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Biological lectures delivered at the Marine biological laboratory of Wood's Hole ... 1890-[1899] (Volume 6) online

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into the vesicular form of the ordinary resting nucleus at the
same time, and keep step throughout the process. This indi-
cates a new general condition of the egg-cytoplasm; for, while
we might explain the enlargement of the egg-nucleus alone as
part of the usual sequence of mitosis, due to purely localized
conditions of the egg-substance, we can only explain the effect
simultaneously produced on both nuclei by the assumption
that the entire egg-cytoplasm is entering on a new phase of its

The first movements of the germ-nuclei begin after each has
enlarged considerably. In its early movements the egg-nucleus
is preceded by the sphere-substance, which moves towards the



centre of the egg, and always towards the side opposite to that
in which the sperm-nucleus lies (Fig. 15). Thus the egg-nucleus
first moves away from the sperm-nucleus, which immediately
takes up its march in an amoeboid manner towards the centre
of the egg, and in the general direction of the egg-nucleus. I
may finish at once with the movements of the germ-nuclei by

saying that they ultimately
come together in the cen-
tre of the egg. Whether
or not the first movement
of the egg-nucleus away
from the sperm-nucleus is
passively brought about by
the sphere-substance, the
behavior of the germ-nuclei
shows that their meeting
is not entirely due to mu-
tual attraction, but partly,
at least, to a common tend-
ency to seek a dynamic cen-
tre of the egg.

To. return to the sphere-
substance : its invariable migration towards the side of the
egg opposite to that in which the sperm-nucleus lies indicates
one of two things, either that the sperm-nucleus has driven
it away, or else that it is moving along lines of orientation of
the egg-substance. To suppose that the minute sperm-nu-
cleus could exercise such an effect seems impossible, and the
corollary of the second alternative is that the sperm-nucleus
has occupied throughout its entire resting period a definite
position within the egg, which can only be explained on the
assumption of a definite orientation of the egg at the time of
fertilization, not only polar, but also corresponding to one of
the other chief axes of the embryo. The first cleavage-plane
passes very nearly through the point at which the sperm-
nucleus has been resting during the maturation.

During its migration towards one side of the egg, the sphere-
substance has undergone a remarkable change of form; it has

FIG. 15. Unto. Early movements of the germ-
nuclei and of the sphere-substance.



FIG. 16. Unto. Outline of section in the direction
of the line ruled across Fig. 15 ; to show the elon-
gation of the sphere-substance.

elongated greatly in a Jiorizontal direction, at rigJit angles to
the line uniting the germ-nuclei (Fig. i6)> and has tJius marked
out the direction of the first
cleavage-spindle. Now this
elongation has begun, and
the plane of division is indi-
cated before the germ-nuclei
have met.

It might very readily be
assumed that the plane of
the first cleavage is deter-
mined by the copulation-
path of the germ-nuclei, as
is stated to be the case in
the ova of some other ani-
mals (frog, Roux ; Toxo-
pneustes, E. B. Wilson).
But whoever should take
this position for Unio would have to explain how it happens
that the sphere-substance elongates in the plane of the first

cleavage-spindle before the
germ-nuclei come together.
It would be necessary, I
believe, to assume that the
distant sperm-nucleus ex-
ercises an influence on the
direction of migration and
of elongation of the sphere-
substance in the first cleav-
age, but that, in the next
division, the sphere-sub-
stance acts independently;
and this assumption is ab-
surd on the face of it.

The first cleavage-spindle
forms in the centre of the
egg (Fig. 17), in the plane already indicated by the elongation
of the sphere-substance. At first there is a single very minute

FIG. 17. Unio. First cleavage-spindle forming. Basi-
chromatin granules near the ends of the spindle ; oxy-
chromatin granules on the spindle-fibres.



