/. Embryol. exp. Morph. Vol. 43, pp. 55-69, 1978
Printed in Great Britain © Company of Biologists Limited 1978
55
Cleavage in the chick embryo
By R U T H BELLAIRS 1 , F. W. L O R E N Z AND T A N I A D U N L A P
From the Department of Anatomy and Embryology,
University College London and Department of Animal Physiology,
University of California, Davis, California, U.S.A.
SUMMARY
Chick embryos ranging from the stage of first cleavage to that of about 700 cells were
removed from the oviduct and examined by transmission electron microscopy. Beneath the
cell membrane is a yolk-free cortical region containing microfilaments. Beneath this lies
cytoplasm which contains yolk spheres which are graded in size, the dorsal ones being smaller
than the ventral ones. The subgerminal periblast possesses a greater proportion of yolk to
cytoplasm than do the cells proper, but it merges with the cytoplasm at the incomplete
borders of the 'open' cells.
Specialized accumulations of membranes lie in the marginal periblast, and it is suggested
that they play a role in cell membrane formation.
(This paper is dedicated to Professor Silvio Ranzi on the occasion
of his 75th birthday)
INTRODUCTION
The cytological events occurring during mitosis are similar in most types of
vertebrate cell, though they vary in detail. The most usual example of cell
division, however, is probably that found during cleavage in large-yolked
species such as the fowl. According to the classical descriptions, which are based
on the study of paraffin sections by light microscopy (Patterson, 1910; Olsen,
1942), a number of'open cells' develop. An 'open cell' is one which is bounded
by cell membrane on only a part of its surface, and whose cell contents are in
open communication with the yolk (Fig. 1). It is supposed that as each 'open
cell' completes mitosis, one daughter nucleus remains in situ and the other
migrates out together with some cytoplasm into the adjacent yolk, forming a
region known as the periblast, which is part yolk, part cytoplasm. Thus the
periblast lies around the periphery of the embryo (marginal periblast) and
beneath it (subgerminal periblast). The cell membrane now extends down
between the two cell nuclei, so that one of the cells loses its contact with the
periblast and hence with the yolk, and is no longer 'open'. At the same time, a
new ' open cell ' has formed around the other daughter nucleus.
Hitherto, the only detailed modern study of cleavage in the chick embryo
1
Author's address: Department of Anatomy and Embryology, University College London,
Gower Street, London WC1E 6BT, U.K.
56
R. BELLAIRS, F. W. LORENZ AND T. D U N L A P
Cytoplasm
,j
Open cell
Open cell
Subgerminal periblast
Marginal
periblast
Marginal
periblast
Yolk
Fig. 1. Diagram to illustrate the structure of the open cells in early cleavage. (A)
Four stages seen from the dorsal side of the embryo. The yolk is not illustrated but
the periblast is visible (from life). (B) Transverse section through the edge of an
early embryo.
Cleavage in the chick embryo
57
was that of Gipson (1974), although she restricted herself to a description of the
cleavage furrows during the first three divisions. In this paper we have not given
space to those aspects of our results which are a mere confirmation of hers.
Instead, we have investigated the structure of the 'open cells' by light and
electron microscopy. In view of the importance of ribosome formation in
young embryos (see Discussion) we have also attempted to find out whether
nucleoli are present in the early stages and, in addition, we have considered
the nature of the cleavage furrows and the possible mechanism of their formation.
MATERIALS AND METHODS
Thirty-five laying hens were killed, usually by intravenous injection of sodium
barbiturate, but occasionally by ether anaesthesia. The cleaving embryo was
dissected from the oviduct and immediately fixed at room temperature (1520 °C) and at pH 7-0-7-2 with either diluted Karnovsky's fixative (Karnovsky,
1965) or with Burnside's fixative (Burnside, 1971). In some specimens, the
entire yolk was plunged into the reagent and the young embryo dissected
from it. In others, the embryo was removed from most of the yolk before
fixation. After remaining in the fixative for periods varying from 1 h to several
days, the embryo with adherent yolk was washed in cacodylate buffer (pH 7-07-2) and treated with buffered 1 % osmium tetroxide for 1 h at pH 7-0. During
dehydration with ethanols the specimens were stained with Araldite or Epon.
