discharged into the sea water (fig. 116A, B). Further development of the egg, however, awaits the entrance of the sperm (fig. 116C--F). Somewhat similar conditions are found in other Urochordata. In the cephalochordate, Amphioxus, and in the vertebrate group as a whole (with certain exceptions) the first polar body is formed and the spindle for the second maturation division is elaborated before normal sperm entrance (figs. 117C, D; 119D). The second maturation division in the latter instances is dependent upon the activities aroused by sperm contact with the oocyte. In the sea urchin, sperm can penetrate the egg before the maturation divisions occur; but, under these conditions, normal development of the egg does not occur. Normally in this species both maturation divisions are effected before sperm entrance, while the egg is still in the ovary. When the egg is discharged into the sea water, the sperm enters the egg, and this event affords the necessary stimulus for further development (fig. 131).
2) Inequality of Cytoplasmic Division in the Oocyte. When the first maturation division occurs, the two resulting cells are called secondary spermatocytes in the male and secondary oocytes in the female (figs. 67, 69). The secondary spermatocytes are smaller both in nuclear and cytoplasmic volume. They also form a definite nuclear membrane. Each secondary spermatocyte then divides and forms two equal spermatids. In contrast to this condition of equality in the daughter cells of the developing male gamete during and following the maturation divisions, an entirely different condition is found in the developing female gamete. In the latter, one of the secondary oocytes is practically as large as the primary oocyte, while the other or first polar body (polocyte) is extremely small in cytoplasmic content although the nuclear material is the same (fig. 117D). During the next division the secondary oocyte behaves in a manner similar to that of the primary oocyte, and a small second polocyte is given off, while the egg remains large (fig. 117E, F). Unlike the secondary spermatocyte, the secondary oocyte does not form a nuclear membrane. The polar body first formed may undergo a division, resulting in a total of three polar bodies (polocytes) and one egg (ootid).
/. Resume of the Significance of the Meiotic Phenomena
In view of the foregoing data with regard to the behavior of the male and female gametes during meiosis, the significant results of this process may be summarized as follows:
(1) There is a mixing or scrambling of the chromatin material brought about by the crossing over of genic materials from one chromatid to another.
(2) Much chromatin material with various genic combinations is discarded during the maturation divisions in the oocyte. In the latter, two polar bodies are ejected with their chromatin material as described above. The egg thus retains one set of the four genic combinations which were present at the end of the primary oocyte stage; the others are lost. (A process of discarding of chromatin material occurs in the male line also. For although four spermatids and sperm normally develop from one primary spermatocyte, great quantities of sperm never reach an egg to fertilize it, and much of the chromatin material is lost by the wayside.)
(3) A reduction of the number of chromosomes from the diploid to the haploid number is a significant procedure of all true meiotic behavior.
(For more detailed discussions and descriptions of meiosis, see De Robertis, et al., ’48; Sharp, ’34, ’43; Snyder, ’45; White, ’45.)
4. Cytosomal (Cytoplasmic) Maturation of the Gametes a. General Aspects of Cytoplasmic Maturation of the Gametes
During the period when the meiotic prophase changes occur in the nucleus of the oocyte, the cytoplasm increases greatly and various aspects of cytoplasmic differentiation are effected. That is, differentiation of both nuclear and cytoplasmic materials tend to occur synchronously in the developing
Fig. 80. Morphogenesis of guinea-pig and human sperm. (A) Spermatocyte of guinea pig before first maturation division. The Golgi complex with included proacrosomic granules and centrioles is shown. (After Gatenby and Woodger, ’21.) (B) Young sister
spermatids of guinea pig. (C) Later spermatid of guinea pig showing acroblast with proacrosomic granules. (D) Young human spermatocyte, showing Golgi apparatus with proacrosomic granules similar to that shown in (A). (After Gatenby and Beams, ’35.) (E) Spermatid of guinea pig later than that shown in (C), showing acroblast with Golgi substance being discarded from around the acroblast. (F) Later human spermatid, showing Golgi substance surrounding acroblast with acrosome bead. (After Gatenby and Beams, ’35.) (G) Later human spermatid, showing acroblast, with acrosome bead
within, surrounded by a vacuole. (After Gatenby and Beams, ’35.) (H) Later spermatid
of guinea pig, showing outer and inner zones of the acrosome. The inner zone corresponds somewhat to the acrosome bead shown in (G) of the human spermatid. (After Gatenby and Wigoder, Proc. Roy. Soc., London, s.B., 104.)
female gamete. In the male gamete, on the other hand, the meiotic processes are completed before morphological differentiation of the cytoplasm is initiated.
Another distinguishing feature in the morphogenesis of the sperm relative to that of the egg is that the cytoplasmic differentiation of the sperm entails a discarding of cytoplasm and contained cytoplasmic structures, whereas the oocyte conserves and increases its cytoplasmic substance (fig. 68). In regard to the behavior of the cytoplasms of the two developing gametes, it is interesting to observe that the idiosome-Golgi-mitochondrial complex behaves very differently in the two gametes (fig. 68).
A third condition of egg and sperm differentiation involves the possible function of the “nurse cells.†In the vertebrate ovary the follicle cells which surround the egg have much to do with the conditions necessary for the differentiation of the oocyte. The latter cannot carry the processes of differentiation to completion without contact with the surrounding follicle cells. Spermiogenesis also depends upon the presence of a nurse cell. In the vertebrate seminiferous tubule, the Sertoli cell is intimately concerned with the transformation of the spermatid into the morphologically adult sperm, and a close contact exists between the developing sperm element and the Sertoli cell during this period (figs. 65, 66, 73). In the discharge of the formed sperm elements into the lumen of the tubule, the Sertoli cell also is concerned (Chap. 1).
b. Morphogenesis (Spermiogenesis; Spermioteleosis) of the Sperm
1) Types of Sperm. There are two main types of sperm to be found in animals, namely, flagellate and non-flagellate sperm (figs. 74, 75). Flagellate sperm possess a flagellum or tail-like organelle; non-flagellate sperm lack this structure. The flagellate type of sperm is found quite universally among animals; non-flagellate sperm occur in certain invertebrate groups, particularly in the nematode worms, such as Ascaris, and in various Crustacea, notably the lobster, crab, etc. (fig. 75). Flagellate sperm may be either uniflagellate or biflagellate. Single flagellate sperm occur in the majority of animals, while a biflagellate form is found in the platode, Procerodes. However, biflagellate sperm may be found as abnormal specimens among animals normally producing uniflagellate sperm.
