AMER. ZOOL, 16:315-343 (1976).
Comparative Spiralian Oogenesis—Structural Aspects:
An Overview
ERWIN HUEBNER
Department of Zoology, University of Manitoba, Winnipeg, Manitoba R3T 2N2, Canada
AND
EVERETT ANDERSON
Department of Anatomy and Laboratory of Human Reproduction and Reproductive
Biology, Harvard Medical School, Boston, Massachusetts 02115
SYNOPSIS Considerable variety exists in ovarian structure and cellular interaction in spiralians. During their development the eggs of Dwpaha cuprea, are associated with nurse cells;
there are no follicle cells in this species. The nurse cells have prominent nuclei and connect
to oocytes via cytoplasmic bridges through which ribosomes, mitochondria and other inclusions pass. More commonly, follicle cells surround a portion of, or entire, oocytes in many
species of spiralians. Usually, as in Ilyanassa, they have a well developed compliment of
organelles. The structure and distribution of organelles within follicle cells implies that they
are functionally active, but precisely in what manner during oogenesis is poorly understood.
Other cell types, such as Leydig and interstitial cells also seem to play a role in oogenesis.
Within the oocyte, a host of components including yolk, lipid, mitochondria, ribosomes,
membranous cisternae, cortical granules, etc. are accumulated. Autosynthetic yolk formation is prevalent among spiralians. Surface differentiation includes microvillar development. This may be uniform in some eggs or restricted to certain regions (e.g., the animal
hemisphere) in other oocytes. Oocyte-follicle cell interactions change during oogenesis. The
topographical association of the oocyte with other ovarian cells influences subsequent
animal-vegetal polarity and other ooplasmic differences. Examples of ooplasmic localizations are discussed. Conventional EM has revealed no unusual cortical structure in many
oocytes although occasionally microtubules and microfilaments are present.
INTRODUCTION
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Since Wilham Harvey s philosophical
statement Omne vivum ex ovo oogenesis
and development have attracted the attenuon of cell biologists from many disc.phnes.
That embryogenesis begins in oogenesis
was suggested by Wilson in 1896 (cited m
Wilson, 1927) and clearly md.cates the significance of the egg. The early studies of
Driesch with sea urchins and Wilson (cf.
1927) with various molluscs and annelids
i n d i c a t e d t h e existence of two categories of
and mosaic
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exhibit a spiral cleavage patm o s a k
t e r n a n d « o v i d e a c o n v e n i e n t category/the
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diverse groups as turbellarians, nemer~
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tines, annelids and molluscs. The spiral
r
We would like to express our thanks to Dr. G. T.
'
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Taylor, Dept. Physiology, Southern Illinois University cleavage pattern, although more or less
at Carbondale, for providing some micrographs of modified in some members, is t h e primary
Ilyanassa and for helpful discussion.
pattern in prostomous bilateral inverteThis report was supported by grants from the Na- brates (Cathers, 1971). Bover (1971) noted
tional Research Council of Canada and Research
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Board of the University of Manitoba to E. H. and t h a t although spiral cleavage and mosaigrants from the National Institutes of Health, United Cism seem to go hand-in-hand, data On the
States Public Health Service to E. A.
acoel Childia has shown a high degree of
315
316
ERWIN HUEBNER AND EVERETT ANDERSON
regulation. This suggests that mosaicism
is highly specialized and a derivative of indeterminate cleavage.
The arrangement of the spiral cleavage
pattern provides unique morphological
markers for following development, but
the primary interest stems from the fact
that the developmental potency of each
blastomere is restricted early and each
eventually gives rise to a definite portion of
the organism (Wilson, 1927; Costello, 1945,
1948; Clement, 1952, 1956, 1968, 1971;
Davidson, 1968; Cathers, 1971). Numerous
investigators support the notion that this
developmental potency is predelineated
during oogenesis (see Wilson, 1927; Raven,
1961; Davidson, 1968; Cathers, 1971; Reverberi, 1971 a,b,c). The consequence of
this predelineation is that there are relationships between the nature and organization of the ooplasmic constituents and the
developmental capacities. As stated by
Davidson and Hough (1972), "The molecular events underlying the establishment of
cytoplasmic localization patterns remain
completely unknown." Thus a clearer understanding of the events of oogenesis both
from ultrastructural and molecular points
of view is an essential prerequisite to
comprehend the relationship between oocyte organization and development.
Ideally one hopes to link the structures
accumulated during oogenesis with their
subsequent roles in embryogenesis. With
this perspective in mind we have chosen to
examine oogenesis in the following framework: ovarian structure, relationships of
other cell types to the oocyte, events of oocyte differentiation, evidence of ooplasmic
localization, and envelope formation. We
have also included a brief look at the fate of
some oocyte structures during early development. With the volume of literature
and limitation of space, we will of necessity
F1G. 1. Section through the wall of the ovary of the
chiton Mopalia muscosa depicting oocytes (II, III, IV) in
various stages of development (OV, ovarian
epithelium; IT, interstitial tissue) x 100. (from Anderson, 1969)
FIG. 2. Light micrograph of the ovary of Ilyanassa
obsoleta illustrating previtellogenic (PV) and vitellogenic oocytes (VC). Note the prominent oocyte nuclei or germinal vesicles with distinct nudeoli. Also
present are thin follicle cells (F) and the interstitial or
be restricted to selected examples and will
include those primarily in the molluscs and
annelids. The review of the literature is not
inclusive and for general reviews on oogenesis the reader is referred to Raven (1961),
Srivastava (1965), Stegner (1967), N0rrevang (1968), Cathers (1971), Reverberi
(1971), Biggers and Schuetz (1972) and
Longo and Anderson (1974).
MATERIALS AND METHODS
The procedures are essentially those
previously reported in Anderson and
Huebner (1968), Anderson (1969), and
Taylor and Anderson (1969). Previously
unpublished data included on Diopatra
cuprea were prepared as cited in Anderson
and Huebner (1968). The data on the
nudibranch Cratenapilata are from material
fixed in a modified Karnovsky's fixative
(Huebner and Anderson, 1972) with sea
water being used instead of cacodylate buffer and gluteraldehyde neutralized prior to
use.
OVARIAN STRUCTURE
Not unexpectedly ovarian structure of
spiralia covers a diversity ranging from solitary egg development in certain acoels
(Raven, 1961; Boyer, 1972) to distinct and
varied organs with various cell types associated with oocyte development. The latter is referred to as alimentary oocyte development (Wilson, 1927; Raven, 1961).
The ovary is a highly compartmentalized
morphogenetic system.
The ovary of the rotifer Asplanchna has a
solid structure consisting of a syncytial association of vitellaria cells with oocytes and
a partially enveloping syncytial follicular
tissue (Bentfield, 1971a, 19716). The elongate solid ovaries of nematodes have ooLeydig cells (LC). x 250. (Micrograph courtesy of G.
Taylor)
FIGS. 3 and 4. These toliiidine blue stained epon
sections of the gonadsof the nudibranch Cratenapilata
illustrate various stages of oocyte growth (I, II, III, IV,
V). Note the prominent germinal vesicle and dark
nucleolus, amphinucleolus (*); follicle cell (F); and
sperm in an adjacent tubule (S). X600, x800, inset
X800.
