On Growth and Form

It has been shewn that ordinary nuclei, isolated in a living or fresh state, easily flow together; and this fact is enough to suggest that they are aggregations of a particular substance rather than bodies deserving the name of particular organs. It is by reason of the same tendency to confluence or aggregation of particles that the ordinary nucleus is itself formed, until the imposition of a new force leads to its disruption. Apart from that invisible or ultra-microscopic heterogeneity which is inseparable from our notion of a “colloid,” there is a visible heterogeneity of structure within both the nucleus and the outer protoplasm. The former, for instance, contains a rounded nucleolus or “germinal spot,” certain conspicuous granules or strands of the peculiar substance called chromatin, and a coarse meshwork of a protoplasmic material known as “linin” or achromatin; the outer protoplasm, or cytoplasm, is generally believed to consist throughout of a sponge-work, or rather alveolar meshwork, of more and less fluid substances; and lastly, there are generally to be detected one or more very minute bodies, usually in the cytoplasm, sometimes within the nucleus, known as the centrosome or centrosomes.

* The elongated or curved ‘‘macronucleus” of an Infusorian is to be looked upon as a single mass of chromatin, rather than as an aggregation of particles in a fluid drop, as in the case described. It has a shape of its own, in which ordinary surface-tension plays a very subordinate part. the cell, meaning thereby a symmetry of visible structure about a particular axis. For instance, whenever we can recognise in a cell both a nucleus and a centrosome, we may consider a line drawn through the two as the morphological axis of polarity : in an epithelium cell, it is obvious that the cell is morphologically symmetrical about a median axis passing from its free surface to its attached base. Again, by an extension of the term “polarity,” as is customary in dynamics, we may have a “radial” polarity, between centre and periphery; and lastly, we may have several apparently independent centres of polarity within the single cell. Only in cells of quite irregular, or amoeboid form, do we fail to recognise a definite and symmetrical “polarity.” The morphological “polarity” is accompanied by, and is but the outward expression (or part of it) of a true dynamical polarity, or distribution of forces; and the “lines of force” are rendered visible by concatenation of particles of matter, such as come under the influence of the forces in action.

When the lines of force stream inwards from the periphery towards a point in the interior of the cell, the particles susceptible of attraction either crowd towards the surface of the cell, or, when retarded by friction, are seen forming lines or “‘fibrillae’’ which radiate outwards from the centre and constitute a so-called “aster.” In the cells of columnar or ciliated epithelium, where the sides of the cell are symmetrically disposed to their neighbours but the free and attached surfaces are very diverse from one another in their external relations, it is these latter surfaces which constitute the opposite poles; and in accordance with the parallel lines of force so set up, we very frequently see parallel lines of granules which have ranged themselves perpendicularly to the free surface of the cell (cf. fig. 97).

A simple manifestation of “polarity” may be well illustrated by the phenomenon of diffusion, where we may conceive, and may automatically reproduce, a “field of force,” with its poles and visible lines of equipotential, very much as in Faraday’s conception of the field of force of a magnetic system. Thus, in one of Leduc’s experiments*, if we spread a layer of salt solution over a level plate of glass, and let fall into the middle of it a drop of indian ink, or of blood, we shall find the coloured particles travelling outwards from the central “pole of concentration” along the lines of diffusive force, and so mapping out for us a “monopolar field” of diffusion: and if we set two such drops side by side, their lines of diffusion will oppose, and repel, one another. Or, instead of the uniform layer of salt solution, we may place at a little distance from one another a grain of salt and a drop of blood, representing two opposite poles: and so obtain a picture of a “bipolar field” of diffusion. In either case, we obtain results closely analogous to the “morphological,” but really dynamical, polarity of the organic cell. But in all probability, the dynamical polarity, or asymmetry of the cell is a very complicated phenomenon: for the obvious reason that, in any system, one asymmetry will tend to beget another. A chemical asymmetry will induce an inequality of surface-tension, which will lead directly to a modification of form; the chemical asymmetry may in turn be due to a process of electrolysis in a polarised electrical field; and again the chemical heterogeneity may be intensified into a chemical “polarity,” by the tendency of certain substances to seek a locus of greater or less surface-energy. We need not attempt to grapple with a subject so complicated, and leading to so many problems which lie beyond the sphere of interest of the morphologist. But yet the morphologist, in his study of the cell, cannot quite evade these important issues; and we shall return to them again when we have dealt somewhat with the form of the cell, and have taken account of some of the simpler phenomena of surface-tension.