Conjugate sperm are produced in certain animal species. For example, two sperm heads adhere closely together in the opossum (fig. 125), also in the beetle, Dytiscus, and in the gastropod, Turritella. Many sperm heads become intimately associated in the grasshopper to form the so-called “sperm boat†(fig. 76). However, all conjugate sperm normally separate from each other in the female genital tract.
2) Structure of a Flagellate Sperm. The flagellate sperm from different species of animals vary considerably in size, shape, and morphological details. Some possess long, spear-shaped heads, some have heads resembling a hatchet, in others the head appears more or less cigar-shaped, while still others possess a head which resembles a spatula (fig. 74). The spatula-shaped head is found in the sperm of the bull, opossum, man, etc. The description given below refers particularly to the spatula-shaped variety. Although all flagellate sperm resemble one another, diversity in various details is the rule,
Fig. 81. Later stages of human spermatogenesis. (Redrawn from Gatenby and Beams,
1935.)
Fig. 82. Stages of guinea-pig spermatogenesis. Observe dual nature of the acrosome; also, middle-piece bead (kinoplasmic droplet). (A-C redrawn from Gatenby and Beams, 1935; D redrawn from Gatenby and Woodger, ’21.)
and the description given below should be regarded as being true of one type of sperm only and should not be applied to all flagellate sperm.
A fully differentiated spatulate sperm of the mammals possesses the following structural parts (fig. 77).
a) Head. Around the head of the sperm there is a thin, enveloping layer of cytoplasm. This cytoplasmic layer continues posteriad into the neck, middle piece, and tail. Within the cytoplasm of the head is the oval-shaped nucleus. Over the anterior half of the nucleus the apical body or acrosome is to be found, forming, apparently, a cephalic covering and skeletal shield for the nucleus. The caudal half of the nucleus is covered by the post-nuclear cap. This also appears to be a skeletal structure supporting this area of the nucleus; moreover, it affords a place of attachment for the anterior centrosome and the anterior end of the axial filament.
In human and bull sperm the acrosome is a thin cap, but in some mammalian sperm it is developed more elaborately. In the guinea pig it assumes the shape of an elongated, shovel-shaped affair (fig. 82), while in the mouse and rat it is hatchet or lance shaped (fig. 74N). In passerine birds the acrosome is a pointed, spiral structure often called the perforatorium (fig. 78). On the other hand, in other birds, reptiles, and amphibia it may be a simple, pointed perforatorial structure (figs. 74, 78, 79). In certain invertebrate species, it is located at the caudal or lateral aspect of the nucleus (figs. 75, 78).
b) Neck. The neck is a constricted area immediately caudal to the posterior nuclear cap and between it and the middle piece. Within it are found the anterior centriole and the anterior end of the axial filament. In this particular region may also be found the so-called neck granule.
c) Connecting Body or Middle Piece. This region is an important portion of the sperm. One of its conspicuous structures is the central core composed of the axial filament and its surrounding cytoplasmic sheath. At the distal end of the middle piece, the central core is circumscribed by the distal, or ring centriole. Investing the central core of the middle piece is the mitochondrial' sheath. The enveloping cytoplasm is thicker to some degree in this area of the sperm than that surrounding the head.
d) Flagellum. The flagellum forms the tail or swimming organ of the sperm. It is composed of two general regions, an anterior principal or chief piece and a posterior end piece. The greater part of the axial filament and its sheath is found in the flagellum. A relatively thick layer of cytoplasm surrounds the filament and its sheath in the chief-piece region of the flagellum, but, in the caudal tip or end piece, the axial filament seems to be almost devoid of enveloping cytoplasm. The end piece often is referred to as the naked portion of the flagellum.
In figure 79 is shown a diagrammatic representation of a urodele amphibian sperm. Two important differences from the mammalian sperm described above are to be observed, namely, the middle piece is devoid of mitochondria and is composed largely of centrioles 1 and 2, and the tail has an elaborate undulating or vibratile filament associated with the chief piece.
3) Spermiogenesis or the Differentiation of the Spermatid into the Morphologically Differentiated Sperm. The differentiation of the spermatid into the fully metamorphosed sperm is an ingenious and striking process. It involves changes in the nucleus, during which the latter as a whole contracts and in some forms becomes greatly elongated into an attenuant structure. (See figs. 79B-F; 85L-P.) It also is concerned with profound modifications of the cytoplasm and its constituents; the latter changes transform the inconspicuous
Spermatid into a most complicated structure. Some of these changes are outlined below.
a) Golgi Substance and Acroblast; Formation of the Acrosome. The Golgi substance or parts thereof previously associated with the idiosome of the spermatocyte (fig. 80A) proceeds to form the acrosome of the developing spermatid as follows: In the differentiating human sperm, the Golgi substance of the spermatocyte (fig. SOD) becomes aggregated at the future anterior end of the nucleus, as shown in fig. 80F, where it forms an acroblast within a capsule of Golgi substance. This acroblast later forms a large vacuole within which is the acrosomal “bead†(figs. 68B; 80G). The acrosomal bead proceeds to form the acrosomal cap, shown in figure 81 A, and the latter grows downward over the anterior pole of the nucleus (fig. 81 A, B). Most of the Golgi substance in the meantime is discarded (fig. 81 A, B). (Sec Gatenby and Beams, ’35.)
In the guinea pig the acroblast together with other Golgi substance, migrates around the nucleus toward the future anterior pole of the latter where the acroblast takes up its new position (fig. 80B, C, E). (See Gatenby and Woodger, ’21.) As shown in figure 80E, the acroblast is composed of inner and outer acrosomal substances. These inner and outer areas of the acroblast give origin respectively to the inner and outer zones of the acrosome (fig. 82). The peripheral or surrounding Golgi material of the acroblast detaches itself meanwhile from the developing acrosome (fig. 80E, H) and drifts downward toward the posterior end of the sperm. Eventually it is discarded with the excess cytoplasm and some mitochondrial material. In some animal species (e.g., grasshopper) the acrosomal substance arises from a multiple type of acroblast (Bowen, ’22). (See fig. 83.) Nevertheless, the general process of acrosome formation is similar to that outlined above.
b) Formation of the Post-nuclear Cap. All spatulate sperm of mam
Fig. 83. Formation of the acrosome from a multiple acroblast in the grasshopper. (After Bowen, Anat. Rec., 24.)