centrosome at each pole, almost in contact with the nuclear
membrane, then a group of centrosome granules imbedded in
a ground substance ; then comes a clearing of the centre, accom-
panied by peripheral arrangement and subsequent fusion of all
the centrosome granules, excepting one, which remains in the
centre. Thus is established a hollow sphere (" centrosome ")
with an included centrosome ("centriole") (Fig. 18). During
the early stages of the metamorphosis of the centrosomes,
chromatin granules from the nucleus are found in their im-
mediate neighborhood, and, as the spindle forms, they become

closely pressed to the
centrosomes (Fig. 17).
The entire spindle then
moves directly along the
prolongation of its axis,
thus parallel to the di-
rection of elongation of
the sphere-substance,
to one side of the egg
(Fig. 1 8), until the cen-
trosome of one end
comes almost in con-
tact with the peripheral
layer of protoplasm.
Then comes the meta-
phase (Fig. 18), and, in
a late stage of the anaphase, the sphere begins to enlarge,
and a reticulum (or vesicular substance) with nodal micro-
somes (Fig. 19) develops in it in place of the single centro-

The egg now begins to elongate at right angles to the plane
of division, and the entire spindle, including the enlarged
spheres, shifts towards the centre of the egg a short distance,
and finally comes to rest (Fig. 19). The spindle acts as though
oscillating through a point of equilibrium, at first shifting too
far in one direction (Fig. 18), then swinging back (Fig. 19), and
possibly undergoing lesser shiftings before coming to rest for
the first cleavage.

FIG. 18. Migration of the spindle to one side of the egg ;
metaphase ; for the sake of clearness only a few of the
chromosomes were drawn in.


To what extent this centre of equilibrium is determined by
the primary orientation of the cytoplasm, and how much by the
secondary distribution of the sphere-substance, it is impossible
to say ; but the orientation of the cytoplasm is, in either event,
the primary factor, inasmuch as it determines the distribution
of the sphere-substance.

The first cleavage-furrow now forms rapidly, and, as it forms,
the spheres undergo an enlargement, migration, and change
of shape analogous to that preceding the first cleavage. We
shall follow it only in the larger cell. The sphere-substance

FIG. 19. Unto. Secondary shifting of the first cleavage-spindle. Beginning of
growth of the sphere-substance.

in this cell elongates during the early stages of reconstitution of
the nucleus in a horizontal direction parallel to the axis later
taken by the second cleavage-spindle in this cell (Fig. 20). At
the same time the entire cell elongates in the same direction
and becomes slightly constricted in a plane and position corre-
sponding precisely with the next cleavage-plane ; this, before
the nucleus is reconstituted, or has moved away from the neigh-
borhood of the first cleavage-wall. It is as though the cytoplasm
were making an attempt at division, which is rendered abortive
by the stage of development of the nucleus. Later both cells
round off and then become applied together, and the second
cleavage-spindle forms and moves into the position of the division
already indicated.

6 4


Conklin (4) was the first to call attention to the possible
importance of the sphere-substance in the cleavage of the egg.
He summarizes his results on the egg of Crepidula thus : " After
the first two cleavages the sphere-substance is differently dis-
tributed to the different cells, the entire sphere-substance of
one generation always going into those cells of the next genera-
tion which lie nearest the animal pole. This differential distri-
bution of the spheres has been followed through every cleavage
up to the twenty-four-cell stage. As the form of the cleavage

is perfectly constant, it
follows that the sphere-
substance of any genera-
tion goes into certain
definite cells which have a
perfectly constant origin
and destiny. This differ-
ential distribution of the
spheres is not caused by
their specific weight, since

their movements are the

t IG. 20. Unto, birst cleavage ; the elongation of the
sphere-substance in the larger cell and of the cell
itself marks the plane of the second cleavage. In a ,

slightly later stage there is a well-marked constriction Same in WnatCVCr pOSltlOn
across the cell in the position of the future second flg Cell ITiaV be DlaCed It
cleavage-spindle in this cell.

seems to be the result of

a form of polarity which, like that of the egg itself, is not
the result of gravity.

" The centrosomes do not, apparently, arise from the sphere-
substance of the previous division, but some distance from it,
and the sphere-substance never divides, but each sphere ulti-
mately grows ragged at its periphery and gradually fades out
into the general cytoplasm.

"The differential distribution of the spheres and their
subsequent conversion into cytoplasm suggest that they
may be important factors in the differentiation of cleav-
age cells, and if further investigation should establish the
fact that they are, in part, composed of the oxychro-
matin of the nucleus, it would furnish a basis in fact for
certain well-known speculations of de Vries, Weismann,
and Roux."