COMPARATIVE SPIRALIAN OOGENESIS—STRUCTURE
SVf
318
ERWIN HUEBNER AND EVERETT ANDERSON
cytes circumferentially associated with the
central common rachis by means of intercellular bridges (Foor, 1967; McLaren,
1973). In some molluscs there is a relatively
simple structure appearing as a single saclike ovary in amphineurans as illustrated in
Figure 1 (see also Anderson, 1969; Richter
and Gotting, 1974) or there are complex
highly branched tubular gonads as in the
ovary of Ilyanassa (Figure 2; see also
Coggeshall, 1970; Hess, 1971; Starke,
1971; Luchtel, \912a,b). In those species
that are hermaphroditic, both sperm and
eggs may develop in the same acini as in
Limnaea (Raven, 1961; Joosse and Reitz,
1969) and Planorbis (Starke, 1971) or they
may be localized in separate but adjacent
acini as noted in the nudibranch Cratena
(Figs. 3, 4). In some polychaetes the ovaries
are fairly non-defined and irregular (Andrews, 1891; Anderson and Huebner,
1968) while in others they are well defined
(Potswald, 1967; Olive, 1971). Olive (1971)
noted the array of gonad morphologies and
locations in 14 families of polychaetes and
described the histological details of the
gonads of Cirratulus which extend posteriorly into the coelom from each septum. A
similar morphology exists in the polychaete
Diopatra where loops of cells extend into
and eventually break off into the coelom
(Andrews, 1891). For further examples
and discussion of gonad structure the
reader may consult Raven (1961),
Harrison-Matthews (1962), Joosse and
Reitz (1969), and Olive (1971). The report
of Luchtel (1972a, 1972ft) on pulmonate
molluscs provides an excellent example of
gonad morphogenesis and how the male
and female compartments of the hermaphroditic gonad became isolated.
In the case of solitary egg production
very little cell association (with no nurse
cells) exists and only rarely are follicle cells
found (Boyer, 1972). In most of the examples noted above however, the oocyte differentiates in intimate association with
other cell types of the ovary. It is known
from the studies of Raven (1970) that much
of the pattern of early development is
somehow determined by the underlying
pattern in the egg and this pattern in turn is
determined or influenced by cellular inter-
actions during oogenesis. Thus, our attention is drawn to cellular interactions existing within the ovary.
CELLULAR INTERACTIONS
Nurse cells
Although a variety of terms has been
applied to cells which are associated with
oocytes by confluent cytoplasmic bridges
we shall consider them collectively as nurse
cells. This term has been widely used to
define such cells in the meroistic ovaries of
insects. In insects nurse cell-oocyte interaction is highly developed (Davidson, 1968;
Huebner and Anderson, 1972). The prevalence of such associations is more restricted
in various spiralians (see Wilson, 1927).
Many of the classical descriptions of nurse
cells that have been reported (cf. Wilson,
1927; Raven, 1961) require re-examination
in order to determine whether or not a true
syncytial relationship exists.
Actual syncytial associations consist of a
compartmentalized system presumed to
have arisen from numerous incomplete cell
divisions (see Anderson and Huebner,
1968; King, 1970).
In the annelids Enchytreus (Dumont,
1969) and Eisenia (Chapron and Relexans,
1971) and the archiannelid Dinophilus
(Grun, 1972) the various compartments of
this syncytium develop as a group of oocytes in synchrony presumably with intercellular exchange of ribosomes, mitochondria and other organelles. In other
cases, more reminiscent of the situation in
insects, only a few or even a single compartment of the syncytium differentiates
into an oocyte whereas the others develop
into nurse cells. We have no information on
what factors trigger a particular compartment to become the oocyte. In Drosophila
King (1970) speculates that the number of
bridges that enter certain compartments is
related to which cell will become the oocyte.
Apparently this is not so in Diopatra since all
compartments except the end ones have 2
bridges each. Although Bentfeld (1971a,
19716) did not describe the ontogeny of the
rotifer ovary, the tissue referred to as the
vitellaria is associated with the few develop-
COMPARATIVE SPIRALIAN OOGENESIS—STRUCTURE
FIGS. 5-9. Phase contrast micrographs illustrating the
development of strings of cells (eggs and nurse cells)
from the coelom of the polychaete Diopatra. Note the
enlarging oocyte (O) and attached nurse cells (N). The
oocyte has a prominent germinal vesicle and distinct
nucleolus. Figs. 5-8, X120; Fig. 9, X140.
FIG. 10. This phase-contrast micrograph illustrates
the stratification of organelles in a mature Diopatra
319
cuprea oocyte. Note the eccentric germinal vesicle
(GV): animal pole (A); and yolk containing vegetal
pole (V). The nurse cells are no longer attached, x 160.
FIG. 11. This micrograph of a living mature Nereis
limbata is included to contrast it with Figure 10. A
similar stratification of ooplasmic components is not
seen here. x280.
320
ERWIN HUEBNER AND EVERETT ANDERSON
COMPARATIVE SPIRALIAN OOGENESIS—STRUCTURE
321
ing oocytes by intercellular bridges and
hence, may be considered nurse cells.
In this communication we have selected
the polychaete Diopatracuprea as an illustrative example to outline the structure and
function of nurse cells (see Andrews, 1891;
Allen, 1961; Anderson and Huebner,
1968). Briefly, loops of cells protruding
from the septal ovary break off and float in
the coelom. Within this chain of cells one
(approximately the middle one) becomes
the oocyte and the rest nurse cells. As the
egg enlarges, the laterally attached chains
of nurse cells are displaced to one pole (the
vegetal pole) eventually detaching when
the oocyte is mature (Figures 5-10, 12, 13,
H).
Ultrastructural examination of the nurse
cells reveals that they possess a microvillous
surface (Fig. 16), presumably to facilitate
uptake of substances from the coelomic
fluids. Each cell has a prominant nucleus
with a pronounced bipartite nucleolus (Fig.
15). As seen in Figures 13 and 14, ribosomes occupy the intercellular bridges. The
structure of the nucleus and cytoplasm of
the nurse cell suggests that one of its major
functions is the production of ribosomal
RNA for the oocyte (Allen, 1961; Anderson
and Huebner, 1968). There is a structural
similarity between the granular areas one
sees in Diopatra nurse cell cytoplasm (Fig.
18) and the granular basophilic areas seen
in the amphineuran oocyte (Fig. 17) which
lacks nurse cells. The amphineuran oocyte
nucleus produces all of the RNA of the
gamete (Anderson, 1969). The nurse cell
cytoplasm also has numerous mitochondria, lipid spheres (Figs. 13, 15) and
pigment granules. All of these structures
are frequently seen in the bridges between
the nurse cells and oocyte. Even in a fairly
far removed taxonomic group such as the
rotifers, the vitellaria (here considered
equivalent to nurse cells) transport ribo-
somes, lipid and mitochondria to the oocyte
(Bentfeld, 1971a, 19716). Cortical granules
are also passed into the oocyte. Presumably,
as has been shown in the cecropia moth by
Woodruff and Telfer (1973), the flow of
material through the intercellular bridges
in other species is unidirectional.