Fig. 84. The mitochondrial nebenkern and its elaborate development in Brachynema. (After Bowen, J, Morphol., 37 and Biol. Bull., 42.) (B-I) Division of the nebenkern
(A) and its elaboration into two attenuant strands extending posteriad into the flagellum.
mals possess a nucleus which has an acrosomal cap over its anterior aspect and a post-nuclear cap covering its posterior area. Both of these caps tend to meet near the equator of the nucleus (fig. 77).
The exact origin of the post-nuclear cap is difficult to ascertain. In the human sperm it appears to arise from a thickened membrane in association with centriole 1 (fig. 80G, post-nuclear membrane). This membrane grows anteriad to meet the acrosomal cap (fig. 81A-C). In the sperm of the guinea pig, a series of post-nuclear granules in the early spermatid appear to coalesce to form the post-nuclear cap (fig. 82A-C).
c) Formation of the Proximal and Distal Centrioles; Axial Filament. While the above changes in the formation of the acrosome are progressing, the centriole (or centrioles) of the idiosome move to the opposite side of the nucleus from that occupied by the forming acrosome, and here in this position the proximal and distal centrioles of the future sperm arise. In this area the neck granules also make their appearance (figs. 68B; 80F-H). The axial filament arises at this time and it probably is derived from the two centrioles simultaneously (fig. 80F, H). The centrioles soon become displaced along the axial filament, the caudal end of which projects from the surface of the cell membrane (fig. 80F-H). The axial filament grows outward posteriorly from the cell membrane in line with the two centrioles and the acrosome-forming material. The anterior-posterior elongation of the sperm thus begins to make its appearance (fig. 80H). The anterior centriole retains a position close to the nuclear membrane, but the posterior or ring centriole moves gradually posteriad toward the cell surface (figs. 81, 82A-C).
d) Mitochondrial Material and Formation of the Middle Piece OF THE Sperm. The behavior of the mitochondria in the formation of sperm varies greatly. In the spatulate sperm described above, a portion of the mitochondrial substance becomes aggregated around the axial filament in the middle-piece area (figs. 77, 82D). In certain amphibian sperm the middle piece appears to be formed mainly by centrioles 1 and 2 (fig. 79D-F). In certain insects the mitochondrial body or nebenkern, divides into two masses which become extended into elongated bodies associated with the flagellum (fig. 84). Some of the mitochondrial substance is discarded with the Golgi substance and excess cytoplasmic materials.
e) The Cytoplasm, Axial Filament, Mitochondria, and Tail Formation. Synchronized with the above events, the cytoplasm becomes drawn out in the posterior direction, forming a thin cytoplasmic layer over the sperm head, and from thence posteriad over the middle piece and the chief piece of the flagellum. However, the end piece of the flagellum may be devoid of investing cytoplasm (fig. 77). As the cytoplasm is elongating posteriorly over the contained essential structures of the forming sperm, much of the cytoplasm and Golgi substance and some mitochondria are discarded and lost from the sperm body. It may be that these discarded bodies form a part of the essential substances of the spermatic (seminal) fluid. (See Chap. 1.) (See figs. 66; 68B-E; 81; 82; 85M-0.)
The centralized core of the tail is the axial filament which arises in relation to centrioles 1 and 2 and grows posteriad through the middle piece and tail
(figs. 80F-H; 81A-C; 82A-~C; 85M-P). A considerable amount of mitochondrial material may also enter into the formation of tail (fig. 84).
A peculiar, highly specialized characteristic of many sperm tails is the development of a vibratile membrane associated with the axial filament (fig. 79E, F). Its origin is not clear, but it probably involves certain relationships with the mitochondrial material as well as the cytoplasm and axial filament.
In the formation of the human and guinea-pig sperm, the nucleus experiences only slight changes in shape from that of the spermatid. However, in many animal species, spermiogenesis involves considerable nuclear metamorphosis as well as cytoplasmic change (figs. 69, 79, 85).
In summary it may be stated that while the various shapes and sizes of mature flagellate sperm in many animal species, vertebrate and invertebrate,
Fig. 85. Spermatogenesis in the common fowl. Observe extreme nuclear metamorphosis. (After Miller, Anat. Rec., 70.) (A) Resting spermatocyte. (B) Early leptotene stage. (C, D) Synaptene stage. (E) Pachytene stage. (F, G) Diplotene stage. (H) Diakinesis. (I) First division, primary sperm. (J-P) Metamorphosing sperm.
are numerous, there is a strong tendency for spermiogenesis to follow similar
lines of development. Deviations occur, but the following comparisons between
mammalian and insect spermiogenesis, somewhat modified from Bowen (’22),
illustrate the uniformity of transformation of the basic structures of the
primitive meiocyte:
Mammalian Sperm
Insect Sperm
Nucleus — head
Nucleus — head
Centrioles — originally double and arranged in a proximal-distal formation. The axial filament arises from both centrioles
Centrioles — same as in mammals
Mitochondria — form an elaborate sheath for the anterior portion of the axial filament
Mitochondria — form a somewhat similar sheath for the axial filament
Idiosome and Golgi apparatus (acroblast portion) — gives origin to a vesicle which contains a granule, the acrosome granule, which is involved in the production of the acrosome
Idiosome and Golgi apparatus — much the same as in mammals
Excess Golgi substance — cast off with excess cytoplasm
Excess Golgi substance — cast off with excess cytoplasm
Excess cytoplasm — cast off — may be part of seminal fluid or possibly may be engulfed by Sertoli cells
Excess cytoplasm — cast off — may be part of seminal fluid or possibly may be engulfed by epithelial cells of the sperm cyst wall
c. Cytoplasmic Differentiation of the Egg
The cytoplasmic differentiation of the egg involves many problems. These problems may be classified under three general headings, viz.:
(1) Formation of the deutoplasm composed of fats, carbohydrates and proteins,
(2) development of the invisible organization within the true protoplasm or hyaloplasm, and finally,
(3) formation of the vitelline or egg membrane or membranes.
In view of the complexity of these three problems and of their importance to the egg in the development of the new individual, the mature oocyte or egg is in a sense no longer a single cell. Rather, it is a differentiated mass of protoplasm which is capable, after proper stimulation, to give origin to a new individual composed of many billions of cells. As such, the differentiation of the oocyte within the ovary represents a relatively unknown period of embryological development.