We are now ready, and in some measure prepared, to study the numerous and complex phenomena which usually accompany the division of the cell, for instance of the fertilised egg. Division of the cell is essentially accompanied, and preceded, by a change from radial or monopolar to a definitely bipolar polarity. In the hitherto quiescent, or apparently quiescent cell, we perceive certain movements, which correspond precisely to what must accompany and result from a “polarisation” of forces within the

cell: of forces which, whatever may be their specific nature, at least are capable of polarisation, and of producing consequent attraction or repulsion between charged particles of matter. The opposing forces which were distributed in equilibrium throughout the substance of the cell become focussed at two “centrosomes,” which may or may not be already distinguished as visible portions of matter; in the egg, one of these is always near to, and the other remote from, the “animal pole” of the egg, which pole is visibly as well as chemically different from the other, and is the region in which the more rapid and conspicuous developmental changes will presently begin. Between the two centrosomes, a spindle-shaped

Fig. 41. Caryokinetic figure in a dividing cell (or blastomere) of the Trout’s ege. (After Prenant, from a preparation by Prof. P. Bouin.) figure appears, whose striking resemblance to the lines of force made visible by iron-filings between the poles of a magnet, was at once recognised by Hermann Fol, when in 1873 he witnessed for the first time the phenomenon in question. On the farther side of the centrosomes are seen star-like figures, or “asters,” in which we can without difficulty recognise the broken lines of force which run externally to those stronger lines which lie nearer to the polar axis and which constitute the “spindle.” The lines of force are rendered visible or “material,” just as in the experiment of the iron-fil ngs, by the fact that, in the heterogeneous substance of . the cell, certain portions of matter are more “permeable” to the acting force than the rest, become themselves polarised after the

fashion of a magnetic or “ paramagnetic”’ body, arrange themselves in an orderly way between the two poles of the field of force, cling to one another as it were in threads*, and are only prevented by the friction of the surrounding medium from approaching and congregating around the adjacent poles. As the field of force strengthens, the more will the lines of force be drawn in towards the interpolar axis, and the less evident will be those remoter lines which constitute the terminal, or extrapolar, asters: a clear space, free from materialised lines of force, may thus tend to be set up on either side of the spindle, the so-called “Biitschli space” of the histologistst. On the other hand, the lines of force constituting the spindle will be less concentrated if they find a path of less resistance at the periphery of the cell: as happens, in our experiment of the iron-filings, when we encircle the field of force with an iron ring. On this principle, the differences observed between cells in which the spindle is well developed and the asters small, and others in which the spindle is weak and the asters enormously developed, can be easily explained by variations in the potential of the field, the large, conspicuous asters being probably correlated with a marked permeability of the surface of the cell.

The visible field of force, though often called the “nuclear spindle,” is formed outside of, but usually near to, the nucleus. Let us look a little more closely into the structure of this body, and into the changes which it presently undergoes. Within its spherical outline (Fig. 42), it contains an “‘alveolar” * Whence the name “mitosis” (Greek uiros, a thread), applied first by Flemming to the whole phenomenon. Kollmann (Biol. Centralbl. 1, p. 107, 1882) called it divisio per fila, or divisio laqueis implicata. Many of the earlier students, such as Van Beneden (Rech. sur la maturation de lceuf, Arch. de Biol. tv, 1883), and Hermann (Zur Lehre y. d. Entstehung d. karyokinetischen Spindel, Arch. f. mikrosk. Anat. xxxvul, 1891) thought they recognised actual muscular threads, drawing the nuclear material asunder towards the respective foci or poles; and some such view was long maintained by other writers, Boveri, Heidenhain, Flemming, R. Hertwig, and many more. In fact, the existence of contractile threads, or the ascription to the spindle rather than to the poles or centrosomes of. the active forces concerned in nuclear division, formed the main tenet of all those who declined to go beyond the “contractile properties of protoplasm” for an explanation of the phenomenon. (Cf. also J. W. Jenkinson, Q. J. M. S. xuvut, p. 471, 1904.)