If a relationship exists between nurse cell
products and subsequent embryonic development, it is unknown at present. We
can only assume that various types of RNA
are transported to the oocyte and utilized
during early embryogenesis. Certainly the
production of mitochondria for oocytes by
nurse cells has important implications for
embryonic development, particularly in
view of the localization of these organelles
in some oocytes. Despite such speculations
one clear aspect of the influence of the
nurse cells on localization in the oocytes can
be found in Diopatra. In this polychaete the
point of nurse cell attachment will become
the future vegetal pole of the egg.
FIG. 12. This light micrograph of an epon embedded
specimen shows a Diopatra oocyte (O) with its attached
nurse cells (N). A cytoplasmic bridge (CBO) is shown
connecting the oocyte to one of the chains of nurse
cells. X400. (from Anderson and Huebner, 1968)
FIG. 13. This electron micrograph and inset show a
portion of the nurse cell (N), oocyte (O), and cytoplasmic bridge (CBO) of Diopatra. Lipid droplets (L)
and mitochondria are present in the nurse cell and
numerous ribosomes in the intercellular bridge. The
border of the bridge possesses a local thickening (LT)
and the oocyte a prominent primary envelope (PE).
x 12,000. (from Anderson and Huebner, 1968)
FIG. 14. Present here is an intercellular bridge (CB)
between two nurse cells. Note the thickened border
(LT), dense areas (DA), and plasmalemma (PM).
x 30,000.
Follicle cells
Follicle cells are more frequently found
associated with oocytes than are nurse cells
(Figs. 1-4, 19, 20; see also Raven, 1961).
Follicular cells may encompass the oocyte
entirely (Anderson, 1969) or partially (Raven, 1961; Dumont, 1969; Taylor and Anderson, 1969). They are not in cytoplasmic
continuity with the oocyte, and in the
majority of cases, consist of distinct single
cells, although exceptions may be found
(Bentfeld, 1971a; Arnold and WilliamsArnold, 1976). There are examples of cells
enveloping or associated with oocytes referred to in the literature by various other
terms. For example, Newton (1970) describes an unusual cell termed the "accessory cell" which surrounds the oocyte of a
fresh-water turbellarian. Further elucidation of the structure and function of this
322
ERWIN HUEBNER AND EVERETT ANDERSON
cell and other enveloping cells is required
in order to determine if they can be included in the category designated by the
term "follicle cell."
As noted by Taylor and Anderson (1969)
follicle cells of molluscs usually constitute a
thin layer around the smallest oocytes close
to the tubular wall (Figs. 2, 3 and 4; see also
Raven, 1961; Bedford, 1966). Presumably
these develop from undifferentiated auxiliary cells (Starke, 1971; Luchtel, 1972). As
oocytes enlarge, they protrude into the
lumen thereby leaving only their basal area
in contact with follicle cells and the rest of
their surface associated either with other
oocytes or the free lumen. The oocytes of
the annelid Enchytraeus have follicle cells
only on their distaj surface (Dumont, 1969).
Consequently, in this species there are regional differences of membrane to membrane association around the circumference of the oocyte during its development.
As will be discussed below, animalvegetal polarity often corresponds to the
orientation of the oocyte within the ovary.
In 1lyanassa that portion of the oocyte adjacent to the follicular epithelium becomes
the yolk-laden vegetal pole (Taylor and
Anderson, 1969). The localization of subcortical areas in Limnaea has also been related to the follicle cell-oocyte interaction
(Raven, 1970). The precise nature of the
subcortical accumulations has not been resolved (Luchtel, 1976).
The presence of junctions between follicle cells and oocytes can be noted (Figs. 19,
21, 22; see also Anderson, 1969; Taylor
and Anderson, 1969; Coggeshall, 1970). In
I lyanassa these two cell types are in apposition during early oogenesis with separation
resulting in wide interfacial clefts occurring
in later stages (Taylor and Anderson,
1969). It appears that similar processes also
occur in Lymnaea (Recourt, 1961) and Aplysia (Coggeshall, 1970). In view of the
studies of Raven (see 1970) implicating follicle cells in influencing ooplasmic localiza-
tions, closer attention must be paid to
membrane substructure in those regions
where follicle cells and oocytes are closely
associated.
Often the follicle cells form a rather simple squamous-like epithelium. In some instances, as in the amphineurans, they may
become highly sculptured and elaborate
(Selwood, 1968; Anderson, 1969; Richter
and Gotting, 1974). Follicle cells in the
gastropod Viviparus are elongate with narrow base on the basement membrane
(Bottke, 1972). In certain amphineurans
portions of the ovarian epithelium grow
around the oocyte to become follicle cells
while others grow as folds or sheets between developing oocytes and become
interstitial tissue (Fig. 1; see also Anderson,
1969). The initially squamous follicle cells
of these amphineurans surround the oocyte throughout oogenesis.
Histochemical and ultrastructural
studies of follicle cells indicate that they are
highly active cells (Taylor and Anderson,
1969; Bottke, 1972). Figures 2, 3 and 4
reveal the overall features of the follicle
cells with their ovoid nucleus and basophilic
cytoplasm. Electron micrographs (Figs. 19,
20) show the presence of numerous ribosomes and a well developed rough endoplasmic reticulum and Golgi. In I lyanassa,
large accumulations of glycogen have also
been noted in follicle cells (Taylor and Anderson, 1969). These cells may function as
intermediaries between the oocyte and surrounding tissues or blood plasma. Their
structure implies an active role in oogenesis. The possibility exists that they are engaged in the regulation of flow and/or modification or production of materials to
either be incorporated into the ooplasm or
the oocyte surface. The presence of endoplasmic reticulum and Golgi has led Bottke
(1972) to suggest that the follicle cells of
Viviparus provide a storage site and also
function in reabsorption of degenerating
oocytes. Follicle cells of the nudibranch
FIGS. 15, 16 and 18. Ultrastructure of the nurse cells
of Diopatra; nurse cell nucleus (NU), bipartite nucleolus (NC), lipid body (L), mitochondria (M), microvillous border (MV), and clumps of granular cytoplasmic material (GR). x 12,000; X9000; x35,000.
FIG. 17. This is an electron micrograph of a portion of
a young oocyte of the chiton, Chaetopleura apkulata.
Note the smooth oolemma (OL), thin follicle cell (F),
prominent nucleus with the distinct nucleolus (NC),
the clumps of granular material in the cytoplasm (GR),
and mitochondria (M). x 10,500. (from Anderson,
1969)
COMPARATIVE SPIRALIAN OOGENESIS—STRUCTURE
i^W^^li
323
324
20
ERWIN HUEBNER AND EVERETT ANDERSON
COMPARATIVE SPIRALIAN OOGENESIS—STRUCTURE
Cratena also appear to function in oocyte
reabsorption (Huebner, unpublished).
Bottke (1972) notes the presence of microtubules within the follicle cells of Viviparus
which are possibly related to the unusual
shape of these cells.
The precise role(s) of follicle cells is not
fully understood. They have been shown to
be responsible for producing the intricate
secondary coat around chiton oocytes (Anderson, 1969).
MISCELLANEOUS CELL TYPES
Included in this category are the surrounding parenchyma cells, as in the case
of the turbellarian Hydrolimax (Newton,
1970) or other specialized cell types within
the ovary. With respect to the latter situation many of the cells in Planorbis, which are
referred to as auxiliary (Starke, 1971), are
similar to follicle cells of other spiralians.