Sections were stained with lead citrate and examined with either a Siemens
Elmiskop 1 or with an A.E.I.-801 electron microscope.
Thick sections were routinely obtained from the same blocks and were
stained with a 1 % aqueous solution of toluidine blue and examined by light
microscopy. Twenty-three additional specimens were embedded in soft plastic
and serially sectioned at 4 jam. They were stained with acid fuchsin and toluidine
blue according to the technique of Ruddell (1971).
Three further specimens were treated at 37 °C with a 1 % solution of aamylase made up in 0-85 % saline at pH 6-0. Each specimen was cut into three
pieces, one being treated for 1 h, another for 3 h, and the third for 6 h. Control
specimens were incubated with saline for corresponding periods. After treatment, specimens were washed briefly in fresh saline and processed in the same
way as the main group of embryos.
In all, 82 specimens were examined ranging in development from the stage
of first cleavage to about 700 cells. In the earliest stages when all the cells were
still open it was sometimes difficult to decide how many were present, especially
since the furrows soon became irregular. The following criteria were therefore
adopted :
No. of cleavage furrows
1
2
3
4
5
No. of cells
2
4
6
9
12
58
R. BELLAIRS, F. W. LORENZ AND T. D U N L A P
Where possible, the terminology of Gipson (1974) has been used to describe the
ultrastructural details.
RESULTS
The general relationship of the cleaving embryo to the vitelline membrane is
shown in Figs. 2 and 3. A fine granular material lies between the vitelline
membrane and the embryo, and extends into the furrows between the cells.
The structure of the vitelline membrane of the fowl has been described elsewhere
(Bellairs, Harkness & Harkness, 1963; Jensen, 1969) and will not be discussed
here.
Until about the 16-celled stage, all cleavage cells are 'open' (Romanoff, 1960)
and in direct communication with the periblast (Figs. 1, 2). When these 'open'
cells are examined by electron microscopy three major zones are apparent:
the cell membrane, the cortical region and the yolky cytoplasm (Fig. 3). The
last grades into the periblast which is rich in yolk spheres and poor in cytoplasm, and this in turn grades into the totally non-cytoplasmic yolky region
(see Fig. 1). By the 16-celled stage the most centrally situated cells have become
completely surrounded by cell membranes, though those in the more peripheral
regions remain 'open'. Eventually, the 'open' cells disappear.
Gipson (1974), who examined only 2-, 4- and 8-cell stages, classified the
furrows as being V-shaped, U-shaped or pendulum-shaped, in longitudinal
section. We have not found it possible to distinguish between the V-shaped
and the U-shaped since many of the variations which occur may be the result
of different angles of sectioning. We prefer to rename both of these types as
shallow furrows (Figs. 4, 5). The deeper, pendulum-shaped furrows are characterized in section by a narrow neck which swells into a bulbous region at its
deepest extent. We have also seen an additional type of furrow not described
by Gibson, which is characterized by relatively smooth walls and apparently
by tight junctions (Fig. 3). We have named this slit-like. This too is found in
later stages and is probably restricted to the most peripheral regions; it does
not seem to be related to the stage of the embryo. We have seen all three types
of furrow at all stages examined from the 2-cell to the 700-cell stages. Many of
the furrows, especially the pendulum-shaped ones, possess highly elaborate
FIGURES 2 AND 3
Fig. 2. Light micrograph of a 4 /*m Ruddell's plastic section through an open cell
of an embryo with 12 cells, p: periblast; s: mitotic spindle; t: transverse furrow;
vm : vitelline membrane. Arrow points to junction of open cell and ventral periblast
x 160. Inset: enlargement of mitotic spindle x 1500.
Fig. 3. Electron micrograph of a section through a slit-like furrow at the periphery
of an embryo with 12 cells. The regions of contact between the walls of the furrow
are probably tight junctions (arrowed). Note the three regions; cell membrane (1),
cortical region (2) and yolky cytoplasm (3). x 6000. vm: vitelline membrane.