Fig. 86. Young oogonia of the fowl entering the growth (oocyte) stage. (A) Idiosome from which the Golgi substance has been removed and stained to show the centrosphere (archoplasm). The centrosome has two centrioles. (B) Idiosome with surrounding Golgi substance. The mitochondria surround the Golgi substance and the nucleus. (After Brambell, ’25.)
Fig. 87. The so-called mitochondrial yolk body in the developing egg of the fowl. (A) Oocyte from 11 -week-old chick, showing mitochondrial cloud and Golgi substances I and II. (B) Oocyte from ovary of adult fowl, showing both types of Golgi substance and mitochondrial cloud. (C) Oocyte from ovary of adult fowl, showing the appearance of the mitochondrial yolk body within the mitochondrial cloud. (D) Oocyte from ovary of adult fowl, showing fragmentation of Golgi substance 1 and the association of the resulting Golgi granules around the mitochondrial yolk body. (After Brambell, ’25.)
Fig. 88. Portion of follicle and periphery of oocyte from ovary of the adult bird,
showing the mitochondria and their transformation into the M-yolk spheres of Brambell.
(After Brambell, ’25.)
Before considering the various aspects of cytoplasmic differentiation of the oocyte, it is best for us to review the types of vertebrate and other chordate eggs in order to be able to visualize the various goals toward which the developing oocyte must proceed.
1) Types of Chordate Eggs. Eggs may be classified according to the amount of deutoplasm (yolk, etc.) present in the cytoplasm as follows:
a) Homolecithal (Isolecithal) Eggs. True homolecithal eggs in the phylum Chordata are found only in the mammals, exclusive of the Prototheria. Here the deutoplasm is small in amount, and is present chiefly in the form of fat droplets and small yolk spherules, distributed in the cytoplasm of the egg (figs. 118A, B; 147A).
b) Telolecithal Eggs. In the telolecithal egg the yolk is present in considerable amounts and concentrated at one pole. Telolecithality of the egg in the phylum Chordata exists in various degrees. We shall arrange them in sequence starting with slight and ending with very marked telolecithality as follows:
(1) Amphioxus and Styela. In Amphioxus and Styela from the subphyla Cephalochordata and Urochordata, respectively, the yolk present is centrally located in the egg before fertilization but becomes concentrated at one pole at the time of the first cleavage where it is contained for the most part within the future entoderm cells (figs. 132D, 167A).
(2) In many Amphibia, such as the frogs and toads, and also in the Petromyzontidae or fresh-water lampreys among the cyclostome fishes, the yolk present is greater in amount than in the preceding eggs. As such, it is concentrated at one pole, the future entodermal or vegetal pole, and a greater degree of telolecithality is attained than in the eggs of Amphioxus or Styela (fig. 141 A).
(3) In many Amphibia, such as Necturus, also in Neoceratodus and Lepidosiren among the lung fishes, and in the cartilaginous ganoid fish, Acipenser, yolk is present in considerable amounts, and the cytoplasm of the animal pole is smaller in comparison to the yolk or vegetal pole (figs. 150, 151, 152).
(4) In the bony ganoid fishes, Amia and Lepisosteus, as well as in the Gyrnnophiona (legless Amphibia) the yolk is situated at one pole and is large in quantity (figs. 153B-F; 154).
(5) Lastly, in a large portion of the vertebrate group, namely, in reptiles, birds, prototherian mammals, teleost and elasmobranch fishes, and in the marine lampreys, the deutoplasm is massive and the protoplasm which takes part in the early cleavages is small in comparison. In these eggs the yolk is never cleaved by the cleavage processes, and development of the embryo is confined to the animal pole cytoplasm (figs. 46, 47).
2) Formation of the Deutoplasm. The cytoplasm of the young oocyte is small in quantity, with a clear homogeneous texture (figs. 68A; 86A, B). As the oocyte develops, the cytoplasmic and nuclear volumes increase (fig. 68F), and the homogeneity of the cytoplasm is soon lost by the appearance of deutoplasmic substances (fig. 68G, H). In the oocyte of the frog, for example, lipid droplets begin to appear when the oocyte is about 50 /x in diameter (fig. 72A). (See Brachet, ’50, p. 53.) A little later glycogen makes its appearance, and finally yolk protein arises.
The origin of fat droplets and yolk spherules has been ascribed variously to the activities of chondriosomes (mitochondria and other similar bodies), Golgi substance, and of certain vacuoles. Most observers place emphasis upon the presence of a so-called “yolk nucleus†or “yolk-attraction sphere†situated near the nucleus of many oocytes as a structure associated with fat and yolk formation. In general, two types of yolk bodies have been described. One is the yolk nucleus of Balbiani and the other the mitochondrial yolk body of Brambell. The yolk nucleus of Balbiani (fig. 86A, B) consists of the following:
- a central body, the centrosphere or archoplasmic sphere within which one or more centriole-like bodies are found, and
- surrounding this central body, a layer of Golgi substances and chondriosomes (i.e., mitochondria, etc.).
This cytoplasmic structure probably is related to the idiosome of the oogonia (fig. 68A).
The formation of the deutoplasm, according to the theory associated with the Balbiani type of yolk nucleus is as follows: The surrounding pallial layer of Golgi substance and mitochondria moves away from the central portion (i.e., away from the centrosphere) of the yolk nucleus and becomes scattered and dispersed as small fragments within the cytosome (fig. 68F, G). The yolk nucleus as an entity thus disappears, and its fragments become immersed within the substance of the cytoplasm. Coincident with this dispersion of yolk nuclear material, rapid formation of small yolk spherules and fat droplets occur (fig. 68H). It appears thus that the formation of the deutoplasm composed of fat droplets and yolk spherules is directly related to the activities of the Golgi substance and chondriosomes.
Fig. 89. (A) Cytoplasm of oocyte, showing formation of a second kind of yolk (the M-C-yolk) in a vacuole surrounding the M-yolk sphere. (After Brambell, ’25.) (B)
Passing of Golgi substance from the follicle cells into the ooplasm of developing oocyte of the fowl. (After Brambell, '25.)
Fig. 90. Diagrams showing contrasting theories explaining the organization of polarity
of the cytoplasm of the fully developing egg or oocyte. Diagram at left shows polarity
explained according to quantitative differences, while the diagram to the right shows
qualitative differences. A = animal pole; V = vegetal pole. E represents a substance or
a factor, while EN-1, EN-2, etc., represent different quantities of substance E distributed
from pole to pole. SEC, SEN and SM are different chemical substances assumed to be
responsible for the determination of the ectoderm, entoderm, and mesoderm of the developing embryo. (After Barth: Embryology, New York, Dryden Press.)