Between developing oocytes in the chiton
ovary shelves of interstitial tissue can be
found. The cells which constitute this tissue
are highly basophilic, and contain numerous ribosomes, glycogen, lipid droplets and
rough endoplasmic reticulum. Presumably
they have a nutritive function (Anderson,
1969). The large Leydig cells surrounding
the ovarian tubules of Ilyanassa, although
structurally different from chiton interstitial tissue cells, are also rich in glycogen
and are thought to be nutritive or storage
structures (Taylor and Anderson, 1969).
The acini of the ovotestis of Aplysia are
surrounded by small muscle cells
(Thompson and Bebbington, 1969,
Coggeshall, 1970) which respond to neurosecretory bag cell products and function in
egg laying (Coggeshall, 1972).
The efforts of the variety of cell types
associated with the oocyte (nurse cells, follicle cells and miscellaneous cells), whether in
regulation of flow, transport, secretion of
precursors, possibly modification of the
surface of the gamete or production of enFIG. 19. Portion of a follicle cell in Ilyanassa showing
the nucleus (N), prominent endoplasmic reticulum
(ER), well developed Golgi complex (GC), and numerous mitochondria (M). x22,000 (micrograph courtesy
of G. Taylor)
325
velopes, are to ensure the orderly development and release of the egg.
OOCYTIC ASPECTS
General comments
During its growth the oocyte accumulates
the necessary nutritional substances, collectively known as yolk, and the necessary
cytoplasmic machinery to sustain itself and
permit early development to occur. Although probably a consequence rather
than a cause of localization, ooplasmic
components are often parceled out unequally to various blastomeres during early
cleavage. Indeed, in some cases a heterogeneous organization of ooplasm is seen in
the unfertilized egg or in the zygote immediately after fertilization (Wilson, 1927;
Costello, 1945, 1948; Davidson, 1968; Anderson and Huebner, 1968).
It has been shown that one can displace
ooplasmic inclusions without greatly altering development (Clement, 1968, 1971).
Nevertheless, the stratified or localized accumulations one sees in the mature eggs of
some organisms (Humphreys, 1962; Anderson and Huebner, 1968; Taylor and
Anderson, 1969; Reverberi, I97la,b,c;
and see Fig. 10) or the ooplasmic segregation at meiosis or fertilization in others
(Costello, 1945, 1948; Henzen, 1966) may
have some significance. Ooplasmic components, of themselves, may not causally
influence developmental potency but
rather may promote or enhance processes
within certain blastomeres, and thereby aid
in establishing developmental paths which
could, perhaps less efficiently, occur without the presence of the various gross inclusions that are displaced in centrifuged eggs.
The statement " . . . the nature of mosaicism will not be revealed by conventional
methods of electron microscopy" by Boyer
(1972) is probably true.
With this view in mind it is important to
have information on the type and nature of
FIG. 20 and inset. Portion of a follicle cell of the
nudibranch Cratena from a region as indicated in the
light micrograph inset. Note the abundance of endoplasmic reticulum (ER) present. X30.000; inset X300.
326
ERWIN HUEBNER AND EVERETT ANDERSON
structures contained in the oocyte. For convenience we shall first survey the types of
structures present and then examine
examples of oocytes where specific ooplasmic localizations occur and finally potential
sites for the inherent developmental pattern.
Oocyte stages
The precise method and number of
stages used to delineate the continuum of
oogenesis in a variety of spiralian species
depends primarily on the personal choice
of criteria of individual investigators (Anderson and Huebner, 1968; Anderson,
1969, Coggeshall, 1970; Olive, 1971 to cite
a few). All stages may occur within the
gonad (Anderson, 1969) or, as in some
polychaetes, there may be early ovarian
stages and subsequent coelomic stages
(Anderson and Huebner, 1968; Olive,
1971). But over a wide spectrum of spiralians, oogenesis may be divided into previtellogenic and vitellogenic phases. During
previtellogenesis cellular organelles such as
ribosomes and mitochondria accumulate
whereas during vitellogenesis yolk granules
accumulate within the oocyte. Envelope
formation around the oocyte may occur
during or after these phases.
Oocyte inclusions
Yolk granules and yolk formation. Both the
amount and distribution of these varied
substances differ widely among spiralians
(Taylor and Anderson, 1969; Bottke,
1973). Even considering one species such as
the archiannelid Dinophilus there is a difference in the size of protein yolk spheres
when comparing female forming oocytes
with male forming oocytes (Gru'n, 1972).
Most studies have dealt with the protein or
protein-carbohydrate yolk spheres. The
quantity of yolk in oocytes can vary from a
small amount as in the viviparous gastropod Viviparus (Bottke, 1973a) to a very
large amount as in Ilyanassa (Taylor and
Anderson, 1969).
Since autosynthesis is the prevalent
method of yolk production in a primitive
turbellarian, it may evolutionarily represent the original system (Boyer, 1972).
There are relatively few examples in the
spiralia of oocytes that rely as significantly
on heterosynthetic (from outside the ovary)
sources of yolk compared with the insects
(Huebner and Anderson, 1972). Where
such sources exist the material is incorporated via the process of endocytosis (Dumont, 1969). A survey of the literature
shows autosynthetic yolk formation to be
very wide spread among the groups under
consideration. This can involve synthesis
within the cisternae of the endoplasmic reticulum as suggested in the turbellarian
Prostheceraeus (Boyer, 1972), the planarian
Dugesia (Grasso and Quaglia, 1970; see also
Gremigni and Benazzi, 1969), the mollusc
Aplysia (Coggeshall, 1970) and the archiannelid Dinophilus (Gru'n, 1972). Or it may
require the conjoined efforts of the endoplasmic reticulum and Golgi complex as
suggested in the mussel Anodonta (Beams
and Sekhon, 1966), the gephyrean
Priapulus (N0rrevang, 1965), the annelids
Diopatra (Anderson and Huebner, 1968)
and Enchtraeus (Dumont, 1969) and the
molluscs Ilyanassa (Taylor and Anderson,
1969), Chaetopleura and Mopalia (Anderson,
1969) and Viviparus (Bottke, 1973a).
Numerous earlier studies have referred
to a structure called the "Balbiani body" or
yolk nucleus, which is thought to function
in yolk production (Rebhun, 19566). The
precise organelle composition of the Balbiani body varies in different species. Since
the term can refer to a variety of subcellular
components, it conveys no information and
should be abandoned (Longo and Ander-
FIG. 21. Desmosome-like structures (MAF) between and less developed ER in the oocyte (O). There are
follicle cells (F) and desomosme-like structures (MA V) regions of close association and some interfacial clefts
between oocyte (OC) and follicle cells in early stage III (IF) present between the oocyte and follicle cell.
oocytes of the chiton, Mopalia muscosa. x20,000. (from x22,000. (micrograph courtesy of G. Taylor)
FIG. 23. Portion of a stage II oocyte of the chiton,
Anderson, 1969)
FIG. 22. Section showing association (arrows) of the Chaetopleura apiculala. Microvilli (MV), Golgi complex
follicle cell (F) with an oocyte in an Ilyanassa ovary. (GC), cortical granules (CG), and cortical filaments
Notetheendoplasmicreticulum (ER) in the follicle cell (FO). x 20,000. (from Anderson, 1969)
COMPARATIVE SPIRALIAN OOGENESIS—STRUCTURE
327
328
ERWIN HUEBNER AND EVERETT ANDERSON
son, 1970, 1974). In the case of Spisula, a
clam, the so-called "yolk nucleus" (Rebhun,
19566) is seen to consist of regions of endoplasmic reticulum and yolk (Longo and
Anderson, 1970; Sachs, 1972).
