The mode of generation of the reproductive elements and their relation to the cells of the parental tissues, is a problem which has always made heavy demands on the labours of microscopical investigators, whether zoological or botanical ; and although an immense literature has grown about the subject since Flourens thought embryos were formed “tout d’un coup,” at the moment of the fusion of the sexual elements, it may be safely asserted that not until the last few years has any very definite knowledge been acquired, and only in a limited number of cases has it yet reached any high degree of accuracy.

With Plates 15 and 16

In a certain number, however, we do know what is actually done during the origin of these cells; and this knowledge is a priceless gift to the biologist, as there is little doubt that the apprehension of karyokinesis, both in its relation to the “Reductions-Theilung “and the ordinary division of somatic cells, has brought him face to face with the actual mechanical expression of hereditary transmission and the problems connected with it.

The modus operandi of the forces which bring about these changes, however, or any serious attempt to ascertain whether they be modifications of ordinary physical phenomena at all, or whether the whole “fleeting show “of attractions, repulsions, and nuclear metamorphoses must be looked upon as something outside the physical domain, as ordinarily understood, is still a fundamental, though quite legitimate problem for microscopical inquiry.

In attempting to obtain a clear conception of the premises from which we have to start in such inquiries, the once absorbing question of the nature of the wide structural differences often apparent in the male and female cells will be found to have lost, if not much of its importance, at any rate most of its original characters. The conjoint labours of Hertwig,1 Ishikawa,2 vom Rath,3 and others, as well as the curious observations of Weismann concerning the non-specialisation of certain spermatozoa, have completely changed the scenes in this direction; and it is matter for rejoicing that the shifting, at any rate in this particular, tends towards a simplification in the gradual banishment of apparent difference in such elements, and of their associated mechanical complexity, to the rank of a purely physiological importance.

Opinion to-day is almost unanimous that the ova and spermatozoa are strictly similar objects, that even the most modified spermatozoon still carries about with it the dwarfed representatives of cell structure; and our knowledge of its development is sufficiently advanced to recognise in the head the reduced nucleus, the kytoplasm in the tail ; and lastly, it appears probable from Hermann’s,4 and more especially from Fick’s5 investigations, that the hitherto enigmatical “Mittel Stiick “is in reality nothing less than the attraction sphere.6

Although both the eggs and the spermatozoa are cells, and similar cells, the final karyokineses which produced them are different from the preceding divisions in the cells of the genital epithelium, whether male or female. As is now well known, this change in the divisional phenomena appears in the extrusion of the polar bodies in the egg, and the “Reductions Theilung” in the spermatozoa. The existence of these phenomena constitutes the empirical though important ground for several well-known theories concerning the physiological value of the “Reductions Theilung” as a preparatory balancing of the “hereditary substance” in two cells, whose fusion forms the starting-point of a succeeding generation.

Nearly all the readings of this riddle actually offered, have sprung from one of two sources: either they have come back upon us as the new-clad ghosts of Balfour’s famous interpretation of the polar bodies; or have accepted as sufficient explanation of the facts, Weismann’s theoretical conceptions of the necessity of a reduction in the quantity or the quality, or both, of the hereditary substance (chromatin).

The continuous processes of assimilation and growth in the resting cells of any tissue, although beyond our actual scrutiny, offer nothing antagonistic to the generally adopted notion, that they are the result of the passive influence of an immensely complicated structure, operating under certain rather limited conditions. But the final dissolution of these conditions, when assimilation and growth in the individual can presumably go no further,1 and their re-establishment under more favorable circumstances, is quite another matter.

In this procedure, the essential material constituents of the cell are accurately halved and separated, and we may presume that the complicated mechanical basis of the resting cell’s activity, or at any rate the power to return to it at some future time, goes with them. What are the means by which this material distribution is effected ? According to more recent investigation, an essential agent seems to be two centres of an alternately attractive and repulsive nature (centrosomes) ; but concomitantly with, and independently of, their operation, other and no less important changes accrue within the cell, perhaps most notably, the evolution of the fine chromatic reticulation of the resting nucleus into a limited number of chromosomes at the nuclear periphery, their number being constant for any particular species—a fact making them of great practical importance in all questions relating to heredity.