59
Cleavage in the chick embryo
**S'
«41» . * * .
W'.£
^i#;*
is
S5^Ä
P-^^
:fj&%^"
,
«
#
•
*
•«•*>
fi
60
R. BELLAIRS, F. W. LORENZ AND T. D U N L A P
•;e>:'o-d
$•'©''••>:
<9:
P./
&X.èó%
: :
0^:': '.:'-':.^^-.^.'--'.
X'
o • ;
O"
®P.^'0#P®-:°^©.P<9'
«life*
J-J® $•:<>•
mmm
fêÖ
.•<?...
.'o
\-y-0:
:
^^^!s^i^
o-
:--. : ®V-:->!«; : V/Î);Ô?Fig. 4. Diagram to illustrate the three types of cleavage furrow. (A) Slit-like;
(B) shallow; (C) pendulum-shaped.
Cleavage in the chick embryo
61
Fig. 5. Electron micrograph of a section through a shallow furrow from an embryo
with about 16 cells, x 1500.
Fig. 6. Electron micrograph of a section through the base of a pendulum-shaped
furrow to show the region of the furrow-base-body taken from an embryo of 16
cells. X1400.
Fig. 7. Electron micrograph of a section through the junction of an open cell and
the periblast in an embryo of 64 cells (see Fig. 1). Note the extension of furrowbase-body-like membrane, x 1500.
Fig. 8. Electron micrograph of a section through an embryo with about 700 cells.
Note the membranous projections reminiscent of the furrow-base-body, x 4000.
structures at the base (Figs. 6, 11). Typically, each consists of a furrow-basebody with branching processes extending from it. (For a full description, see
Gipson, 1974.) Examination of many thick sections by light microscopy has
convinced us that there is only one region of base-body per furrow, though our
electron micrographs have demonstrated that in that location the furrow-basebody may have several attachments (Fig. 6).
5
EMB 43
62
R. BELLAIRS, F. W. LORENZ AND T. D U N L A P
Fig. 9. Electron micrograph of a section through a patch of membranous vesicular
reticulum in an embryo of about eight cells. It lies just beneath the cortex and
distal to the cleavage furrows and may therefore be considered to be part of the
periblast, x 5200.
Fig. 10. Electron micrograph of a section through microvilli in the neck of a pendulum-shaped furrow, x 12500.
By the 64-cell stage the open cells are restricted to the peripheral region. The
cell membrane of each open cell is continuous with the cellular membrane
covering the periblast. Fig. 7 is a micrograph of a section at the angle between
an open cell and periblast and shows a region of projections resembling a
furrow-base-body. Similar projections are shown in Fig. 8 on the surface of
fully formed cells from late cleavage (about 700 cells).
FIGURES
11-13
Fig. 11. Electron micrograph of a section to show the root region (arrowed) which
lies ventral to the furrow-base-body. Taken from an embryo of 16 cells, x 6250.
Fig. 12. Electron micrograph of a section in the region where the periblast merges
into the yolk. Note the large closely packed yolk granules, each with a densely
stained core (c) and a less dense peripheral region (p); note also the mitochondria
(w). x6250.
Fig. 13. Electron micrograph of a section through the nucleus («) in an embryo of 64
cells. Note the absence of nucleoli, x 5000.
Cleavage in the chick embryo
63
5-2
64
R. BELLAIRS, F. W. LORENZ AND T. D U N L A P
The cell membrane along the dorsal surface of the open cells exhibits many
pinocytotic pits and vesicles and is also extended out as microvilli. Beneath
the cell membrane lies a cortical zone. This is free of yolk and usually about
\ /im thick. It contains an array of microfilaments which appear to run parallel
to the surface of the membrane, though it is not always easy to see them in the
older cleavage stages. This suggests that they are frequently not arranged to
run in a predominantly single direction. In the microvilli (Fig. 10), however,
they tend to lie in conspicuous parallel bundles and are sometimes interspersed
with fine granules which individually measure about 10 nm across and appear
to be ribosomes. Occasionally glycogen granules are present in the cortex.