Thus, putting Conklin's results 1 on the movements of the
sphere-substance and my own together, it would appear that,
whereas in the first two cleavages this substance is divided
between the cells in proportion to their size, in the formation
of the generations of ectomeres the substance enters special
cells. This would coincide very closely with the differential
value of the cleavages, the first four cells possessing ectoblastic,
entoblastic, and, in part, mesoblastic materials, while the three
subsequent divisions of the macromeres separate ectoblastic
portions. This tends, it seems to me, to strengthen Conklin's
conclusion that the sphere-substance may be an important
factor in the differentiation of cells.

Finally, I do not believe that the process of nuclear or cell-
division is ever in itself an act of differentiation. That it is
not, in certain cases at any rate, is shown beyond the possibility
of any doubt by examples of non-determinate cleavage, such as
that of the fish-egg, in which the cleavage-planes bear no con-
stant relation to each other or to the embryonic parts, and,
still more strikingly, in the case of ciliate Infusoria, where the
entire process of development takes place without any cell-
division. If my observations are correctly interpreted in
what has preceded, the essential process in early embryo-
formation proceeds on the basis of a definite orientation and
organization of the egg-substance, carried forward and elab-
orated by certain intercellular processes, in which the produc-
tion of special substances which have been acted on by the
chromatin may play an important role. Now the distribution
of these substances is not dependent on cell-division, though by
this they may be isolated in separate cells ; but it is conceiv-
able that the cleavage-planes may, so to speak, ignore the lines
of orientation of the egg and of distribution of specific parts of
it ; thus it may be that determinism in the cleavage is no
measure of the degree of organization of the egg, as Whitman
has so ably argued.

It is quite possible that there is no sharp distinction between

1 In Crepidiila, apparently, the substance of the spheres is not divided in the
second cleavage, but passes into special cells. See lecture by Dr. Conklin in this


determinate, and indeterminate cleavage, and that one grades
into the other, the apparent difference being due to insufficient
knowledge, as Conklin and Eisig (5) have suggested. But there
can be no doubt in the mind of any one as to the existence of
a very real difference between determinate and indeterminate
types of cleavage, who has compared, for instance, the cleavage
of the egg of an annelid, possessing a perfectly definite and
unvarying mode of cleavage and cell-lineage of organs, with
that of a fish, in which slight alterations of the external condi-
tions cause the very greatest variations in cleavage, so that
often the cells of two eggs of the same species cannot be
homologized, and no definite cell-lineage of organs exists. The
explanation of this difference, it seems to me, is a prospective
one. It is dependent, I believe, on the actual number of cells
composing the embryo at the time that the first larval or
embryonic organs come into service. In other words, I would
think of determinate cleavage as an adaptation to a condition
in which the functional activity of organs begins with a rela-
tively small number of cells, and in which, therefore, each cell
is of special importance.

August, 1898.



1. CHILD, C. M. A Preliminary Account of the Cleavage of Arenicola

cristata, with Remarks on the Mosaic Theory. Zool. Bulletin. Vol. i.

2. CONKLIN, E. G. Cleavage and Differentiation. Biological Lectures.

Delivered at the Marine Biological Laboratory. 1896, 1897.

3. CONKLIN, E. G. The Embryology of Crepidula. Journ. of Morph.

Vol. xiii. 1897.

4. CONKLIN, E. G. See summary of a paper delivered before the Ameri-

can Society of Morphologists at the Ithaca meeting, December, 1897.
Science. March, 1898.

5. EISIG, HUGO. Zur Entwicklungsgeschichte der Capitelliden. Mitth.

aus der Zool. Station zu NeapeL Bd. xiii. 1898.

6. LILLIE, FRANK R. Preliminary Account of the Embryology of Unio

complanata. Journ. of Morph. Vol. viii. 1893.

7. LILLIE, FRANK R. Embryology of the Unionidae. Journ. of Morph.

Vol. x. 1895.

8. LILLIE, FRANK R. Centrosome and Sphere in the Egg of Unio. Zool.

Bulletin. Vol. i. 1898.

9. MEAD, A. D. The Early Development of Marine Annelids. Journ.

of Morph. Vol. xiii. 1897.

10. TREADWELL, A. D. The Cell-Lineage of Podarke obscura. Prelimi-

nary Communication. Zool. Bulletin. Vol. i, No. 4. 1897.