The presence of endoplasmic reticulum
and Golgi in the oocytes of Mopalia and
Diopatra is shown in Figures 23, 24, 26, 28,
29. The papers by Anderson and Huebner
(1968), Anderson (1969) and Taylor and
Anderson (1969) elaborate and present
ultrastructural evidence on the role of these
organelles in yolk production and need not
be reviewed here.
Eventually the successive fusions of vesicles from the synthetic organelles result in
the formation of electron dense proteincarbohydrate yolk spheres. Their internal
structure can consist of a homogeneous
granularity (Fig. 24; see also Anderson and
Huebner, 1968) or may be bipartite (Figs. 4,
31) with a central crystalline portion as in
Ilyanassa (Taylor and Anderson, 1969) or
have other varied structures as in Limnaea
(Bluemink, 1969). In the annelidEnchtraeus
some material is derived heterosynthetically by pinocytosis (Dumont, 1969).
Here the material incorporated by pinocytosis forms the dense matrix of the bipartite yolk spheres.
Intramitochondrial yolk formation has
been observed in the mollusc Planorbis
(Favard and Carasso, 1958; see also N0rrevang, 1968 for further discussion). Presumably the mechanism of formation is
similar to the process described ultrastructurally in amphibians (Massover, 1971).
In addition to containing nutritional
moieties certain yolk spheres have also been
shown to contain lysosomal enzymes
(Dalcq, 1963; Dalcq and Pasteels, 1963;
Bluemink, 1969). It is possible that these
enzymes are activated during development
and function within the confines of the yolk
sphere thereby releasing "nutrients" for
the embryo. The change in electron density
and peripheral vacuolation seen in the cortex of yolk spheres in early cleavage stages
FIGS. 24-26. Ooplasm of growing oocytes of the polychaete, Diopatra cuprea. As seen in Figure 25 the various components are randomly dispersed. Primary envelope (PE), endoplasmic reticulum (ER), mitochondria (M), yolk (Y), lipid body (L), multivesicular body
of Diopatra (Fig. 48) may be a consequence
of such intra-yolk enzyme activity.
The liquid droplets (Figs. 24, 29) that
accumulate during oogenesis are as prominent as dense yolk spheres; however, we
have no information as to their origin.
Their presence in the nurse cells of Diopatra
(Fig. 13) and occasionally in nurse celloocyte bridges suggests that nurse cells may
provide some of the lipid spheres. Glycogen can also be considered a form of yolk
and certainly ample evidence exists of its
presence in spiralian oocytes (Taylor and
Anderson, 1969; Bottke, 1973a).
Other membrane-bound granules and inclu-
sions. Along with the various yolk inclusions
one often notes the presence of pigment
granules (Fig. 26 and Anderson and
Huebner, 1968). These are synthesized by
the endoplasmic reticulum and Golgi complex.
Another structure often present is the
multivesicular body. As Figure 25 suggests
the vesicles in Diopatra eggs are likely
formed by an inpocketting of the surface
membrane. Multivesicular bodies (MVB)
have been reported in the oocytes of other
spiralians (N0rrevang, 1965, 1968; Pasteels, 19666). Dohmen and Verdonk (1974)
comment on their presence in the polar
lobes of some animals. The function of
multivesicular bodies in oogenesis is unknown.
Not all spiralian oocytes (e.g., Ilyanassa,
Taylor and Anderson, 1969; Diopatra, Anderson and Huebner, 1968; Priapulus,
N0rrevang, 1965, 1968) possess cortical
granules or cortical alveoli. Cortical
granules have been observed in the rotifer,
Asplanchna (Bentfeld, 197la,b); the turbellarian, Prostheceraeus (Boyer, 1972); the annelids, Pornatocerous, (Gwynn and Jones,
1971); Nereis (Pasteels, 1966c; Fallon and
Austin, 1967; Dhainaut, 1969); the
echuroid worm, Urechis (Gould-Somero
and Holland, 1974) and the molluscs,
Chaetopleura and Mopalia (Anderson, 1969)
and Spisula (Longo and Anderson, 1970).
(MVB-note inpocketing *), and pigment granules (P).
In Figure 26 the prominent Golgi complex (GC) is
associated with vesicles and forming pigment granules
(P). x 15,000; x20,000; x25,000.
COMPARATIVE SPIRALIAN OOGENESIS—STRUCTURE
329
330
ERWIN HUEBNER AND EVERETT ANDERSON
31
FIGS. 27-31. Periphery of Diopatra oocytes illustrating first, and major layer (MP) produced later, with microsuccessive stages of primary envelope formation.
villi (MV) protruding and indications (arrows) of the
Present are aspects of the minor layer (MI) produced fusion of vesicles of endoplasmic reticulum. Figure 31
COMPARATIVE SPIRALIAN OOGENESIS—STRUCTURE
331
Cortical granules vary in their histo- received little attention in studies of spirachemical staining properties (Anderson, lian oogenesis. Filaments can be seen in
1969; Boyer, 1972) but generally contain a Mopalia oocytes (Fig. 23; see also Anderson,
mucopolysaccharide and protein. In most 1969). The significance of filaments within
instances they are produced by the endo- the oocyte (particularly the cortex) is unplasmic reticulum and Golgi of the oocyte. known. From the studies of Raff (1972) on
In the rotifer Asplanchna they are synthe- the effects of microtubule and microsized by the nurse cell-like vitellarium and filament disrupting agents on polar lobes of
transported to the oocyte (Bentfeld, 1971). Ilyanassa and Arnold and Williams-Arnold
In some instances, some or all cortical (1974) on the effects of cytochalasin on the
granules or alveoli are released at fertiliza- squid egg cortex, it is clear that a retion (Fallon and Austin, 1967; Bentfeld, examination of the distribution and struc19716; Gould-Somero and Holland, 1974) ture of microfilaments in spiralian oocytes
whereas in others they are retained until would be fruitful. Besides simple supporlater (Humphreys, 1967; Longo and An- tive function they may also have the funcderson, 1969, 1970; Sachs, 1971).
tion of directing and implementing ooGru'n (1972) outlines the production and plasmicflowand segregation. Microtubules
storage of mucopolysaccharide granules in are present in the stalks which attach dethe oocytes of the archiannelid Dinophilus. veloping Anodonta oocytes to the ovarian
These are released after oviposition and wall (Beams and Sekhon, 1966).
form the egg capsule.