On the other hand, the interpretation and description of the yolk body and its subsequent activities given by Brambell (’25) present a different view. According to the latter author, the yolk body is composed entirely of mitochondria; the Golgi substance and centrosphere are absent. Yolk formation proceeds as follows: As the young oocyte grows, the mitochondria increase in number and form the mitochondrial cloud (fig. 87A, B). The transitory mitochondrial yolk body differentiates within this cloud (fig. 87C). The mitochondrial yolk body ultimately breaks up into a mass of mitochondria, and the latter becomes dispersed in the cytoplasm of the oocyte (figs. 68F, G; 87D). Some of these dispersed mitochondria transform directly into yolk spheres (figs. 68H, 88, 89). Following this, another kind of yolk is formed in vacuoles surrounding these original yolk spheres (figs. 68H, 89 A, yolk spheres plus vacuoles). The fat droplets (C-yolk) within the ooplasm are formed according to Brambell “possibly under the influence of Golgi elements†(fig. 68H, fat droplets). Relative to the function of the yolk nucleus and its mitochondria, Brachet (’50), p. 57, considers it significant at the beginning, but its real importance is still to be understood.
The relationship, if any, of the oocyte nucleus to the deposition of yolk materials is not apparent. One must not overlook the real probability that the germinal vesicle (i.e,, the enlarged nucleus of the oocyte) may be related to the increase and growth of the cytoplasm and to yolk formation, for it is at this time that the chromatin threads surrender their normal diplotene appearance and become diffusely placed in the germinal vesicle. They also lose much of their basic chromatin-staining affinities while the Feulgen reaction is diminished (Brachet, ’50, p. 63). With regard to the possible function of the germinal vesicle in yolk synthesis, the following quotation is taken from a publication by Brachet (’47):
It is well worth pointing out that Puspiva (1942), using a very delicate and precise technique, found no correlation between the dipeptidase content of the nucleus and the onset of vitellus synthesis: such a correlation exists, however, in the case of the cytoplasm where dipcptidase increases markedly when the first yolk granules make their appearance. These results suggest that there is not evidence that the nucleus is the site of an especially active metabolism; cytoplasmic dipeptidase probably plays a part in yolk protein synthesis; if the nucleus controls such a synthesis, it works in a very delicate and still unknown way.
However, the means by which protein synthesis is effected still is a problem which awaits explanation (Northrop, ’50). (The interested student should consult Brachet, ’50, Chap. Ill, for a detailed discussion of the cytochemistry of yolk formation.)
Another aspect of the problem of cytoplasmic growth and differentiation of the oocyte presents itself for further study. Brambell (’25) concluded from his observations that Golgi substance passes from the follicle cells into the ooplasm of the growing bird oocyte and contributes to the substance of the peripheral layer (fig. 89B). Palade and Claude (’49) suggest that at least some of the Golgi substance be identified as myelin figures which develop “at the expense of lipid inclusions.†Thus it may be that the Golgi substance which Brambell observed (fig. 89B) passing from the follicle cells to the oocyte represents lipid substance. In the growing oocyte of the rat, Leblond (’50) demonstrated the presence of small amounts of polysaccharides in the cytoplasm of the oocyte, while the surrounding zona pellucida and follicle cells contained considerable quantities. These considerations suggest that the blood stream using the surrounding follicle cells as an intermediary may contribute food materials of a complex nature to the growing cytoplasm of the oocyte.
The localization of the yolk toward one pole of the egg is one of the movements which occurs during fertilization in many teleost fishes. In these forms, the deutoplasmic materials are laid down centrally in the egg during oogenesis, but move poleward at fertilization (fig. 122). A similar phenomenon occurs also during fertilization in Amphioxus and Styela among the protochordates. In many other fishes and in the amphibia, reptiles, birds, and monotrematous mammals, the yolk becomes deposited or polarized toward one pole of the oocyte during the later stages of oocyte formation, as the cytoplasm and the germinal vesicle move toward the other pole (figs. 68H, 72F). The polarization of the deutoplasmic substances thus is a general feature of the organization of the chordate egg.
3) Invisible Morphogenetic Organization Within the Cytoplasm of the Egg.
Two general categories of substances are developed within the cytoplasm of the oocyte during its development within the ovary, viz.:
(1) the visible or formed cytoplasmic inclusions, and
(2) an invisible morphogenetic ground substance.
The former group comprises the yolk spherules, fats, and other visible, often pigmented bodies which can be seen with the naked eye or by means of the microscope. The morphogenetic ground substance probably is composed of enzymes, hormones, and various nucleocytoplasmic derivatives enmeshed within the living cytoplasm. However, although we may assume that the basic, morphogenetic ground substance is composed of enzymes, hormones, etc., the exact nature of the basic substance or its precise relationship to the various formed inclusions of the cytoplasm is quite unknown (see Fankhauser, ’48, for discussion). More recent experiments demonstrate that the yolk or deutoplasmic material not only serves as a reservoir of energy for embryonic development but also is in some way connected with the essential, basic organization of the egg.
Although we know little concerning the exact nature of the morphogenetic organization of the egg or how it forms, studies of embryological development force upon us but one conclusion, to wit, that, during the period when the oocyte develops in the ovary, basic conditions are elaborated from which the future individual arises (Fankhauser, ’48). Within the cytoplasm of the mature egg of many chordates, this inherent organization is revealed at the time of fertilization by the appearance of definite areas of presumptive organ-forming substances. For example, in the egg of the frog and other amphibia, the yolk pole is the stuff from which the future entodermal structures take their origin; the darkly pigmented animal or nuclear pole will eventually give origin to epidermal and neural tissues; and from the zone between these two areas mesodermal and notochordal tissues will arise (fig. 119K). Similar major organ-forming areas in the recently fertilized egg have been demonstrated in other chordates, as in the ascidian, St ye la, and in the cephalochordate, Amphioxus. In the eggs of reptiles, birds, and teleost and elasmobranch fishes, while the relationship to the yolk is somewhat different, major organ-forming areas of a similar character have been demonstrated at a later period of development (Chaps. 6-9). This suggests that these eggs also possess a fundamental organization similar, although not identical, to that in the amphibian egg.
