KARYOKINESIS
In 1873, Hermann Fol, writing of the eggs of Geryonia, thus describes the
phenomenon of karyokinesis: "On either side of the residue of the nucleus
there appears a concentration of plasma, thus forming two perfectly regular
star-like figures, whose rays are straight lines of granulations. There are
other curved rays which pass from one star or centre of attraction to the
other. The whole figure is extraordinarily distinct, recalling in a
striking manner the arrangement of iron filings surrounding the poles of a
magnet. Sachs' theory is that the division of the nucleus is caused by
centres of attraction, and I agree with him, not on theoretical grounds,
but because I have actually seen these centres of attraction."
Since the discovery of Hermann Fol, a great number of explanations have
been given, all of them theoretical, to account for the figures and
phenomena of karyokinesis. Many of these so-called explanations are
mechanical, while others invoke the aid of magnetism or electricity to
account for the resemblance of the figures of karyokinesis to the magnetic
or electric phantom or spectre. Among the authors who have dealt with this
question we may mention Hartog of Cork, Gallardo of Buenos Ayres, and
Rhumbler of Goettingen.
In 1904 I presented to the Grenoble Congress, and in 1906 to the Lyons
Congress, a series of photographs and preparations of experimental
karyokinesis. I showed how, in a solution analogous to that found in the
natural cell, the simple processes of liquid diffusion, without the
intervention of magnetism or electricity, may reproduce with perfect
accuracy and in their normal sequence the whole of the movements and {90}
figures which characterize the phenomenon of karyokinesis. This experiment
consists not merely in the production of a certain figure, such as is
obtained in the magnetic spectre, but in the reproduction of the movement
itself, and of all the successive forms which are seen in the natural
phenomenon. These are evolved before the eyes of the spectator in their
regular order and sequence.
I may here reproduce the text of my communication at Grenoble: "Until I
introduced the conception of a field of diffusion, there was no proper
means of studying the phenomena of diffusion, which obey the laws of a
field of force as expounded by Faraday. Moreover, no one suspected the
possibility of reproducing by liquid diffusion a spectre analogous to the
electro-magnetic phantom. Guided by this theory of a diffusion field of
force, I have been able to reproduce experimentally the figures of
karyokinesis by simple diffusion. With regard to the achromatin spindle,
Professor Hartog has shown that the two poles of the spindle are of the
same sign, and not of opposite signs as was at first supposed. In the
process of karyokinesis the two centrosomes, _i.e._ the two poles of the
achromatin spindle, repel one another. They must therefore be poles of the
same sign. An electric or magnetic spectre showing a spindle between two
poles of the same sign is unknown; such a thing would appear to be an
absolute impossibility. What is impossible in electricity and magnetism,
however, is quite possible in the artificial diffusion field; we can here
have a spindle between two poles which repel one another--that is, between
poles of the same sign. Fig. 31 is a photograph of such a spindle produced
by diffusion. On either side are two poles of concentration, which
represent the centrosomes, each pole being surrounded by a star-like
radiation. These poles being alike, repel one another. In the preparation
one may see the distance between the two poles slowly increase, the poles
gradually separating from one another just as do the centrosomes of an ovum
during karyokinesis. This preparation, then, which is produced entirely by
diffusion, presents a perfect resemblance to the achromatin spindle in
karyokinesis. {91}
[Illustration: FIG. 31.--Diffusion figure representing karyokinesis.
Achromatin spindle between two similar poles of concentration.]
"The spindle of which we give a photograph in Fig. 31 was made by placing
in salt water a drop of the same solution pigmented with blood or Indian
ink, and placing on either side of this central drop a hypertonic drop of
salt solution more lightly coloured. After diffusion had gone on for some
minutes, we obtained the figure which we have photographed. I would draw
your attention to the equatorial plane, which shows that the spindle is not
formed by lines of force passing from one pole to the other, as would be
the case between two poles of contrary sign, but by two forces acting in
opposite directions. On either side the pigment of the central drop has
been drawn towards the hypertonic centre nearest to it. In the median line,
however, the pigment is attracted in opposite directions by equal forces,
and therefore remains undisturbed, marking the position of the equatorial
plane. This observation applies equally to the equatorial plane in natural
karyokinesis, whose existence is thus readily explained.
"It is hardly necessary to insist on the fact that liquid preparations like
these are of extreme delicacy and sensitiveness, and require for their
production, and still more for their photography, the greatest care and
skill, which can only be acquired by long practice. {92}
"We are able to produce by diffusion not only the achromatin spindle, but
also the segmentation of the chromatin, and the division of the nucleus. If
in the saline solution we place a coloured isotonic drop between two
coloured hypertonic drops, all the figures and movements of karyokinesis
appear successively in their due order. The central drop, representing the
nucleus between the two lateral drops or centrosomes, first becomes
granular. Next we see what appears to be a rolled-up ribbon analogous to
the chromatin band, which soon breaks into fragments analogous to the
chromosomes. These arrange themselves around, and are gradually attracted
towards the centrosomes, where they accumulate to form two pigmented
nuclear masses. A partition then makes its appearance in the median line,
and this partition becomes continuous with the boundary of the spheres
around the centrosomes. Finally we have two cells in juxtaposition, each
with its nucleus, its protoplasm, and its enveloping membrane. I have been
able to photograph these successive stages of the segmentation of the
chromatin just as I have those of the achromatin spindle" (Fig. 32).
[Illustration: FIG. 32.--Four successive stages in the production of
artificial karyokinesis by diffusion.]