Now I wish to call attention to some points in the spermatogenesis of Branchipus, which may appear to throw much light on portions of these successive stages; and although it is at present hopelessly inadequate to illuminate the greater problems which arise from it, I have taken extra trouble to be sure of my. ground here, because I conceive that the existing theoretical solutions of these problems must one day find powerful confirmation or the reverse, in a true appreciation of the character of the processes which underlie the karyokinetic metamorphosis.

The male gland in Branchipus is a rather straight biramous tube extending up the tail, and a short distance further up the body. If spermatozoa are free in the lower portion all tbe stages of spermatogenesis are visible as we pass up. The difference in phase amongst the cells may be taken to represent the zones of Hertwig, van Beneden, and Julin.1

The spermatocytes, however, break away from the walls in groups (fig. 21), their individual components being all in the same phase. But as this phase, characteristic of each group as a whole, is not often similar to those on either side of it, we cannot strictly divide the tube into ascending zones. When the ordinary somatic division has come to an end spermatogenesis sets in among the cells lining the hollow of the tube. These somewhat minute elements rapidly increase in size, and their nuclei pass into a spirem, with a peculiar and characteristic grouping of their chromatic elements all on one side, just as in Hermann’s beautiful figures of spermatocytes in the salamander (fig. 3)1

Sometimes before, and always during the course of these changes, bodies answering to the centrosomes in all peculiarities except their number, which is abnormally great, make their appearance in the angular mass of protoplasm at the bases of the characteristic cells represented in figs. 1—4, a.

Merely for the sake of clearness, and to keep these bodies out of confusion with the true and enormous centrosomes appearing later, as well as to separate them from another type of body, to which I shall have to refer at length, I have provisionally collected these bodies under the term pseudosomes.

As the spermatogenesis proceeds, the lop-sided chromatic arrangement of the spirem rapidly gives place to ten chromosomes, all arranged on the nuclear periphery, and these ten chromosomes in turn become transversely constricted to form the well-known dumb-bell elements (figs. 8—12), so that we have ten double or twenty single chromosomes, which rapidly arrange themselves in the disc-like equatorial plate seen in optical section (fig. 11).

At this period of the metamorphosis (Flemming’s metakinesis) a number of most remarkable bodies make their appearance, more or less exclusively related to the cell periphery, but connected one to another and to the inner group of chromosomes by fine strands, which remain uncoloured by reagents ; and, as their relation to these fine threads suggests the nodal points in a net, I have termed them dictyosomes (figs. 11—13, d).

The constriction between the dumb-bell-like heads of the chromosomes becomes more and more pronounced, and they ultimately separate, passing in opposite directions towards the relatively colossal centrosomes now occupying the spindle apices (figs. 12, 17, 19, c, d). I have separated the dictyosomes from the centrosomes, not because they appear to be in any way essentially distinct, but because they originate at a later period in the division, and the two sets of structures might otherwise be confused.

Respecting the relation between the pseudosomes, centrosomes, and dictyosomes I shall speak later on.

The two nuclear groups now separate as in fig. 14, and the first reduction division is completed. The small elements thus formed never again regain the character of a resting cell,1 but there are appearances of irregular division, resulting in the formation of two excessively small spheroidal bodies, each presumably containing the equivalent of five chromosomes (figs. 15, 16). This procedure must be looked upon as constituting the second “Reduction Theilung,” and the resulting elements are the mature spermatozoa.

With the above broad facts of spermatogenesis kept well in view I proceed to a more minute description of the successive stages of the karyokinesis related to it, more especially with a view to determining the nature of the bodies I termed dictyosomes and pseudosomes in the previous description, and which at first seemed to appear, disappear, and reappear in a quite bewildering fashion. The original small spermatocytes are similar in all essentials to the least specialised elements of the somatic tissues.