4) Polarity of the Egg and Its Relation to Body Organization and Bilateral Symmetry of the Mature Egg. One of the characteristic features of the terminal phase of egg differentiation in the chordate group is the migration of the gerrninal vesicle toward the animal pole of the egg (figs. 72F, 119A). As stated aboveTTn many vertebrate eggs the deutoplasmic jnaterial becomes situated at the opposite pole, known as the vegetal (vegetative) or yolk pole, either before fertilizatloiToTsEortty after. The relatively yolk-free protoplasm aggregates at the animal pole. Consequently the maturation divisions of the egg occur at this pole (fig. 1 19 A, B, D). The formation of a definite polarity of the egg, therefore, is one of the main results of the differentiation of the oocyte.
Various theories have been suggested in an endeavor to explain polarity in the fully developed egg or oocyte. All these theories emphasize qualitative and quantitative differences in the cytoplasmic substances extending from one pole of the egg to the other (fig. 90).
The animal and vegetal poles of the egg have a definite relationship to the organization of the chordate embryo. In Amphioxus, the animal pole becomes the ventro-anterior part of the embryo, while in the frog the animal pole area becomes the cephalic end of the future tadpole, and the yolk pole comes to occupy the posterior aspect. In teleost and elasmobranch fishes the yolk-laden pole lies in the future ventral aspect of the embryo, and it occupies a similar position in the reptile, bird, and prototherian mammal (see fig. 215). Studies have shown that the early auxiliary or trophoblastic cells in eutherian mammals lie on the ventral aspect of the future embryo. Consequently, it is to be observed that the various substances in mature vertebrate and protochordate eggs tend to assume a polarized relationship to the future embrydnic axis and body organization.
Many vertebrate and protochordate eggs possess a bilateral symmetry
which becomes evident when the fertilization processes are under way or
shortly after their conclusion. The appearance of the gray crescent in the
frog’s egg (fig. 119K) and in other amphibian eggs during fertilization and
the similar appearance of the yellow crescent in the fertilized egg of the
ascidian, Styela (fig. 132D) serve to orient the future right and left halves
of the embryo. Conditions similar to that of Styela, but lacking the yellow
pigment, are present in Amphioxus. Similarly, in the chick, if one holds the
blunt end of the egg to the left, and the pointed end to the right, the early
embryo appears most often at right angles, or nearly so, to the axis extending
from the broader to the smaller end of the egg, and in the majority of cases
the cephalic end of the embryo will appear toward the side away from the
body of the observer. There is some evidence that the “yolk†or egg proper
is slightly elongated in this axis. It appears, therefore, that the general plane
of bilateral symmetry is well established in the early chick blastoderm, although
the early cleavages do not occur in a manner to indicate or coincide with
this plane. In prototherian mammals, a bilateral symmetry and an anteroposterior orientation is established in the germinal disc at the time of fertilization, soon after the second polar body is discharged (fig. 136).
5) Membranes Developed in Relation to the Oocyte; Their Possible Sources of Origin. A series of membranes associated with the surface of the oocyte are formed during its development within the ovary. Three general types of such membranes are elaborated which separate from the oocyte’s surface at or before fertilization, leaving a perivitelline space between the egg’s surface and the membrane. They are:
( 1 ) A true vitelline membrane which probably represents a specialization or product of the ooplasmic surface. For a time this membrane adheres closely to the outer boundary of the ooplasm, but at fertilization it separates from the surface as a distinct membrane.
(2) A second membrane in certain chordates is elaborated by the follicle cells. It is known as a chorion in lower Chordata but is called the zona pellucida in mammals.
(3) A zona radiata or a thickened, rather complex, membrane is formed in many vertebrates; it may be considered to be a product of the ooplasm or of the ooplasm and the surrounding follicle cells.
All of the above membranes serve to enclose the egg during the early phases of embryonic development and therefore may be considered as primary embryonic membranes. As such, they should be regarded as a definite part of the egg and of the egg’s differentiation in the ovary. A description of these membranes in relation to the egg and possible source of their origin in the various chordate groups is given below.
a) Chorion in Styela. A previously held view maintained that the chorion
Fig. 91. Formation of the chorion in the egg of Styela. (A) Chorion is shown along
the inner aspect of the follicular epithelium. The test cells lie in indentations of the
peripheral ooplasm. (B) Optical section of an ovulated egg. (Redrawn and modified
from Tucker, ’42.)
Fig. 92. Developing vitelline membranes of Scyllium canicula. Observe that two membranes are present in the young egg; later these membranes fuse into one membrane. (A) Surface area of young oocyte with a vitelline membrane and zona radiata. (B) Slightly older oocyte with the radiate zone not as prominent. (C) Older oocyte with a single, relatively thick, vitelline membrane. (D) Nearly mature oocyte with a thin vitelline membrane. (After Balfour, Plate 25, The Works of Francis Maitland Balfour, ed. by Foster and Sedgwick, London, Macmillan, 1885.)
and “test†cells of the egg of Styela were ejected from the surface cytoplasm
at the time of ovulation (Conklin, ’05). A recent view, however, maintains
that the test cells arise from follicle cells and come to lie in indentations of
the periphery of the egg outside of the thin vitelline membrane (Tucker, ’42).
The chorion is formed by the inner layer of follicle cells and comes to lie
between the test cells and the inner layer of follicle cells in the mature egg
(fig. 91A). At ovulation the chorion moves away from the surface of the
oocyte. At this time also, the test cells move outward from their indentations
in the peripheral ooplasm and come to lie in the perivitelline space between
the egg surface and the chorion (fig. 9 IB). An ooplasmic membrane which represents the thin surface layer of ooplasm is present. However, it does not
separate from the periphery of the egg at fertilization. During its early development, the embryo remains within this chorionic shell. The chorion thus
represents the primary embryonic membrane of this species.