11. WHITMAN, C. O. The Embryology of Clepsine. Quar. Journ. of

Micr. Science. Vol. xviii. 1878.

12. WHITMAN, C. O. The Inadequacy of the Cell-Theory of Development.

Biological Lectures. Delivered at the Marine Biological Laboratory.

13. WILSON, E. B. The Cell- Lineage of Nereis. Journ. of Morph.

Vol. vi. 1892.

14. WILSON, E. B. Considerations on Cell-Lineage and Ancestral Remi-

niscence. Annals of the New York Academy of Sciences. Vol. xi,
No. i. 1808.




THE fundamental problems of development and inheritance
are in the last analysis questions of differentiation. Develop-
ment is progressive differentiation coordinated as to time and
place ; hereditary likeness consists in the repetition by the off-
spring, at certain stages of its life cycle, of definite differentia-
tions of the parent ; and hereditary unlikeness, or variation, is
a modification of these differentiations either as to their char-
acter or as to the time of their appearance. The phenomena of
differentiation are therefore of the greatest interest, and their
causes one of the most important problems of biology.

In many respects the simplest and yet most important phe-
nomena of differentiation occur in the early stages of develop-
ment, while the later differentiations of tissues and organs are
more complicated and less general in character. The polarity
of the egg is one of the earliest differentiations of the de-
veloping organism ; it consists not only in the aggregation of
yolk at one pole and of protoplasm at the other, but also in
the establishment of certain structural peculiarities which in
most cases determine the position and direction of the two
maturation spindles and cause the first two cleavage planes to
pass through the polar axis of the egg. Further, it has a defi-
nite prospective significance, since it is probable that in all ani-
mals it determines the ectodermal and endodermal poles of the
embryo, while in most cases the animal and vegetal poles of
the egg give rise, respectively, to the apical and oral poles of the



larva or adult. This polar differentiation may appear at an
early stage in the ovarian egg, or it may be delayed until after
fertilization. In some cases (insects and cephalopods) not only
the primary axis of the egg, but all the axes and regions of the
future animal are marked out in the ovarian egg ; in other cases
these axes, except the primary one, are not apparent until the
end of cleavage or even after gastrulation.

In the cleavage of the egg, differentiations occur in a
remarkable degree in certain cases, while they appear to be
absent in others. Typically, cell divisions are rhythmical, alter-
nating, quantitatively and qualitatively equal, and consequently
non-differential. The differentiations of cleavage cells are due
to departures from this typical condition in one or more partic-
ulars. In certain animals these departures are very notable, the
cleavages being from the first non-alternating, non-rythmical,
unequal, and qualitatively dissimilar. These differentiations of
cleavage have also a far-reaching prospective significance, since
in certain cases (polyclades, nematodes, rotifers, annelids, mol-
lusks) the principal axes and body regions of the future animal
are marked out by the cleavage planes, and the building mate-
rial of entire organs is segregated into a single cell or group
of cells.

I have repeatedly observed these unequal, non-alternating,
and non-rhythmical cleavages with the feeling that the causes
of such differentiation were almost within sight, and with the
conviction that continued study could not fail to reveal them ;
and yet it must be said that these causes, which seem so near
at hand, generally elude one's grasp. Unequal cleavage is due
to the eccentricity of the mitotic spindle, but why is the spindle
eccentric? Non-alternating cleavage is due to the spindles hav-
ing approximately the same direction during successive cleav-
ages ; but why do the spindles take this peculiar position?
Non-rhythmical cleavage can be referred only to differences in
the substance of cells, but how these differences operate can-
not in most cases be explained. It is hopeless to look for an
answer to the last question that may be asked concerning the
cause of these or of any other phenomena; all that can reason-
ably be expected is that the many different phenomena and


factors of development may some time be harmonized and uni-
fied by the discovery of some common causal principles.


Of the many factors of differentiation which have been pro-
posed within recent years, the majority are in the nature of
simple physical causes. Thus the polar differentiation of the
egg has been attributed to differences in the specific weight of
protoplasm and yolk ; e.g., Hertwig 1 says : " Polar differentiation
consists in this, that the lighter protoplasm collects at one pole
and the heavier yolk substance at the other." Experiments on
the frog's egg led Pfliiger, Born, and Schultze to essentially the
same conclusion.