meshwork (often described, from its appearance in optical section, as a “reticulum’’), consisting of more solid substances, with more fluid matter filling up the interalveolar meshes. This phenomenon is nothing else than what we call in ordinary language, a “froth” or a “foam.” It is a surface-tension phenomenon, due to the interacting surface-tensions of two intermixed fluids, not very different in density, as they strive to separate. Of precisely the same kind (as Biitschli was the first to shew) are the minute alveolar networks which are to be discerned in the cytoplasm of the cell*, and which we now know to be not inherent in the nature of

| (ee | Moral its | Vat boy Cay vy ne ' } / / aS 4 / Sk cele ite ” Sy Ay e— ' ‘nuclear membrane nucleolus Fig. 42. Fig. 43. protoplasm, or of living matter in general, but to be due to various causes, natural as well as artificial. The microscopic honeycomb structure of cast metal under various conditions of cooling, even on a grand scale the columnar structure of basaltic rock, is an example of the same surface-tension phenomenon. * Arrhenius, in describing a typical colloid precipitate, does so in terms that are very closely applicable to the ordinary microscopic appearance of the protoplasm of the cell. The precipitate consists, he says, ‘““en un réseau d’une substance solide contenant peu d’eau, dans les mailles duquel est inclus un fluide contenant un peu de colloide dans beaucoup d’eau...Evidemment cette structure se forme a cause de la petite différence de poids spécifique des deux phases, et de la consistance gluante des particules séparées, qui s’attachent en forme de réseau.” Rev. Scientifique, Feb. 1911.

But here we touch the brink of a subject so important that we must not pass it by without a word, and yet so contentious that we must not enter into its details. The question involved is simply whether the great mass of recorded observations and accepted beliefs with regard to the visible structure of protoplasm and of the cell constitute a fair picture of the actual living cell, or be based on appearances which are incident to death itself and to the artificial treatment which the microscopist is accustomed to apply. The great bulk of histological work is done by methods which involve the sudden killing of the cell or organism by strong reagents, the assumption being that death is so rapid that the visible phenomena exhibited during life are retained or “fixed” in our preparations. While this assumption is reasonable and justified as regards the general outward form of small organisms or of individual cells, enough has been done of late years to shew that the case is totally different in the case of the minute internal networks, granules, etc., which represent the alleged structure of protoplasm. For, as Hardy puts it, “It is notorious that the various fixing reagents are coagulants of organic colloids, and that they produce precipitates which have a certain figure or structure,...

and that the figure varies, other things being equal, according to the reagent used.”” So it comes to pass that some writers* have altogether denied the existence in the living cell-protoplasm of a network or alveolar “‘foam” ; others} have cast doubts on the main tenets of recent histology regarding nuclear structure; and Hardy, discussing the structure of certain gland-cells, declares that “‘there is no evidence that the structure discoverable in the cellsubstance of these cells after fixation has any counterpart in the cell when living.” “A large part of it” he goes on to say “is an artefact. The profound difference in the minute structure of a secretory cell of a mucous gland according to the reagent which is used to fix it would, it seems to me, almost suffice to establish this statement in the absence of other evidence.”

Nevertheless, histological study proceeds, especially on the part of the morphologists, with but little change in theory or in method, in spite of these and many other warnings. That certain visible structures, nucleus, vacuoles, “attraction-spheres” or centrosomes, etc., are actually present in the living cell, we know for certain; and to this class belong the great majority of structures (including the nuclear “spindle” itself) with which we are at present concerned. That many other alleged structures are artificial has also been placed beyond a doubt; but where to draw the dividing line we often do not know fi.

The following is a brief epitome of the visible changes undergone by a typical cell, leading up to the act of segmentation, and constituting the phenomenon of mitosis or caryokinetic division. In the egg of a sea-urchin, we see with almost diagrammatic completeness what is set forth here*. 1. The chromatin, which to begin with was distributed in granules on the otherwise achromatic reticulum (Fig. 42), concentrates to form a skein or-spireme, which may be a continuous thread from the first (Figs. 43, 44), or from the first segmented. In any case it divides transversely sooner or later into a number of chromosomes (Fig. 45), which as a rule have the shape of little

rods, straight or curved, often bent into a V, but which may also be ovoid, or round, or even annular. Certain deeply staining masses, the nucleoli, which may be present in the resting nucleus, do not take part in the process of chromosome formation; they are either cast out of the nucleus and are dissolved in the cytoplasm, or fade away 7 situ. 2. Meanwhile, the deeply staining granule (here extranuclear), known as the centrosome, has divided in two. The two resulting granules travel to opposite poles of the nucleus, and