Beneath the cortex is the cytoplasmic region which contains the same components in both open and closed cells, both in early and later cleavage. It is
characterized by the presence of densely staining granules each about 300 nm
in diameter. These granules are often arranged in clusters and are considered
to be glycogen. In specimens treated with a-amylase many of the granules
sometimes extended into the projections in the furrows.
Other components of the cytoplasm include yolk spheres (Fig. 11), mitochondria, ribosomes, Golgi bodies and annulate lamellae. Granular endoplasmic
reticulum is scarce but smooth surfaced vesicles are common.
Two regions of highly specialized cytoplasm may be noted. The membranous
vesicular reticulum (Fig. 9) tends to lie near the dorsal surface, especially in the
marginal periblast of young cleavage stages. It consists of a dense accumulation
of small vesicles which individually measure between about 0-2 and 0-5 ^m
in diameter and which frequently contain a fine granular material. Larger
vesicles are also present, though these tend to be relatively empty.
The second type of specialization is seen directly beneath many of the
furrow-base-bodies (Fig. 11). It consists of a dense concentration of cytoplasm
which passes like a root deep into the periblast and consists of an aggregation
of Golgi bodies, vesicles and mitochondria (Fig. 4C).
The yolk spheres present in the cytoplasm are usually of the relatively simple
type described by Bellairs (1958) in which each consists of a densely staining
granular core surrounded by a less dense zone (Fig. 11). Some of those lying
most dorsally measure as much as 10 /mi in. diameter and are separated by
cytoplasm. More deeply situated yolk spheres are larger with less cytoplasm
between them. This changing relationship between the size of the yolk spheres
and the amount of cytoplasm is especially noticeable in the open cells where
the morphological size gradient changes in a proximo-distal as well as a dorsolateral manner, until in the region of undoubted periblast (Fig. 12) very few
cytoplasmic structures can be seen among the yolk spheres. Lipid drops of the
type seen in older chick embryos (see Bellairs, 1958) are seldom visible in
these early stages. Similar observations on the intracellular yolk spheres have
been made by Emanuelsson & Von Mecklenburg (1968).
A remarkable feature of the investigation is our inability to identify nuclei in
Cleavage in the chick embryo
65
the early cleavage stages or in the periblast, despite the fact that we have
searched through very many paraffin and plastic sections by light microscopy,
as well as thin sections by electron microscopy. Furthermore, we have seen
mitotic figures only rarely in open cells (Fig. 2). We have, however, frequently
seen nuclei in closed cells (Fig. 13). These nuclei do not appear to possess
nucleoli but are sparsely packed with granules.
DISCUSSION
Many theories have been proposed to explain the mechanism of cleavage in
young embryos (reviewed by Wolpert, 1960). Most of these are based on the
study of echinoderm eggs which possess little yolk and so undergo total (holoblastic) cleavage. Large yolked eggs cleave in a different way (meroblastic
cleavage), forming 'open' cells at least in the early stages. The mechanism of
cell division in large yolked species must therefore differ in some respects from
that in echinoderm eggs, and theories based on echinoderm cleavage cannot be
applied to the chick without some modification. In general, most theories
include the following features: first, that the plane of cleavage is related to
the mitotic apparatus of the cell, but cleavage may nevertheless be initiated in
the absence of the mitotic figure. There is experimental evidence that in many
echinoderms the asters play an essential role (Rappaport, 1973), but we have
no evidence as to the part played by mitotic spindles or asters in the chick.
Second, there is a growth of new cell membrane, and there are many theories
as to how this is brought about (see Singer & Rothfield, 1973). One theory is
that cytoplasmic vesicles fuse with the existing cell membrane (Arnold, 1968,
1969). The presence of many cytoplasmic vesicles in the chick suggests that this
might be the mechanism, though other factors may be involved. For example,
in Xenopus there is an insertion of precursors at intervals along the existing
membrane (Bluemink & DeLaat, 1973) and it is possible that the same process
occurs in the chick.