When stained with orange, gentian violet, or hæmatoxylin, after treatment with Hermann’s or Flemming’s fluid (the best results were obtained from a combination of gentian violet and orange), the somewhat triangular cells2 present a fine reticulate appearance, both within and without the nucleus. The meshes of this reticulum are of fairly equal size in both cases (fig. 1), and a close examination leaves no doubt that the appearance (at any rate in these cells) is produced by a vast number of clear globules, kept apart by some non-miscible intervening fluid ;1 in fact, the whole might fitly be described as a foam structure, or “Schaumplasm” of Bütschli.

I have attempted to give some idea of this appearance in fig. 1, a resting spermatocyte just previous to its division, but the result is not nearly so impressive as I could wish.

Nuclear stains affect to a certain extent the intervening fluid throughout the whole cell, and the stain appears to be related to excessively fine granules suspended in a clear plasma. These cyto-microsomes do not appear to be “varicosities” of the kytoplasmic strands between the globules, but the stain appears to affect microsomes suspended in this intervening fluid. The whole darkened nuclear area suggests a condensation of this staining material, possibly by its own cohesion.

Outside the nucleus there are usually to be found, on the side where there is most cell body, and where vom Rath represents the centrosomes in the resting spermatocytes of Gryllo-talpa, those dark points, whose appearance corresponds in everything but number with the centrosomes as ordinarily understood, and which I collected in the more general description under the term pseudosomes (figs. 1—6, a). No archoplasm is apparent round them, and a close examination suggests that they are simply the expression of a collection of the above staining material (microsomes) in the angular spaces between the spheroids, producing the reticulate appearance (figs. 1—9).

Careful search will, as I have said, often raise the number of these bodies as high as six or eight. The more we look the more difficult it becomes to separate the pseudosomes from the less conspicuous interspaces of fluid between the globules ; both appear to pass insensibly into each other. The appearance and relation of the more conspicuous are very striking, as observation of their subsequent behaviour left no doubt on my mind that they were intimately bound up, if not with the origin, at any rate with a remarkable increase witnessed in the centrosomes ultimately occupying the apices of the spindle figure. It will, however, be well to advance the description of the mitosis a little before discussing this point. The first nuclear differentiation appears at one side of the nucleus as a colourless spot (fig. 2), which grows, driving the chromatic network before it to one side (fig. 3). The individual chromatin bands become shorter and thicker in proportion to this displacement, and nearly all the fine strands of “linin’’ disappear from this area, or, in other words, the spheroids fuse one with another, the fusion being produced by the substance of the clear globules breaking through the walls of intervening fluid one into another. In fact, this fusion spreads just as in soap froth the larger bubbles grow at the expense of the smaller, and the continuance of such a process results in the chromatin being thrown to one side in the form of a crescent (fig. 3), its threads are naturally thickened in proportion to their displacement, and the curious initial figure, which so much struck Hermann in the spermatocytes of the salamander, appears to be a necessary consequence of an intra-nuclear fusion in Branchipus.1

As the intra-globular fluid (with its staining granules) is between the adjacent spheroids, or in any single instance is peripherally disposed towards them, it follows that if the fusion continues until the whole nucleus consists of one or of a small number of spheroids, the intervening staining chromatin will be, as it practically is, all on the periphery.

Secondly, the fewer the globules, the larger and fewer the angular spaces between them (figs. 4, 5), and consequently the more deeply staining intervening matter appears as a limited number of chromosomes connected by fine striæ (linin) ; their actual number will naturally depend on the size of the spheroids compared with that of the nucleus.2

A great deal of importance has been attached directly or indirectly to the number of the chromosomes by all the more recent investigators, and this factor in their origin (in Branchipus) fully bears out the assumption that they are the visual expression of the primary constitution of the cell to which they belong. Nor is this all ; for if we believe, as we have every reason to believe, that the character of the nucleus is the determining factor of the nature of the cell’s activity, that curious variation in the number of the chromosomes in cells of closely allied species would be more intelligible ; for although the frothy structure of the nucleus might be actually or closely similar, a very slight difference in the cellular dimensions would, provided the foam structure remained the same, materially alter the number of the spaces between the globules, and consequently the number of the chromosomes. It is at the same time apparent that this number, as well as the general nuclear characteristics, oscillate within narrow limits for the same species.1

It is interesting to note in this connection that the characters of nuclei, in Arthropods and Annelids, have much in common. They nearly all present the peculiar ball-like chromosomes during metamorphosis, just as they tend to form a reticulate nucleus when at rest. In fact, we might say such nuclei constitute an Annelidean nuclear type.