* Mv description and diagrams (Figs 42—51) are based on those of Professor E. B. Wilson. there each becomes surrounded by a system of radiating lines, the asters; immediately around the centrosome is a clear space, the centrosphere (Figs. 43-45). Between the two centrosomes with their asters stretches a bundle of achromatic fibres, the spindle. 3. The surface-film bounding the nucleus has broken down, the definite nuclear boundaries are lost, and the spindle now stretches through the nuclear material, in which he the chromosomes (Figs. 45, 46). These chromosomes now arrange themselves midway between the poles of the spindle, where they form what is called the equatorial plate (Fig. 47). _ 4. Each chromosome splits longitudinally into two: usually

at this stage,—but it is to be noticed that the splitting may have taken place so early as the spireme stage (Fig. 48). 5. The halves of the split chromosomes now separate from one another, and travel in opposite directions towards the two poles (Fig. 49). As they move, it becomes apparent that the spindle consists of a median bundle of ‘‘fibres,” the central spindle, running from pole to pole, and a more superficial sheath of “mantlefibres,’ to which the chromosomes seem to be attached, and by which they seem to be drawn towards the asters.

6. The daughter chromosomes, arranged now in two groups, become closely crowded in a mass near the centre of each aster (Fig. 50). They fuse together and form once more an alveolar reticulum and may occasionally at this stage form another spireme. central spindie pas a Tia oa ae mantle-fibres > ~ ae ™ a UR re A boundary or surface wall is now developed round each reconstructed nuclear mass, and the spindle-fibres disappear (Fig. 51). The centrosome remains, as a rule, outside the nucleus.

Sea Ss we roa , x i ms \ =e 2 ae : had \ == & faery | (ES Ha = = Sore \ SEY LA | . NR I ie 4 Naa ear, "cellplate Reconstructed daughter-nucles Fig. 50. Fig. 51, 7. On the central spindle, in the position of the equatorial plate, there has appeared during the migration of the chromosomes, a “cell-plate” of deeply staining thickenings (Figs. 50, 51). This is more conspicuous in plant-cells. 8. A constriction has meanwhile appeared in the cytoplasm, and the cell divides through the equatorial plane. In plant-cells the line of this division is foreshadowed by the “cell-plate,” which extends from the spindle across the entire cell, and splits into two layers, between which appears the membrane by which the daughter cells are cleft asunder. In animal cells the cell-plate does not attain such dimensions, and no cell-wall is formed.

The whole, or very nearly the whole of these nuclear phenomena may be brought into relation with that polarisation of forces, in the cell as a whole, whose field is made manifest by the “spindle” and “asters” of which we have already spoken: certain particular phenomena, directly attributable to surface-tension and diffusion, taking place in more or less obvious and inevitable dependence upon the polar system *. At the same time, in attempting to explain the phenomena, we cannot say too clearly, or too often, that all that we are meanwhile justified in doing is to try to shew that such and such actions he within the range of known physical actions and phenomena, or that known physical phenomena produce effects similar to them. We want to feel sure that the whole phenomenon is not sw generis, but is somehow or other capable of being referred to dynamical laws, and to the general principles of physical science. But when we speak of some particular force or mode of action, using it as an illustrative hypothesis, we must stop far short of the implication that this or that force is necessarily the very one which is actually at work within the living cell; and certainly we need not attempt the formidable task of trying to reconcile, or to choose between, the various hypotheses which have already been enunciated, or the several assumptions on which they depend.

Any region of space within which action is manifested is a field of force; and a simple example is a bipolar field, in which the action is symmetrical with reference to the line joining two points, or poles, and also with reference to the “ equatorial” plane equidistant from both. We have such a “field of force” in * The reference numbers in the following account refer to the paragraphs and figures of the preceding summary of visible nuclear phenomena.

the neighbourhood of the centrosome of the ripe cell or ovum, when it is about to divide: and by the time the centrosome has divided, the field is definitely a bipolar one. The quality of a medium filling the field of force may be uniform, or it may vary from point to point. In particular, it may depend upon the magnitude of the field; and the quality of one medium may differ from that of another. Such variation of quality, within one medium, or from one medium to another, is capable of diagrammatic representation by a variation of the direction or the strength of the field (other conditions being the same) from the state manifested in some uniform medium taken as a standard. The medium is said to be permeable to the force, in greater or less degree than the standard medium, according as the variation of the density of the lines of force from the standard case, under otherwise identical conditions, is in excess or defect. A body placed im the medium will tend to move towards regions of greater or less force according as its permeability is greater or less than that of

the surrounding medium*. In the common experiment of placing iron-filings between the two poles of a magnetic field, the filings have a very high permeability; and not only do they themselves become polarised so as to attract one another, but they tend to be attracted from the weaker to the stronger parts of the field, and as we have seen, were it not for friction or some other resistance, they would soon gather together around the nearest pole. But if we repeat the same experiment with such a metal as bismuth, which is very little permeable to the magnetic force, then the conditions are reversed, and the particles, being repelled from the stronger to the weaker parts of the field, tend to take up their position as far from the poles as possible. The particles have become polarised, but in a sense opposite to that of the surrounding, or adjacent, field.