Granular areas. At the onset of vitelloMembrane systems. The presence and role genesis in Ilyanassa, granular perinuclear
of the usual organelles, the endoplasmic bodies appear in the oocyte (Taylor and
reticulum and Golgi, have been noted Anderson, 1969). Similar structures occur
above. Here we draw attention to the annu- in immature chiton oocytes (Fig. 17) but not
late lamellae (AL) found in oocytes. Annu- mature eggs (Anderson, 1969). Figure 18
late lamellae have been reported in numer- shows similar structures are present in
ous molluscan and annelid oocytes (Reb- Diopatra nurse cells. In the polyclad Proshun, 1956«; Durchon and Boilly, 1964; theceraeus nucleus-derived dense material
Anderson, 1969; Dumont, 1969; Taylor remains clumped in localized areas (Boyer,
and Anderson, 1969; Gwynn «/ «/., 1971; 1972). As noted by Anderson (1969), in
Dhainaut, 1973; Bottke, 1973a). The lack chitons these particles are derived from the
of cerebral hormone-stimulated prolifera- nucleus and consist of ribonucleoprotein.
tion of annulate lamellae in Nereis (Dur- Studies by Dhainaut (19706) using 3Hchon and Boilly, 1964; Dhainaut, 1973). uridine labelled Nereis eggs lend support
Dhainaut (1973), using Nereis, suggested to this. In addition, evidence from studies
that annulate lamellae were formed from of 3 H-uridine and 3H-leucine labelling of
endoplasmic reticulum. Gwynn et al. nuclear emissions in mictic eggs of rotifers
(1971), using the annelid Pomatocerous pre- (Bentfeld, 197 la,b) suggests that similar
sented data which suggest that stacks of structural material is composed of riboannulate lamellae produced from the nu- nucleoprotein. Taylor and Anderson
clear envelope in successive waves and act (1969) speculated that the granular areas in
as a means of transporting "sandwiches" of Ilyanassa oocytes were RNA, but Gerin
nuclear material into the ooplasm. The (1971) has noted that the perinuclear
reader is referred to Kessel (1968) and bodies in young Ilyanassa oocytes were not
Wischnitzer (1970) for reviews on the annu- removed by RNAase but rather by pronase
and pepsin. Clarification of whether the
late lamellae.
Microfilaments and microtubules. Until re- bodies described by Gerin (1971) are the
cently these two important structures have same as those reported by Taylor and Andepicts the bifurcation of the microvilli. x40,000;
X50.000; X40.000; x40,000; X40.000.
FIGS. 32 and 33. Tangential and oblique sections of
the envelope showing the fibrillar nature of the major
layer as well as the regular pattern of microvilli (MV)
along the surface (arrows) of the oocyte. x40,000;
x20,000.
332
ERWIN HUEBNER AND EVERETT ANDERSON
FIGS. 34-36. Phase-contrast micrographs illustratinga
developing oocyte with its primary envelope (Fig. 34),
a 4 seteger larva (Fig. 35) and a highly developed larva.
X200; x200; x60.
FIGS. 37 and 38. The electron micrograph in Figure
37 and inset illustrate the nearly mature oocyte envelope with its minor and major layers (MI and MP).
Figure 38 shows the cuticle of a developing larva.
COMPARATIVE SPIRALIAN OOGENESIS—STRUCTURE
333
derson (1969), as well as information on the oogonium and/or from nurse cells (Antheir chemical nature is needed. Gerin derson and Huebner, 1968). In Diopatra
(1972) showed an association of granular oocytes, mitochondria are initially ranmaterial with endoplasmic reticulum dur- domly distributed (Fig. 24); however, by
ing oogenesis. These membrane vesicle the time the oocyte reaches maturity they
structures were later observed in the polar are present in great preponderance in the
lobe of the embryo.
animal pole (Fig. 41) and frequently in the
At present it is unclear if the granular oocyte cortex (Fig. 42). The abundance of
clusters or their dispersed components mitochondria in the oocyte cortex was also
represent localizations which contain noted by Boyer (1972). Most, if not all cytonucleus-derived information to be used in plasmic DNA is located within the mitolater development. Anderson (1969) and chondria (Dawid and Brown, 1970 and
Boyer (1972) suggest that the nuclear ex- Dawid, 1972).
trusions may be related to mosaicism in
Oocyte nucleus. Spiralian oocytes have
spiralian animals.
prominent germinal vesicles and nucleoli
The abundance of ribosomes in many (Wilson, 1927 and Figs. 2,3,4, 9,12,17 and
oocytes is obvious. There may be differen- 34). The shape can vary from pleomorphic
tial distribution of RNA as noted in Diopatra initially, (Bottke, 1973) in some cases to
(Allen, 1961). An unusual association of nearly round or ovoid in others (Taylor and
ribosomes with amylase-sensitive material, Anderson, 1969). Complete discussion of
resulting in clumps, has been reported in the nucleus and nucleolus in spiralians
Ilyanassa (Pucci-Minafra et al., 1969); how- could be the topic for an entire paper; conever, Geuskens and d'Ardoye (1971) have sequently, we refer the reader to Raven
found contradictory results using the same (1961), Reverberi (1971), Dhainaut (1972),
organism.
Bottke (19736), and Kietbowna and Koscielski
(1974) for further discussion of nuMitochondria. In view of the potential role
of mitochondria in embryogenesis, their clear morphology and nucleolar transforabundance and distribution in spiralian mations. Figures 3 and 4 or Cratena illuoocytes is of considerable interest strate the prominence of nucleoli. Figure 4
(Lehmann, 1941; Reverberi, 1958, 1970a, depicts a typical so-called amphinucleolus
b, 1971c; Hersh, 1969; Geuskens and d'Ar- that is reminiscent of the bipartite nucledoye, 1971; Dawid, 1972). In Priapulus olus of Diopatra.
there are five to eight mitochondria in a
young oocyte and approximately 40,000 in Oocyte surface
the mature egg (N0rrevang, 1965). The
major responsibility for synthesizing most,
Generally the oolemma of the very young
if not all mitochondria for the embryo rests oocyte is relatively unspecialized and thus
with the female (Dawid and Brown, 1970; has few microvilli (Fig. 17). As seen in FigDawid, 1972). As in Priapulus, mito- ures 21 and 22 the smooth early oocyte
chondria of spiralian oocytes display varied membrane can form junctional areas with
configurations ranging from elongate, to follicle cells. Differentiation of the
"donut-shaped," "cup-shaped," and oolemma is usually accompanied by the de"dumbbell-shaped" (N0rrevang, 1965; An- velopment of microvilli (Anderson and
derson and Huebner, 1968; Anderson, Huebner, 1968; Dumont, 1969; Fallon and
1969; Taylor and Anderson. 1969; Du- Austin, 1969; Gwynn and Jones, 1971;
mont, 1969 and Bottke, 1973a). The Longo and Anderson, 1970). N0rrevang
dumbbell-shaped mitochondria (Fig. 27) (1968) noted that the microvilli are small
are thought to represent division figures. and few in number in polychaetes. HowMitochondria of the mature egg are de- ever, as evidenced by the studies cited
rived from pre-existing mitochondria of above and Figures 27 to 31 presented here,
Microvilli (MV) penetrate the fibrillar cuticle with its
outer and an inner portions (MI and MP). The inset
demonstrates the fibrillar nature and microvilli of the
adultcuticle. X30.000; inset, x40,000; X22 000- inset
x22,000.