The differentiations of cleavage are commonly attributed to
factors of a similar character ; thus the direction of cell division
is said to be due to the fact that the mitotic spindle lies in the
direction of least resistance (Pfluger) or in the longest axis of
the protoplasmic mass (Hertwig) ; the shape and position of
cells, and consequently to a certain extent the direction of divi-
sion, are said to be due to the rectangular intersection of cleav-
age planes (Sachs), or to the principle of smallest surfaces
(Berthold) ; the rate of division and the relative size of daughter-
cells are commonly attributed to the mechanical influences of
inert yolk (Balfour, Hertwig). Finally, the differences in the
quality of cells have been referred to intercellular reactions
(Hertwig, Wilson),* which, in some cases at least, are regarded
as of a physical rather than of a physiological character.

It has been repeatedly shown that none of these principles
are of universal application, and it seems doubtful whether in
any case they are the real causes of the phenomena in ques-
tion. How little gravity has to do with polar differentiation
is well shown in the eggs of many gasteropods where the eggs
lie in all possible positions in the egg capsules with their pri-
mary axes turned in all possible directions, and yet the polar
differentiation occurs as perfectly and as rapidly in one position
as in another. That gravity can have nothing directly to do

i The Cell, p. 215.


with this differentiation is further shown by the fact that the
protoplasm which lies at the animal pole in early stages of
cleavage lies at the vegetal pole of the macromeres in later
stages ; while the yolk, which originally lay at the vegetal pole,
comes to lie at the animal pole of the macromeres ; in short,
the polarity of the macromeres is reversed during cleavage, and
this always happens in the same way, irrespective of the posi-
tions in which the eggs may lie.

How far these mechanical principles fall short of explaining
the differentiations of cleavage has been pointed out by Mead
('94), Kofoid ('94), Lillie ('95), McMurrich ('95), Wheeler ('95),
Zur Strassen ('95), Castle ('96), Jennings ('96), Conklin ('92, '94,
'97), and many others. Much has been said, and very justly, of
the difficulty of explaining by simple mechanical causes non-
alternating and unequal cleavages, and yet it ought not to be
forgotten that the causes of alternating and equal cleavages
have not been given. To say merely that cell divisions are typi-
cally alternating and equal affords no causal explanation. If we
knew why cleavages are usually alternating and equal, we should
probably be able to explain why they are sometimes neither.
How little alternating cleavage has to do with the mere diver-
gence of centrosomes in planes successively at right angles to
one another will be apparent further on, where we shall see
that the centrosomes do not preserve their original positions
in the daughter-cells, and that the direction of their divergence
bears no constant relation to the direction of the cell division ;
and how little yolk has to do with the inequality of division is
shown by the fact that in the formation of the polar bodies we
have two very unequal divisions of the egg, while the first and
second cleavages, are frequently equal ; the first, second, and
third divisions of the macromeres thus formed are usually very
unequal, while the fourth and fifth divisions of these cells are
again nearly equal ; finally, among the micromeres, which fre-
quently contain no yolk, the early divisions are more frequently
unequal than equal. These same considerations apply in the
main to the rate of division ; in most cases of determinate
cleavage it bears no constant relation to the presence or absence
of yolk in the cells. In every case of determinate cleavage


these so-called "laws" are repeatedly set at naught, and this
fact has led those who have studied such cleavage to the con-
clusion that no simple physical explanation of these processes
of differentiation is possible, and that their cause must be found
in the structure of the protoplasm or in some physiological


The causes of differentiation are frequently referred to the
structure of the germinal protoplasm, as if this were a sat-
isfactory explanation. But to say that polarity and differen-
tiations of cleavage are due to the constitution of the egg is
merely a form of words which means little or nothing. In the
same way it might be said that all the multifarious aspects of
the universe are the results of the constitution of matter. To
refer vital phenomena to the constitution of protoplasm and
there to rest is merely to juggle with words. The phenomena
in question must be analyzed and their immediate causes deter-
mined step by step before any " explanation " can be thought of.
If, then, those who attempt to explain differentiation as the re-

Online LibraryMass.) Marine Biological Laboratory (Woods HoleBiological lectures delivered at the Marine biological laboratory of Wood's Hole ... 1890-[1899] (Volume 6) → online text (page 6 of 29)