Again, the characters of the Mammalian nuclei are very constant, but they nevertheless differ in minor details even from those of the Amphibia. In fact, the difference between these two latter is as small as that between them both, and the former, is great. The comparative study of nuclei is well worthy of more minute attention; suffice it, however, at the present moment to point out that such generalisations would have weight in our conceptions of heredity.

Of the regular occurrence of a peculiar intra-nuclear fusion in Branchipus the appearances leave no doubt, or that it is primarily instrumental in bringing about the conversion of the fine chromatic network of the resting nucleus first into the lop-sided figure described by Hermann, and probably has a good deal to do with the origin of the ten chromosomes on the nuclear periphery (figs. 4—8). But the initial impulse which starts such a fusion is an entirely different matter. This might rise from a variety of causes, from a gradual increase of internal pressure caused by osmotic action, or it might be produced by some change in that polarity supposed to exist between the centrosomes lying close to its exterior; and it is curious to note in this connection that the fusion in Branchipus does start from that side where the most marked pseudosomes exist (figs. 3, 4, 6, a, a), and, if we may put the same interpretation on the metamorphosis of other cells, Hermann’s, vom Rath’s, and possibly Flemming’s figures would be in complete accordance with such a view. I very much doubt, however, if either of these suppositions will be found to be the explanation of the origin of the fusion in the first instance. But, if once started, we have seen that the nuclear metamorphosis during karyo- kinesis, from the resting stage up to that at which a limited number of chromosomes exist on the periphery, is, to a certain extent, the logical consequence of its progress.

I have arranged the succeeding description in the light of this conception because, since it has helped us thus far, we might pre-suppose it’ useful in the elucidation of other karyo- kiuetic phenomena; and, unless I have done very indifferent justice to the appearances before me, this supposition should be fully justified.

The ten ellipso-spherical chromosomes (figs. 5, 7) which have arisen from an irregular transverse splitting, or, rather, running into drops of the thickened chromatic network, after it was brought by the progressive fusion to the nuclear periphery, become rapidly constricted in the middle to form the dumb-bell figures characteristic of these and many other Arthropod nuclei (figs. 7—12).

Each cell now contains ten double or twenty single chromosomes, i. e. double the ordinary number (figs. 7,10,12) ; and it is interesting to compare such nuclear figures and their origin with others, like those of Salamander, in which the chromatin is arranged in a succession of bands or irregular annuli, set more or less transversely to the long nuclear axis. These appearances would be produced by a similar fusion of globules, in Such a manner that a few diaphragm-like membranes of the intervening fluid with its staining microsomes were left across the long axis of the nucleus. In such a case the chromatin would inevitably be arranged as it always is in the re-entrant solid angles. Interesting artificial reproductions of the nuclear figures may be seen by watching the growth of bubbles, and I have given in fig. 18 some drawings of the ultimate configurations produced by the growth of bubbles in a fine froth. The lines of foam left as bands along the position of the ruptured walls would represent the chromatic loops, and it will be seen that they show a marked tendency to contract into more or less rounded bodies.

The existence of nuclei in groups of four or five, each with their ten dumb-bell chromosomes, gives a very striking appearance to the testes of Branchipus (fig. 21) ; and while the condition characterises one of the longest phases of the whole nuclear division, its final metamorphosis occurs with the utmost rapidity, the cells appearing as if transformed by magic into a complete spindle figure. Intermediate phases are, however, to be found, and it appears that the fusion or running together of the globules continues, breaking through the old nuclear boundary at several points into the surrounding kyto- plasmic network (fig. 8), so that the clear mass of nuclear plasm appears to spread out on all sides (figs. 8, 11, 19). The result of this is that chromosomes are at last left hanging in a clear central space by a few irregular strands of this kytoplasmic network, into which the fusion has not yet broken (figs. 10—12, 19). These irregular strands are ultimately reduced to fine threads (figs. 11, 12, 17, 19), and their peripheral extremities are related to dark bodies which can be nothing but the pseudosomes of which I have already spoken in an earlier phase of the metamorphosis (figs. 10, 11, 12, da.). These pseudosomes appear now to have increased in size somewhat, their relation to the spaces of the intra-globular network being more pronounced, and we are naturally led to the conclusion that such an increase is brought about by the massing of the staining material in these angular spaces, owing to the progressive fusion tending to lessen their number and increase their size, just as it did with respect to the chromatin within the old nuclear limits.