A third feature of all cleaving embryos is the presence of microfilaments
running parallel to the surface membrane. These structures have been described
in many species, and by finding them in the chick, we confirm the observations
of Gipson (1974). They are also present in the embryos of cephalopods (Arnold
1968,1969,1971), which resemble the chick in undergoing meroblastic cleavage.
It has often been suggested that each furrow is produced by contraction of
the microfilaments (e.g. Arnold, 1968, 1971; Szollosi, 1970; Bluemink, 1973;
Perry, 1975) and there is some evidence that if the microfilaments are cut
(Arnold, 1971) furrow formation is impaired. The process may also be upset
by treating the embryo with cytochalasin B (Arnold & Williams-Arnold, 1974;
Bluemink & DeLaat, 1973), though the effect on the microfilaments may then
be an indirect one (Bluemink, 1971). It is generally considered that contraction
of the microfilaments leads to the folding and wrinkling of the cell membrane
66
R. BELLAIRS, F. W. LORENZ AND T. D U N L A P
which is so characteristic a feature not only of cleaving eggs (e.g. Bluemink,
1971; Monroy & Baccetti, 1975; Arnold, 1969) but also of many cells in
mitosis (e.g. Schroeder, 1968; Scott & Daniel, 1970). It may be noted, however,
that many of the projections, especially those in the necks of the pendulumshaped furrows, possess many more clearly arranged bands of microfilaments
than do the walls of the furrows, and this suggests that the projections are actively
motile structures rather than merely folds pulled passively into this shape. It
seems possible that they play a role in preventing the two sides of the furrow
becoming pulled widely apart. The tight junctions in the walls of the furrow
probably also aid in keeping the developing cells in contact. Although gap
junctions are present shortly after incubation of the laid egg begins (Bellairs
Breathrach & Gross, 1975), they do not appear to be present in the early
cleavage stages. There is extensive evidence in the literature that electrical
coupling of cells takes place via the gap junctions (Gilula, 1974) but it seems
unlikely that specialized contacts would be required for cell communication
when body cells are still present.
The furrow-base-body appears to be associated with the production or
accumulation of new membrane. Gipson (1974) has suggested that it is analogous to the midbody of cleaving holoblastic eggs. This seems unlikely, since
the furrow-base-body does not resemble the midbody found in the chick
embryo at later stages (Bellairs & Bancroft, 1975). Furthermore, a midbody
proper is formed at the point of constriction of two daughter cells, but there is
no comparable constriction in the cleavage stages of open cells. We suggest
that the furrow-base-body is related to the burrowing of the furrow into the
periblast, and may be associated with the change of direction which occurs
when the furrow begins to extend laterally to form the ventral border of the
cells. Arnold (1969) has described a specialized region at the base of the cleavage
furrows in the cephalopod, Loligo, which consists of a network of tubules,
though he was unable to ascribe a function to it. Both the cephalopod tubular
network and the avian furrow-base-body occupy comparable positions in
meroblastic eggs; each is a highly membranous structure. Nothing is known
about the role of the root region beneath the furrow-base-body, nor about the
processes which prevent the furrow from passing deeper and deeper toward the
yolk. It seems likely, however, that a furrow reaches its maximum depth when
the proportion of yolk to cytoplasm passes a certain threshold. A similar
situation appears to hold in Loligo (Arnold, 1971) though there the yolk and
the cytoplasm are sharply separated from one another.
The nature of the periblast and of the open cells in the chick has received
little attention in recent years. The term periblast is used to cover the regions
which lie both ventral and peripheral to the closed cells. The cytoplasmic components of the ventral periblast appear to be similar to those of the embryo
proper, though the relative proportions of yolk and cytoplasm gradually
change between the cells and the periblast and the yolk. In particular, small
Cleavage in the chick embryo
67
patches of glycogen granules and mitochondria are visible in the regions
which would otherwise be considered to be entirely non-cytoplasmic. The
specialized membranous structures illustrated in Fig. 9, however, have been
seen only in the new membrane (as discussed above). Significantly, we have
never been able to identify nuclei in either of the periblast regions whether by
light or by electron microscopy. Patterson (1910) also failed to find nuclei in
the periblast. Bekhtina (1960) reported on the basis of a light microscopic
study that the cleavage nuclei of the chick were found at the depths of the
furrows. We have not seen nuclei in these regions and in our opinion, Bekhtina
mistook the pendulum at the base of the furrow for a nucleus. Nuclei are
notoriously difficult to distinguish in light microscope sections of yolky material.