334
ERWIN HUEBNER AND EVERETT ANDERSON
%
GV
•is
-b
i
•
• *
-
. *
.'?•
COMPARATIVE SPIRALIAN OOGENESIS—STRUCTURE
microvilli are very numerous on many such
oocytes and often in a regular array (Figs.
32, 33). Initially microvilli may be rather
simple morphologically; however, some
may become complex with their tips extending through the egg envelopes and
branching. This increase in surface area of
the Diopatra oocyte probably facilitates increased uptake of precursors through the
oolemma from coelomic fluids.
Interesting changes occur in the
oolemma during Ilyanassa oogenesis
(Taylor and Anderson, 1969). At the onset
of vitellogenesis the maculae adhaerentes
which were interconnecting young oocytes,
disappear and interfacial clefts enlarge.
Short microvilli appear on the surface of
the oocyte protruding into the lumen. This
is the future animal pole. Histochemical
staining for mucopolysaccharide has shown
that only the microvillar oolemma area has
positive material on its surface (Taylor and
Anderson, 1969). This may represent regional chemical differences in the oolemma. The reader is referred to Pasteels
(1966fl) for discussion of microvillar and
cortical movements in early mollusc development.
335
ary and tertiary to designate envelopes
formed by the oocyte, follicle cells and
extraovarian sources respectively was originally proposed by Ludwig (1874). Figures
27 to 33 and 37 and inset illustrate stages in
primary envelope formation in the polychaete Diopatra cuprea. The envelope precursors are synthesized within the endoplasmic reticulum and released by
exocytosis. The fully formed coat has a
major and minor layer and the minor layer
has sublayers (Fig. 37) as originally described by Anderson and Huebner (1968).
Microvilli, arranged in a regular pattern
(Figs. 32, 33), penetrate through the envelope and branch at their tips. This envelope produced during oogenesis of
Diopatra is analogous to internal structures,
such as yolk, because it is utilized and modified throughout the embryonic stages, and
eventually becomes a portion of the cuticle
of the larva and finally the adult (Figs. 3438). Thus, in Diopatra the animal does not
hatch out of its egg envelopes but utilizes it
fully as a part of its external surface.
Regional localizations or stratification
Subsequent to or during vitellogenesis,
egg envelopes are deposited on the surface
of many oocytes. These are virtually
nonexistent in some species (Taylor and
Anderson, 1969). In others they may be
homogeneous or composed of intricate
sub-structures (Anderson, 1969; Gwynn
and Jones, 1971; Fallon and Austin, 1967;
Anderson and Huebner, 1968; Longo and
Anderson, 1970; Noda and Takashima
published in N0rrevang, 1968). The reader
is referred to Anderson (1969) for discussion of the highly ornate coat of chiton oocytes which consists of a primary and secondary envelope and sculptured outer
layer consisting of follicle cells.
The usage of the terms primary, second-
Timing. As alluded to earlier, there are
definite examples of localization or stratification of ooplasmic materials in the mature egg (Allen, 1961; Raven, 1961; Humphreys, 1962; Anderson and Huebner,
1968; Taylor and Anderson, 1969; Longo
and Anderson, 1969; Reverberi, 197077,6,
1971a,c). In other instances this localization
is not manifested until after release from
the ovary, meiosis or fertilization (Costello,
1945, 1948; Raven, 1963, 1967, 1970; Henzen, 1966). Comparisons of the mature
eggs of two polychaetesD/o/?«/ra and Nereis
(Figs. 10, 11) illustrate this point. Costello
(1945, 1948) clearly demonstrated ooplasmic movements and ooplasmic segregation
in Nereis after fertilization. Raven (1963,
1967, 1970; see also Hess, 1971) showed in
Lymnaea that the appearance of the subcortical accumulations did not appear until the
FIG. 39. Section through the animal region of a postvitellogenic Ilyanassa oocyte showing lipid spheres (L),
mitochondria (M), very few yolk spheres (Y), and nucleus (GV). x22,000 (micrograph courtesy of G.
Taylor)
FIG. 40. This electron micrograph reveals the presence of yolk spheres (Y), mitochondria (M), and some
lipid spheres (L) located in the vegetal hemisphere of a
postvitellogenic Ilyanassa egg. X8.000. (micrograph
courtesy of G. Taylor)
Envelope formation.
336
ERWIN HUEBNER AND EVERETT ANDERSON
oocyte was in the oviduct. Animal-vegetal stituents of the mature llyanassa egg are
differences were present earlier. In Tubifex uniformly distributed; however, Taylor
the localizations of the polar plasms occurs and Anderson (1969) subsequently showed
at meiosis (Lehmann, 1948; Henzen, 1966). that definite animal-vegetal differences did
Where evidence of ooplasmic segrega- exist in mature ovarian eggs. (Figs. 39, 40).
tion can be shown to exist, there are differ- The animal pole contains lipid droplets,
ences in the timing of this appearance. This mitochondria and relatively few small yolk
was originally noted by Whitman in 1878 spheres, whereas, the vegetal area has
who showed that regional differences in numerous yolk spheres, mitochondria and
eggs of Clepsine appeared only after mat- other ooplasmic organelles (Taylor and
uration. In the case of the nemertine Cere- Anderson, 1969). Presumably this pattern
bra tulus, Freeman (1974) has been able, ex- is altered after fertilization since Clement
perimentally, to induce precocious forma- and Lehmann (1956, see also Clement,
tion of localization in egg fragments that 1971) reported an abundance of mitochondria in the animal pole and Crowell
specify apical tuft and gut.
Types of localizations. The establishment of (1964) noted the presence of very few mitopolarity in many yolk-laden eggs as chondria in the polar lobe. Geuskens and
animal-vegetal differences is related to the d'Ardoy (1971), however, suggested that
future cephalo-caudal axis of development mitochondria are important components
(Reverberi, 197 \a). This polarity of the egg of the polar lobe. Perhaps some of the mitomay be manifested by a variety of chondria from the oocyte vegetal pole rephenomena including distribution of yolk main there during early development.
or mitochondria, or position of the germi- Gerin (1972, see also Dohmen and Vernal vesicle (Wilson, 1927). It was Mark donk, 1974) have drawn attention to mem(188 1) who first stated that the primary po- brane vesicles which in llyanassa form durlarity of the egg might be determined by ing oogenesis and are localized later in the
"the topographical relation of the egg polar lobe. The study of Dohmen and Ver(which is still in an indifferent state) to the donk (1974) on the gastropod Bithynia has
remaining cells of the maternal tissue from shown the presence of an RNA positive
which it differentiated." As further discuss- cup-shaped mass of small vesicles referred
ed by Hess (1971) and Raven (1961, 1970) to as the vegetal body in the polar lobe.
the localization seen in the oocyte or early Dohmen and Verdonk (1974) are of the
embryo is influenced by its cellular inter- opinion that this material is a morphoactions during oogenesis. Hess (1971) genetic cytoplasm. This further emphasizes
noted that presumably materials such as that we should re-examine the nature and
yolk, mitochondria, endoplasmic re- distribution of vesicles of this type during
ticulum, as well as products such as various oogenesis. No detailed study on their distypes of RN A and metabolites are unevenly tribution within the mature ovarian egg has
distributed during ooplasmic segregation. been made. For evidence of localizations in
The distributions are stabilized by the or- Mytilus one can consult Humphreys (1962),
derly process of cleavage. The pattern is Reverberi (1967, 1971ft).
highly stable and is predetermined genetiThe oocyte of Diopatra cuprea shows recally by the mother (Raven, 1970; Hess, gional differences in substructure as seen in
1971).