Proportionately to the extension of this fusion, the tension along the achromatic lines, on which the chromosomes are suspended, becomes greater as the dark points (pseudosomes) at their peripheral extremities retreat with the vanishing achromatic network and its contained microsomes towards the cell’s circumference (figs. 10—12, 19).

If we now try to realise what is actually taking place, it will become apparent that the traction towards the periphery through these points (pseudosomes) along the achromatic threads, and ultimately upon the chromosomes themselves, will tend to set itself along some axis across the nuclear figure as a whole, and the points (pseudosomes) chosen will be those on opposite sides which have, so to speak, the best foothold in the periphery. The remaining points (pseudosomes) will tend to glide as they do (figs. 9,10,12) towards the extremes of such an axis, and a spindle figure will be finally set up (figs. 9, 12, 22, 23). From the figures just referred to, it will be apparent that the coalescences of the points of attachment of the distal extremities of the achromatic fibres (pseudosomes) become marked out as centrosome-like bodies which travel away towards the cell’s circumference, and finally come to rest on its extreme margin (figs. 11, 22, 23). In other words, these centrosomes are virtually derived from a fusion of some of the pseudosomes, and these were in turn seen to originally correspond to the angular spaces in a network exterior to the nucleus.1

In cells a trifle more advanced than those represented in the preceding figures showing (fig. 19) the area of clear fluid produced by the massing of the original diffuse staining material within the nucleus into the small space of the chromosomes, by the process I have described—it will be seen that this space, which in the first stage of the spindle figure represents the nuclearplasm, and retains the spherical character of the original nuclear contour, has become very much enlarged (figs. 11, 12, 23), not only in the direction of the spindle axis, but laterally all round, so that it appears as a continually increasing irregular area, occupying by far the greater part of the cell’s substance. Round this irregular space a rind of the original kytoplasmic reticulum still remains (figs. 11, 12, 19, 23), and it will be noticed that at the junction of this rind and the clear fluid within (fig. 19) a number of small staining points exist, related to the angular spaces between the clear globules and the non-miscible intervening fluid.

These bodies grow continually, and their size marks the progress of the fusion of the clear central mass of fluid with the similar constituents of the peripheral rind, just as the thickening of the chromatin bands was the measure of the fusion proceeding within the original nuclear limits.

They continually stain more and more deeply with orange and gentian violet, as the diffuse staining material dispersed through what remains of the kytoplasmic network is swept before the progress of the fusion into nodal points until, simultaneously with its extension through the whole cell, they are left as some twenty conspicuously dark bodies regularly arranged on the periphery (figs. 13, 17). Close examination reveals, however, that the fusion is not really complete, but that fine achromatic threads connect these bodies one to another (figs. 13—19) and to the inner group of chromosomes in the manner described in an earlier part of my paper—a fact which led me to devise the term dictyosome as expressive of these peculiar relations.

It will be seen that these dictyosomes appear at a definite point in the karyokinetic metamorphosis, viz. the later phases of the spindle figure ; and the cells which present these conditions are comparatively large spherical bodies, which have become so much altered in their refractive characters that one is reminded of Flemming’s words when describing a similar change witnessed in the dividing cells of the salamander :

“Bekommt man unwillkürlich den Eindruck, als sei die Zelle wahrend ihrer Theilung durch und durch mit einer be- sonderen Substanz durchtrânkt oder—um mich vorsichtiger auszudriicken—als besitze sie durch und durch eine besondere physikalische oder chemische Beschaffenheit.”