Similarly, we have failed to find nuclei at all in the open cells of the early
stages, though we have on one occasion found a mitotic spindle in an open cell
of a 12-cell embryo. Gipson (1974), though not discussing the problem, did not
illustrate or mention the nuclei, whilst Emanuelsson (1965) reported a failure
to find mitotic figures in embryos with less than 30 cells. Negative results being
notoriously unsatisfactory, the situation needs further analysis, but we would
like to emphasize that if it were established that nuclei were not present in all
the open cells or in the periblast, then traditional ideas on the mechanism of
expansion of the early cleavage embryo would need revision. It has sometimes
been suggested that amitosis can occur during cleavage in birds (Patterson,
1910; Emanuelsson, 1965), but we would be reluctant to accept this idea without
firmer evidence.
One possibility is that the cytoplasm of the early embryo begins to undergo
division as a result of activation by the many accessory sperm nuclei still
present at this stage, and that the open cells only later become colonized by
embryonic nuclei. Certainly, natural parthenogenesis in birds is now well
established (Olsen, 1966; Lorenz, 1975) and this implies a ready ability to
divide without a full chromosomal complement.
By contrast, nuclei are easily seen at later stages of cleavage when closed cells
are present (Fig. 13). At first they appear to lack nucleoli and these have not
been seen by us until about mid-cleavage. This supports the findings of Wylie
(1972) that RNA synthesis does not begin to take place until cleavage is well
advanced.
We wish to thank Mr R. Moss and Miss Doreen Bailey for their skilled technical help.
REFERENCES
J. M. (1968). An analysis of cleavage furrow formation in the egg of Loligo pealii.
Biol. Bull. 135, 413.
ARNOLD, J. M. (1969). Cleavage furrow formation in a telolecithal egg {Loligo pealii). 1.
Filaments in early furrow formation. /. Cell Biol. 41, 894-904.
ARNOLD, J. M. (1971). Cleavage furrow formation in a telolecithal egg {Loligo pealii). II.
Direct evidence for a contraction of the cleavage furrow. /. exp. Zool. 176, 73-86.
ARNOLD,
68
R. B E L L A I R S ,
F. W . L O R E N Z
A N D T. D U N L A P
ARNOLD, J. M. & WILLIAMS-ARNOLD, L. D. (1974). Cortical-nuclear interactions in cephalopod development: cytochalasin B effects on the informational pattern in the cell surface.
J. Embryol. exp. Morph. 31, 1-5.
BEKHTINA, V. G. (1960). The early stages of cleavage in the chick embryo. Arkiv. Anat.
Gistol. i Embryol. 38, 77-85. (In Russian: translation No. 5050 of National Lending
Library G.B.).
BELLAIRS, R. (1958). The conversion of yolk into cytoplasm in the chick blastoderm as shown
by electron microscopy. / . Embryol. exp. Morph. 6, 149-161.
BELLAIRS, R. & BANCROFT, M. (1975). Midbodies and beaded threads. Am. J. Anat. 143,
393-398.
BELLAIRS, R., HARKNESS, M. & HARKNESS, R. D. (1963). The vitelline membrane of the hen's
egg: a chemical and electron microscopical Study. / . Ultrastruct. Res. 8, 339-359.
BELLAIRS, R., BREATHNACH, A. S. & GROSS, M. (1975). Freeze-fracture replication of junc-
tional complexes in unincubated and incubated chick embryos. Cell Tiss. Res. 162, 235-252.
BLUEMINK, J. G. (1970). The first cleavage of the amphibian egg. An electron microscope
study of the onset of cytokinesis in the egg of Ambystoma mexicanum. J. Ultrastruct.
Res. 32, 142-166.