* If the word permeability be deemed too directly suggestive of the phenomena of magnetism we may replace it by the more general term of specific inductive capacity. This would cover the particular case, which is by no means an improbable one, of our phenomena being due to a “surface charge” borne by the nucleus itself and also by the chromosomes: this surface charge being in turn the result of a difference in inductive capacity between the body or particle and its surrounding medium. (Cf. footnote, p. 187.)

the centrosomes the nucleus appears to act as a more or less permeable body, as a body more permeable than the surrounding medium, that is to say the “cytoplasm” of the cell. It is accordingly attracted by, and drawn into, the field of force, and tries, as it were, to set itself between the poles and as far as possible from both of them. In other words, the centrosome-foci will be apparently drawn over its surface, until the nucleus as a whole is involved within the field of force, which is visibly marked out by the “‘spindle” (par. 3, Figs. 44, 45).

If the field of force be electrical, or act in a fashion analogous » to an electrical field, the charged nucleus will have its surfacetensions diminished*: with the double result that the inner alveolar meshwork will be broken up (par. 1), and that the spherical boundary of the whole nucleus will disappear (par. 2). The break-up of the alveoli (by thinning and rupture of their partition walls) leads to the formation of a net, and the further break-up of the net may lead to the unravelling of a thread or “spireme” (Figs. 43, 44).

Here there comes into play a fundamental principle which, — in so far as we require to understand it, can be explained in simple words. The effect (and we might even say the object) of drawing the more permeable body in between the poles, is to obtain an “easier path” by which the hnes of force may travel; but it is obvious that a longer route through the more permeable body may at length be found less advantageous than a shorter route through the less permeable medium. That is to say, the more permeable body will only tend to be drawn in to the field of force until a point is reached where (so to speak) the way round and the way through are equally advantageous. We should accordingly expect that (on our hypothesis) there would be found cases in which the nucleus was wholly, and others in which it was only partially, and in greater or less degree, drawn in to the field between the centrosomes. This is precisely what is found to occur in actual fact. Figs. 44 and 45 represent two so-called “types,” of a phase which follows that represented in Fig. 43. According to the usual descriptions (and in particular to Professor

* On the effect of electrical influences in altering the surface-tensions of the colloid particles, see Bredig, Anorganische Fermente, pp. 15, 16, 1901. E. B. Wilson’s*), we are told that, in such a case as Fig. 44, the “primary spindle” disappears and the centrosomes diverge to opposite poles of the nucleus; such a condition bemg found in many plant-cells, and in the cleavage-stages of many eggs. In Fig. 45, on the other hand, the primary spindle persists, and subsequently comes to form the main or “central” spindle; -while at the same time we see the fading away of the nuclear membrane, the breaking up of the spireme into separate chromosomes, and an ingrowth into the nuclear area of the “astral rays,” —all as in Fig. 46, which represents the next succeeding phase of Fig. 45. This condition, of Fig. 46, occurs in a variety of cases ; it is well seen in the epidermal cells of the salamander, and is also on the whole characteristic of the mode of formation of the “polar bodies.”’ It is clear and obvious that the two “types” correspond to mere differences of degree, and are such as would naturally be brought about by differences in the relative permeabilities of the nuclear mass and of the surrounding cytoplasm, or even by differences in the magnitude of the former body.

But now an important change takes place, or rather an important difference appears; for, whereas the nucleus as a whole tended to be drawn in to the stronger parts of the field, when it comes to break up we find, on the contrary, that its contained spireme-thread or separate chromosomes tend to be repelled to the weaker parts. Whatever this difference may be due to,— whether, for instance, to actual differences of permeability, or possibly to differences in “surface-charge,”’—the fact is that the chromatin substance now behaves after the fashion of a “diamagnetic” body, and is repelled from the stronger to the weaker parts of the field. In other words, its particles, lying in the inter-polar field, tend to travel towards the equatorial plane thereof (Figs. 47, 48), and further tend to move outwards towards the periphery of that plane, towards what the histologist calls the ‘“‘mantle-fibres,” or outermost of the lines of force of which the spindle is made up (par. 5, Fig. 47). And if this comparatively non-permeable chromatin substance come to consist of separate portions, more or less elongated in form, these portions, or separate “chromosomes,” will adjust themselves longitudinally,