This change appears in the spermatocytes of Branchipus to be the direct result of the collecting of the primarily diffuse staining material of the resting nucleus into ten chromosomes, and of that existing in the kytoplasm without into distinct chromatic bodies (dictyosomes). Both these changes are apparently due to a progressive fusion or running together of the clear globules which, begun within the nucleus, formed the chromosomes on its surface, and extending, swept the diffuse staining material of the cell body together into some twenty dictyosomes on its circumference. In this brief and necessarily crude manner I hope to have made the main drift of the investigation up to this point clear. I have dealt with the development of the chromosomes in Branchipus, and shown reason to believe that it is in a measure dependent on a fusion of the globules which give rise to the reticulate appearance. The progress of such a fusion would produce the one-sided figure described by Hermann in the spermatocytes of the salamander, and tend ultimately to form a limited number of chromosomes all on the nuclear periphery; and we have seen that during these changes there exist in the resting spermatocytes those dark points (pseudosomes) whose appearance corresponds in everything but number with the centrosomes of previous authors.

We have seen also that the fusion producing such wide changes in the nucleus spreads beyond it, leaving the chromosomes suspended to the pseudosomes. These pseudosomes retreat with the remnant of the original network as it vanishes towards the periphery, and in connection with this motion an axis tends to be set up round which the spindle figure gathers, while at its apices some of the pseudosomes coalesce to build up the colossal centrosomes.

Lastly, just as the fusion within the nucleus brought about the massing of the chromatin into a limited number of chromosomes, so also the extra-nuclear fusion operating in the same way upon the sparse staining material of the kytoplasm, without the nucleus ultimately collects this, into chromatic bodies in the angular spaces between the enlarged globules. They first appear as an irregular cloud on the outskirts of the fusion(fig. 23,d), grow enormously in size, and acquire a regular distribution on the cell periphery. They still, however, remain connected one to another and to the inner group of centrosomes by fine threads ; the fact that they thus form, as it were, the nodal points in a net, suggesting the term dictyosome as expressive of this peculiar relation. It will, moreover, have become apparent from the description that there is no genetic distinction between the pseudosomes, centrosomes, and dictyo- somes, and my sole reason for using the two new terms is their spccessional appearance.

In thus bringing into prominence the existence in Branchipus of a veritable “Schaumplasm ”and its inter-activities, I would observe that I do so with no predisposition to utilise Bütschli’s conception of such structure as a fundamental interpretation of some of the phenomena of karyokinesis, either in this or any other case, but rather the reverse. Nevertheless the observation that a foam structure is intimately bound up with the phenomena of karyokinesis on the one hand (even in a single type) must materially enhance the value of Bütschli’s ingenious hypothesis that it is sufficient to account for the amoeboid activities of protoplasm on the other.

A very natural objection to the conclusion I have stated may arise out of the apparent whittling process to which it subjects the centrosomes, resolving these bodies into nothing more than the irregular staining material between the globules of a protoplasmic froth.

I wish, however, while concluding this part of my paper, to point out that such an objection is only apparent, and not real.

In a former essay, while discussing the meaning of the difference in the component parts of the spheres apparent in the works of Flemming, Hermann, van Beneden, Boveri, and others, I remarked, “Comparison between the spheres and their constituent parts in various animals might appear pedantic, and, in the present state of our knowledge, unnecessary, if it were not that some of these parts are probably, as we have seen, the fleeting expression of metamorphic phenomena; while others (such as the central body), though dividing, retain their characteristics unimpaired; “and I have ventured to repeat this as showing that the great pioneers of this phase of cytology had already hunted the all-important part of the sphere down to the narrow limits of the centrosome. And the fact that in Branchipus six or eight bodies indistinguishable from one another exist at first, and that these afterwards fuse to augment the size of the two actually chosen to occupy the spindle apices, does not prevent anyone from regarding these two of the six or eight, as endowed with special properties if he pleases, nor does their relation to the interglobular spaces affect the point in any way that I can see. Whether two of these bodies are really to be regarded as different from the rest is a point on which at present I offer no opinion.

It will be seen that the spermatogenesis of Branchipus corresponds in the main with that described by vom Rath in Gryllotalpa, and that the reticulum has disappeared in the ultimate division altogether (fig. 16). Now if, as I have shown, there is reason to believe the reticulum in this particular instance is a mechanical factor in portions of the karyokinesis, all possibility of such phenomena will come to an end with the complete fusion of the clear globules, and there is thus a definite reason why the subdivision goes thus far and no farther, at any rate for a time.