Fig. 93. Vitelline membranes of certain teleost fishes. (After Eigenmann, 1890.) (A)
Pygosteus pungtius. Radial section through micropyle of egg about 0.4 mm. in diameter. (B) Radial section through micropyle of egg of Perea, the perch. (C) Vitelline membranes of Fundulus heteroclitus about 0.8 mm. in diameter.
b) Egg Membranes of Amphioxus. Two surface membranes are formed
and eventually separate from the egg of Amphioxus. The outer vitelline
membrane is elaborated on the surface of the egg and remains in contact
with this surface until about the time of the first maturation division. It then
begins to separate from the egg’s surface. (See Chap. 5.) After the sperm
enters and the second maturation division occurs, a second, rather thick,
vitelline membrane also separates from the egg. The first and second vitelline
membranes then fuse together and become greatly expanded to form the primary embryonic membrane. (See Chap. 5.) A thin ooplasmic membrane
remains at the egg’s surface.
c) Vitelline Membrane and Zona Radiata of Elasmobranch Fishes. In the egg of the shark, Scy Ilium canicula, two egg membranes are formed, an outer and an inner membrane. The outer membrane is a homogeneous vitelline membrane, while the membrane which comes to lie beneath this outer membrane has a radiate appearance and hence may be called a zona radiata. This latter membrane soon loses its radiate appearance and becomes a thin membrane along the inner aspect of the vitelline membrane (fig. 92 A, B). In the mature egg both of these membranes form a thin, composite, vitelline membrane (fig. 92C, D). At about the time of fertilization the latter membrane separates from the egg’s surface; a perivitelline space then lies between these structures and the surface ooplasm of the egg.
d) Zona Radiata of Teleost Fishes. The surface ooplasm in teleost fishes gives origin to a membrane which in many cases has a radiate appearance. In some species this membrane appears to be composed of two layers. This radiate membrane which forms at the surface of the egg of teleost fishes appears to be the product of the ooplasm, and, therefore, should be regarded as a true vitelline membrane. In the perch a true chorion also is formed as a gelatinous or filamentous layer produced external to the radiate membrane by the follicle cells (fig. 93B). In Fundulus heteroclitus there are apparently three distinct parts to the membrane which surrounds the ooplasm of the egg:
( 1 ) a zona radiata,
(2) a thin structureless membrane external to the zona, and,
(3) the filamentous layer whose filaments are joined to the thin membrane around the zona (fig. 93C).
These three layers are probably derived from the ooplasm of the egg (Eigenmann, ’90). Consequently, the filamentous chorion or gelatinous layer, if derived from the egg itself, is not a true chorion in this particular egg.
Fig. 94. Vitelline membrane of an almost mature egg of the frog.
Fig. 95. Zona radiata (zona pellucida) or vitelline membrane of Chrysemys picta. (After Thing, ’18.)
Fig. 96. Zona radiata of the egg of the fowl. (After Brambell, ’25.)
At one end of the forming egg, a follicle cell sends an enlarged pseudopodiumlike process inward to the surface of the egg. As a result of this enlarged extension of the follicle cell to the ooplasmic surface, an enlarged pore-like opening in the zona radiata is formed. This opening persists as the micropyle after the egg leaves the ovary (fig. 93A).
As the teleost egg is spawned, the chorionic layer hardens when it comes in contact with the water. If fertilization occurs, the surface of the egg emits a fluid and shrinks inward from the zona radiata. In this manner, a perivitelline space is formed between the egg, and the zona is filled with a fluid. The egg is thus free to revolve inside of the zona (Chap. 5).
e) Vitelline Membrane (Zona Radiata) in Amphibia. In the amphibia, a vitelline membrane is formed probably by the surface ooplasm, although there may be contributions by the follicle cells of the ovary (Noble, ’31, p. 281). This membrane separates from the egg at the time of fertilization, forming a perivitelline space (fig. 94). The latter space is filled with fluid. Later the vitelline membrane expands greatly to accommodate the developing embryo. A delicate surface layer or membrane forms the outer portion of the ooplasm below the vitelline membrane. In some amphibia the vitelline membrane may have a radiate appearance.
f) Zona Radiata (Zona Pellucida) of the Reptile Oocyte. In the turtle group, the development of the zona radiata (pellucida) appears to be the product of the follicle cells (Thing, ’18). Filamentous prolongations of the follicle cells extend to the surface ooplasm of the developing egg (fig. 95). A homogeneous substance produced by the follicle cells then fills the spaces between these prolongations. The filamentous extensions of the follicle cells in this way produce a radiating system of canals passing through the homogeneous substance; hence the name, zona radiata. Bhattacharya describes Golgi substance as passing from the follicle cells through the canals of the zona radiata into the egg’s ooplasm in the developing eggs of Testudo graeca and Uromastix hardwicki. (See Brambell, ’25, p. 147.)
In contradistinction to the above interpretation, Retzius (’12) describes the homogeneous substance which forms the zona radiata of the lizard, Lacerta viridis, as originating from the ooplasm of the egg.
g) Vitelline Membrane (Zona Radiata) of the Hen’s Egg. The vitelline membrane, as in the turtle groups, appears to form about the young oocyte as a result of contributions from the surrounding follicle cells although the superficial ooplasm of the oocyte may contribute some substance. This occurs before the rapid deposition of yolk within the developing oocyte. It is probable that the follicle cells send small pseudopodium-like strands of cytoplasm through the numerous perforations of the very thin vitelline membrane around the oocyte’s surface into the superficial ooplasm in a similar manner to that which occurs in reptiles. The vitelline membrane (zona radiata) thus assumes a radiate appearance as it increases in thickness (figs. 47, 96).
Fig. 97. Kinoplasmic bead or droplet upon the middle piece of mammalian sperm.
(A) Pig sperm. (After Retzius, Biol. Untersuchungen, New Serfes, 10; Stockholm: Jena.)
(B) Cat sperm. (After Retzius, Biol. Untersuchungen, New Series, 10; Stockholm: Jena.) (C-D) Dog sperm. (C) Upper part of epididymis. (D) Lower or caudal part of epididymis.
When the vitelline membrane thickens, the loci where the cytoplasmic strands from the follicle cells pass through the membrane become little canals or canaliculi. As the oocyte increases in size, a thin space forms between the vitelline membrane or zona radiata and the follicle cells; it is filled with fluid and forms the follicular space. The egg now is free to rotate within the follicle. In consequence, the pole of the egg containing the blastodisc always appears uppermost. Due to the increasing pendency of the egg follicle as the egg matures, the blastodisc comes to rest, a short while previous to ovulation, at the base of the pedicle where the blood vessels are most abundant (fig. 47B). During the latter phases of oocyte development, the vitelline membrane constitutes an osmotic membrane through which all nourishment must pass to the oocyte, particularly in its later stages of growth. Tlie surface ooplasm forms a delicate surface membrane beneath the zona radiata.
h) Membranes of the Mammalian Oocyte. All mammalian oocytes possess a membrane known as the zona pellucida. It is a homogeneous layer interposed between the ooplasm and the follicle cells. By some investigators it is regarded as a product of the oocyte, while others regard it as a contribution of the ooplasm and follicle cells. The majority opinion, however, derives the zona pellucida from the follicle cells. In addition to the zona pellucida, the oocyte of the prototherian mammals has a striate layer lying close to the surface of the oocyte. This striated layer probably is derived from the surface ooplasm. This membrane later disappears, and a perivitelline space occupies the general area between the surface of the oocyte and the zona pellucida (fig. 46; Chap. 5). The zona pellucida separates from the egg surface after sperm contact.