Figures 41, 42 and 43. Yolk granules are
The following are examples which illu- vegetally located, lipid is at the lateral anistrate the types of localizations or uneven mal pole, mitochondria are clustered at the
distributions seen in a few organisms. animal and the germinal vesicle is eccentric
Pucci-Minafra and Collier (1968) in a brief (Anderson and Huebner, 1968). Allen
report suggested that the ooplasmic con- (1961) noted the increased amount of RNA
FIGS. 41-43. Animal pole (Fig 41), cortex (Fig. 42)
and lipid area (Fig. 43) of a Diopatra oocyte. Mitochondria (M) with numerous dumbell shapes (arrow), cortical mitochondria (M in Fig. 42), endoplasmic re-
ticulum (ER), lipid spheres (L) with intervening organelle containing cytoplasm containing a Golgi complex (GC in Fig. 43). x22,000; x28,000; x20,000.
(Fig. 41 from Anderson and Huebner, 1968)
COMPARATIVE SPIRALIAN OOGENESIS—STRUCTURE
337
.%<
338
ERWIN HUEBNER AND EVERETT ANDERSON
49
COMPARATIVE SPIRALIAN OOGENESIS—STRUCTURE
339
in the animal pole. As seen in Figures 44 to
46, 48, and 49 the local heterogeneous
stratification of the Diopatra oocyte is retained during early cleavage. Lipid is found
primarily in the animal hemisphere and
yolk in the vegetal. Areas devoid of these
inclusions are located centrally or slightly
apically. If one examines cells at a later
stage of embryogenesis, such as the trochophore, it is apparent that certain cells have
accumulated structures such as, pigment
granules which were originally found in the
oocyte (Fig. 47). The detailed analysis of the
fate of ooplasmic materials is preliminary
and in progress (Huebner, unpublished
observations). The polar plasms of Tubifex
provide another example of localization
(Penners, 1926; Henzen, 1966, see also Reverberi, 1971a).
Work on Dentalium has shown the vegetal
pole to contain abundant mitochondria,
few microvilli and yolk spheres, and no cortical granules. The animal pole however,
has cortical granules, large yolk spheres
and many microvilli (Reverberi, 1958,
19706, 1971). The polar lobe of this species
has numerous mitochondria. Verdonk
(1968) determined that polar lobe material
is not only localized vegetally in Dentalium
but also bilaterally. The numerous studies
on Limnaea from Raven's laboratory (1961,
1963, 1967, 1970, 1974; see also Hess,
1971) in addition to showing the presence
of the polar plasms have outlined the locations of the distinctive subcortical accumulation and traced their fate. Ultrastructural
study of Limnaea eggs has not revealed any
distinctive cytoplasmic areas corresponding to the subcortical accumulation seen in
the light microscopic studies of Raven
(Luchtel, 1976). Thus, precise information
on the possible nature of these localizations
is still unclear.
Detailed consideration of the fates of
cytoplasmic localizations and the mechanism by which localization occurs can not be
covered in depth here. Studies of Clement
(1968) and Verdonk (1968) have shown
that morphogenetic factors are not present
in the displaceable cytoplasm. However,
Dohmen and Verdonk (1974) pointed out
that ultrastructural studies of displacement
of cytoplasmic components from polar
lobes are lacking. Although larger structures may be moved, they note that in
Bithynia the potentially morphogenetic
vegetal body is not displaced by centrifugation. Certainly much cytological work is required on centrifuged eggs, but many
studies have suggested that the developmental pattern of embryogenesis is imprinted in the oocyte cortex (Arnold, 1968,
1970; Raven, 1970; Cathers, 1971; Hess,
1972; Arnold and Williams-Arnold, 1974).
The studies of Arnold on the squid oocyte
have shown by centrifugation, microbeam
irradiation, topographical ligation and
cytochalasin treatment that the cortex plays
a vital role in pattern formation.
Conklin (1917) suggested that the hyaloplasm or cell surface formed the basis of
localizing activity. He assumed it to be elastic and contractile and, thus, to produce the
so-called flowing movements of living
protoplasm. Mechanisms aiding or causing
cytoplasmic movements, and hence, localizations will presumably include a microfilament mechanism. Raff (1972) has provided insight into the role of microfilaments in Ilyanassa polar lobe formation.
He notes that one has to have informationcarrying determinants synthesized during
oogenesis as well as a geometric matrix to
specify the location. Microfilaments or their
precursors are probably formed during
oogenesis. Techniques used to illustrate the
presence and distribution of microfilaments or actin-like filaments in other cell
types should be applied to the various
stages of oogenesis and during the time of
localization.
Basic structural work laying the founda-
FIGS. 44 and 45. Micrographs of a 2-cell stage Diopatra
embryo showing a living specimen with phase-contrast
and a sectioned toluidine blue stained specimen, respectively. Polar body (PB), lipid rich animal hemisphere (L), yolk rich vegetal area (V), inclusion free
areas and chorion (C). X140; x200.
FIGS. 46,48 and 49. Electron micrographs of the lipid
area (L), vegetal yolk rich area (V), and cytoplasmic
areas, respectively. Note the change in the periphery
of the yolk sphere (Y) compared with earlier micrographs. Mitochondria (M), endoplasmic reticulum
(ER). x 10,000; X30.000; x 10,000.
FIG. 47. Portion of a cell from a Diopatra trochophore
near the apical tuft. Note the large accumulation of
pigment granules (P). x 11,000.
340
ERWIN HUEBNER AND EVERETT ANDERSON
Arnold, J. M. and L. D. Williams-Arnold. 1974.
Cortical-nuclear interactions in cephalopod development: Cytochalasin B effects on the informational pattern in the cell surface. J. Embryol. Exp.
Morph. 31:1-25.
Arnold, J. M. and L. D. Williams-Arnold. 1976. The
egg cortex problem as seen through the squid eye.
Amer. Zool. 16:421-446.
Bailey, T. G. 1973. Them i^ro culture of reproductive
organs of the slug Agriolimax reticulatus (Mull).
Netherlands J. Zool. 23:72-85.
Beams, H. W. and S. Sekhon. 1966. Electron microscope studies on the oocyte of the fresh water mussel
(Anodonta) with special reference to the stalk and
mechanism of yolk deposition. J. Morph. 119:477501.
Bedford, L. 1966. The electron microscopy and cytochemistry of oogenesis and the cytochemistry of
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15:15-37.
Bentfeld, M. E. 1971a. Studies of oogenesis in the
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Bentfeld, M. E. 197li. Studies of oogenesis in the
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Biggers, J. D. and A. W. Schuetz (eds.). 1972. Prologue. In Oogenesis, pp. 1-4, University Park Press,
Baltimore, Md.
Bluemink, J. G. 1969. Are yolk granules related to
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mutants which are expressed by alterations
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This communication has presented the
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oogenesis and the structures present and
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