BLUEMINK, J. G. (1973). Effects of cytochalasin B on surface contractility and cell junction
formation during egg cleavage in Xenopus laevis. Cytobiologie 3, 176.
BLUEMINK, J. G. & DELAAT, S. W. (1973). New membrane formation during cytokinesis in
normal and cytochalasin B treated eggs of Xenopus laevis. I. Electron microscope observations. / . Cell Biol. 59, 89-108.
BURNSIDE, B. (1971). Microtubules and microfilaments in newt neurulation. Devi Biol. 26,
416-441.
EMANUELSSON, H. (1965). Cell multiplication in the chick blastoderm up to the time of laying.
Expl Cell Res. 39, 386-399.
EMANUELSSON, H. & VON MECKLENBURG, C. (1968). Localization of extra-nuclear DNA in
early chick blastoderm cells with electron microscopical autoradiography. Ark. Zool. 22,
(Sec. 2) 155-162.
GILULA, N. B. (1974). Junctions between cells. In Cell Communication (ed. Rody P. Cox).
New York : Wiley.
GIPSON, I. (1974). Electron microscopy of the early cleavage furrows in the chick blastodisc.
J. Ultrastruct. Res. 49, 331-347.
JENSEN, C. (1969). Ultrastructural changes in the avian vitelline membrane during embryonic
development. J. Embryol. exp. Morph. 21, 467-484.
KARNOVSKY, M. (1965). A formaldehyde-glutaraldehyde fixative of high osmolarity for use
in electron microscopy. / . Cell Biol. 27, 137 A.
LORENZ, F. W. (1975). Influence of age on avian gametes. In Aging Gametes (éd. R. J. Blandau). Basel: S. Karger.
MONROY, A. & BACCETTI, B. (1975). Morphological changes of the surface of the egg of
Xenopus laevis in the course of development. I. Fertilization and early cleavage. / . Ultrastruct. Res. 50, 131-142.
OLSEN, M. W. (1942). Maturation, fertilization and early cleavage in the hen's egg. / . Morph.
70, 513-533.
OLSEN, M. W. (1966). Frequency of parthenogenesis in chicken eggs. / . Hered. 57, 23-25.
PATTERSON, J. (1910). Early development of the hen's egg. / . Morph. 21, 101-134.
PERRY, M. M. (1975). Microfilaments in the external surface layer of the early amphibian
embryo. / . Embryol. exp. Morph. 33, 127-146.
RAPPAPORT, R. (1973). Cleavage furrow establishment - a preliminary to cylindrical shape
change. Am. Zool. 13, 941-948.
ROMANOFF, A. L. (1960). The Avian Embryo. New York: Macmillan.
RUDDELL, C. L. (1971). Embedding media for 1-2 micron sectioning. 3. Hydroxyethyl
methacrylate-benzoyl peroxide activated with pyridine. Stain Technology 46, 77-83.
SCHROEDER, T. E. (1968). Cytokinesis: filaments in the cleavage furrow. Expl Cell Res. 53,
272-276.
Cleavage in the chick embryo
69
SCHROEDER, T. E. (1972). The contractile ring. IT. Determining its brief existence, volumetric
changes and vital role in cleaving Arbacia eggs. / . Cell Biol. 53, 419-434.
SCOTT, D. G. & DANIEL, C. W. (1970). Filaments in the division furrow of mouse mammary
cells. / . Cell Biol. 45, 461-466.
SINGER, S. J. & ROTHFIELD, L. I. (1973). Synthesis and turnover of cell membranes. Neurosciences Research Program Bulletin 11, 1-86.
SZOLLOSI, D. (1970). Cortical cytoplasmic filaments of cleaving eggs: a structural element
corresponding to the contractile ring. / . Cell Biol. 44, 192-210.
WOLPERT, L. (1960). The mechanics and mechanisms of cleavage. Rev. Cytol. 10, 163-216.
WYLIE, C. C. (1972). The appearance and quantification of cytoplasmic ribonucleic acid in
the early chick embryo. / . Embryol. exp. Morph. 28, 367-384.
(Received 22 February 1977, revised 1 June 1977)