5. Physiological Maturation of the Gametes a. Physiological Differentiation of the Sperm
Added to the nuclear and cytoplasmic transformations of the sperm described above, a further process of sperm ripening or maturing appears to be necessary. In the mammal, for example, the sperm cell must pass through the epididymis to achieve the ability to fertilize the egg. This is shown by the fact that sperm taken from the seminiferous tubules will not fertilize, although, morphologically, two sperm, one from the testis and one from the epididymis cannot be distinguished other than by the presence in some mammals of the so-called “kinoplasmic droplet†(figs. 82D, 97). These droplets do not appear in great numbers upon ejaculated sperm but are found on sperm, particularly in epididymides. It is possible that these droplets may arise from a secretion from the epididymal cells (CoIIery, ’44). In the dog, these droplets are attached to the neck of the sperm in the caput epididymidis but are found at the posterior end of the middle piece of the sperm in the cauda epididymidis and vas deferens and are probably lost at the time of ejaculation (Collery, ’44). Investigators differ greatly in interpreting the significance of this body. However, these droplets do seem in some way to be directly or indirectly concerned with the physiological maturing of the sperm. In this connection Collery (’44) notes that sperm are probably motile on leaving the seminiferous tubules, but active forward movement is not seen until the bead has reached the junction of middle piece and tail.
In the fowl, Domm (’30, p. 318) suggests the probability that the sperm may undergo an aging or ripening process essential for reproduction somewhere in the reproductive system other than the seminiferous tubules. The work of Lipsett quoted in Humphrey (’45) suggests that the accessory reproductive system also is necessary for a ripening process of the sperm in urodele amphibia.
On the other hand, in the frog, sperm taken from the testis have the ability to fertilize eggs. In this case, the sperm probably undergo a physiological ripening in the testis along with morphological differentiation.
The foregoing considerations suggest that a physiological maturation of the sperm is necessary to enable the sperm to take part in the fertilization process.
b. Physiological Ripening of the Female Gamete
The physiological maturing of the oocyte is linked to factors which influence the developing egg at about the time the maturation divisions occur. Sea-urchin sperm may penetrate the egg before the maturation divisions occur (Chap. 5). However, development does not take place in such instances. On the other hand, sperm entrance after both maturation divisions are completed initiates normal development. In the protochordate, Styela, marked cortical changes transpire at about the time the egg leaves the ovary, and as it reaches the sea water, the germinal vesicle begins to break down. The oocyte becomes fertilizable at about this time. In Amphioxus, although the first polar body is given off within the adult body, the egg apparently is not fertilizable until it reaches the external salt-water environment. The secondary oocyte of the frog presumably must remain within the uterus for a time to ripen in order that ensuing development may be normal. These and other instances suggest that physiological changes — changes which are imperative for the normal development of the egg — are effected at about the time that the maturation divisions occur.
D. Summary of Egg and Sperm Development
From the foregoing it may be seen that the development of the gametes in either sex involves a process of maturation. This maturation entails changes in the structure and constitution of the nucleus and cytoplasm, and, further, a functional or physiological ripening must occur. The comparative maturation events in the egg and sperm may be summarized as follows:
Egg ( in Oogenesis)
Sperm (in Spermatogenesis)
1. Nuclear maturation
a. Homologous chromosomes synapse and undergo profound changes during which parts of homologous chromosomes may be interchanged; ultimately, the chromosome number is reduced to the haploid number
b. Nucleus enlarges, and contained nuclear fluid increases greatly; ultimately the nuclear fluid is contributed to cytoplasm upon germinal vesicle break down
c. Nuclear maturation occurs simultaneously with cytoplasmic differentiation
1. Nuclear maturation
a. (Similar to the female)
b. Nucleus remains relatively small and
enlargement is slight; nuclear fluid
small in amount; during spermiogenesis the nucleus may contract into
a compact mass; considerable elongation of nucleus occurs in many
species
c. Nuclear maturation occurs before spermiogenesis or cytoplasmic differentiation
SUMMARY OF EGG AND SPERM DEVELOPMENT
Egg ( in Oogenesis)
Sperm (in Spermatogenesis)
2. Cytoplasmic maturation
This involves:
2. Cytoplasmic maturation
This involves:
a. Polarization of cytoplasmic materials and the nucleus in relation to the future maturation phenomena; the nucleus becomes displaced toward one pole, the animal pole, and the yolk, and other cytoplasmic materials; in many eggs becomes displaced toward the opposite or vegetal pole
a. Polarization of nucleus and cytoplasmic materials along an elongated antero-posterior axis, with the head, neck, middle piece, and tail occupying specific regions along this axis. The nucleus occupies a considerable portion of the anterior region or head
b. Formation of deutoplasm or stored food material, varying greatly in amount in different animal species. The deutoplasm is composed of fats, carbohydrates, and protein substances
b. Little food substances stored within cytoplasm; food reserve in seminal fluid
c. Cytoplasm increased in amount; formation of basic organ-forming areas or cytoplasmic stuffs from which the future embryo arises
c. Discarding of a considerable amount of cytoplasm, some Golgi elements and mitochondria. Retention of some Golgi elements, centrioles, mitochondria, etc.
d. Formation of primary embryonic membranes
d. No specific membranes formed around sperm, although elaborate membranes for motile purposes are formed in some sperm
3. Physiological maturation or the development of a fertilizable stage
This involves:
a. Formation of an organization which when stimulated by external influences initiates and carries on the processes necessary for normal embryonic development
3. Physiological maturation or the development of the ability to contact and fertilize the egg
This involves:
a. Development of an organization which, when stimulated by proper external substances, responds by a directed movement resulting in locomotion; also capable of being attracted by egg substances
b. Acquisition of ability to enter into a developmental union with a sperm
b. Acquisition of ability to fertilize, i.e., to enter into a developmental union with an egg or oocyte
c. Development of ability to form and secrete gynogamic substances which aid in the fertilization process. (See Chap. 5)
c. Acquisition of ability to produce and secrete androgamic substances which aid in the fertilization process
d. Assumption of an inhibited or dormant condition during which metabolic processes proceed slowly in anticipation of the fertilization event
d. Assumption of an active metabolic state
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