This memoir, written in 1904, clearly asserts the homopolarity of the
centrosomes, and shows that the nuclear division is the result of a bipolar
action, two poles of the same sign exerting their influence on opposite
sides of the nucleus. It also emphasizes the important fact that diffusion,
{93} and as far as we know diffusion alone, is able to produce a spindle
between homologous poles.
A glance at the photograph is enough to show that the spindle is formed
between poles of the same sign. The lines of diffusion radiate from one
centre and converge towards the other centre in curves, giving the double
convergence characteristic of a spindle. The central drop merely supplies
the necessary material, and should have a concentration but slightly less
than that of the plasma, so as not to set up its own lines of diffusion.
The photograph shows clearly that the rays of the spindle traverse the
equator without any break. It has been objected that these lines form not
so much a spindle as two hemi-spindles, but it is clear that these two
hemi-spindles are continuous and form a single sheaf of rays uniting the
two poles of concentration. This is a phenomenon entirely unknown in the
magnetic or electric fields, where two poles of the same sign, one on
either side of a pole of the contrary sign, give two separate spindles. In
a magnetic field it is impossible to make the lines emanating from one pole
converge, except to a pole of opposite sign. Hence if we admit the
homopolarity of the centrosomes, we must also admit that diffusion is the
_vera causa_ of karyokinesis, since, as I showed at the Grenoble Congress
in 1904, diffusion and diffusion alone is capable of producing a spindle
between two poles of the same sign.
_Nuclear Division._--In order to reproduce artificially the phenomena
attending the division of the nucleus, we may proceed as follows. We cover
a perfectly horizontal glass plate with a semi-saturated solution of
potassium nitrate to represent the cytoplasm of the cell. The nucleus in
the centre is reproduced by a drop of the same solution coloured by a trace
of Indian ink, the solid particles of which will represent the chromatin
granules of the nucleus. The addition of the Indian ink will have slightly
lowered the concentration of the central drop, and this is in accordance
with nature, since the osmotic pressure of the nucleus is somewhat less
than that of the plasma. We next place on either side of the drop which
represents the {94} nucleus a coloured drop of solution more concentrated
than the cytoplasm solution. The particles of Indian ink in the central
drop arrange themselves in a long coloured ribbon, apparently rolled up in
a coil, the edges of the ribbon having a beaded appearance. After a short
time the ribbon loses its beaded appearance and becomes smooth, with a
double outline, as is shown in A, Fig. 32. This coil or skein of ribbon
subsequently divides, forming a nuclear spindle, while the chromatin
substance collects together in the equatorial plane as in B, Fig. 32.
A more advanced stage of the nuclear division is shown at C, Fig. 32, where
the chromatin bands of artificial chromosomes are grouped in two conical
sheafs converging towards the two centrosomes. For some considerable time
these conical bundles remain united by fine filaments, the last vestiges of
the nuclear spindle. The final stage is that of two artificial cells in
juxtaposition, whose nuclei are formed by the original centrosomes
augmented by the chromatin bands or chromosomes (Fig. 32, D).
[Illustration: FIG. 33.--Equatorial crown produced by diffusion.]
The resemblance of these successive phenomena to those of natural
karyokinesis is of the closest. The experiment shows that diffusion is
quite sufficient to produce organic karyokinesis, and that the only
physical force required is that of osmotic pressure. If in the cytoplasm of
a cell there are two points of molecular concentration greater than that of
the general mass, the nucleus must necessarily divide with all the
phenomena which accompany karyokinesis. In nature these two centres of
positive concentration are introduced into the protoplasm of the cell by
fecundation--that is, by the entrance of the centrosomes of the sperm cell.
In certain abnormal cases the concentration may be produced in the cell
itself by the formation of two centres of catabolism or molecular
disintegration, since, as we have seen, molecular disintegration raises the
osmotic pressure. This phenomenon, namely the production of karyokinesis
from centres of catabolism, may account for the abnormal karyokinesis of
cancer cells and the like. The subject is one which would well repay
further investigation. {95}
[Illustration: FIG. 34.--A triaster produced by diffusion.]
It has been found in our experiments that in order to obtain the regular
division of the artificial nucleus represented by the intermediary drop,
the latter must have an osmotic pressure slightly below that of the plasma.
This leads to the supposition that a similar condition must obtain in the
natural cell. It may be noticed, moreover, that the grains of pigment
follow the direction of the flow of water, being carried along by the
stream. This would appear to show that the nucleus of a natural cell has
also a molecular concentration less than that of the plasma--a result
either of dehydration of the plasma, or of some diminution in the molecular
concentration of the nucleus.
Other phenomena of karyokinesis may also be closely imitated by diffusion.
For instance, in the diffusion preparation we notice at each extremity of
the equator a V-shaped figure with its apex towards the centre,
corresponding exactly to what in natural karyokinesis is called the
equatorial crown.
We may also produce diffusion figures of abnormal karyokinesis. Fig. 34
represents such a form, a triaster produced by diffusion.
Artificial karyokinesis may also be produced by hypotonic poles of
concentration--that is to say, when the central drop representing the ovum
is positive and the lateral drops representing the centrosomes are negative
with respect to the plasma. In this case, however, the resemblance to
natural karyokinesis is less perfect. {96}
Without attaching to it an importance which is not warranted by
experimental results, it is interesting to note that we have here two
methods of fertilization, hypertonic and hypotonic, _i.e._ by centrosomes
of greater concentration and by centrosomes of less concentration than that
of the plasma of the ovum, and that we have in nature two corresponding
results, viz. two different sexes. It is possible that we have in these two
methods of producing nuclear division the secret of the difference of sex.
* * * * *
{97}