In the ovigenesis proper—that is, in the metamorphosis among those cells which directly produce the eggs—there is nothing special ; but among the cells subsidiary to this process, many points are worthy of attention. Scattered through the egg mother-cells are numerous groups and rows of nuclei, obviously of a different character from those destined to form the eggs (figs. 20, 21, 43, &c.). These nuclei are very irregular in outline, and of a fine reticulate appearance. They show numerous figures of direct division (figs. 20, 43). At the same time it is quite easy to establish a gradational series extending from the true egg, forming nuclei on the one hand to the irregular akinetically dividing elements on the other, the latter class being always intimately and actually concerned in the secretion of a peculiar slimy substance (fig. 44) ; and this slime in turn is ultimately worked up in the lower portion of the tube to form the ornamental egg-case, so that although in the egg formation in Branchipus the primitively similar genital cells (male ova) diverge along two ways, one leading through successive karyokinesis to the final eggs-, they ultimately both cooperate in the perpetuation of the species by the rest being bodily transmuted into the ornamental case in which the eggs are laid. This duality in the ovarian elements is interestingin the sense that it offers a precise parallelism to the dualism caused in the spermatic apparatus by the presence of the akinetically dividing foot-cells, over whose significance so much controversy has at times been raised. In Branchipus the foot-cells are more regularly arranged than the above akinetically dividing elements in the ovary. At the upper end of the gland they occur at intervals of about ten cells in all directions, and, true to their female homologues, are more numerous as we descend towards the genital aperture. Apart from the function of the foot-cells no one can be in doubt as to their homology with the above akinetically dividing elements of the ovary ; and the fact that the latter are intimately bound up with the formation of the slime that makes the egg-case (slime-cells) seems to me to remove all doubt from vom Rath’s theory, that in the spermatogenesis they are concerned in the secretion of a fluid in which the spermatozoa are suspended. The key to the whole position seems to lie in the observation of La Valette St. George, that the mulberry-shaped masses of the spermatocytes in Blatta are produced from one cell, whose residual moiety remains, acquiries distinct characters from the rest, and is not converted into the spermatozoa. From such a starting-point we may see our way through a gradual evolution to meet the physiological necessities of the case, to the complex reproductive apparatus in Branchipus, where two different kinds of cells exist in both sexes, one to form the eggs or spermatozoa, and one to form the case or fluid in which these bodies are respectively suspended or enclosed.

Whether akinetic division is really wholly related to the foot-cells in animals is a controverted question, but from what I have seen in Branchipus (figs. 20, 21) and elsewhere, I am inclined to believe that it is not wholly restricted to these elements, but that there is a general tendency towards the two methods in the two kinds of cells.

To recapitulate, it will be seen then—

I.That in Branchipus the observations bear out the general law as to the similarity of the male and female cells, and that their own specific peculiarities are physiological in origin, having no morphological significance.

II.The derivatives of the primitive genital cells (male ova) are of two kinds, one transformed directly into the reproductive elements, the other into the egg-case or into the fluid in which the spermatozoa are suspended. Karyokinesis is the method of procedure in the one—akinesis in the other.

III. That the divisional phenomena of these cells are intimately related to a protoplasmic structure, which might be fitly described as “Schaumplasma,” and one of the initial physical impulses towards metamorphosis is a fusion of some of the intra-nuclear globules ; and a considerable portion of the complicated karyokinetic figures, with their centrosomes, pseudosomes, and dictyosomes, appear to be the logical as well as the actual consequence of the continuance of this process.

With the foregoing results of observation as a basis of comparison, I made a close examination of the ovigenesis in Apus. Unfortunately the male of this species is practically unknown, ten thousand having been collected without a male appearing in a single instance. This renders the chances of proper fertilisation very rare, and, as all the specimens are equally prolific, we must look either to hermaphroditism or parthenogenesis as the means by which the embryonic development is started. The hermaphrodite character has recently been ascribed to the genital gland of various species of Apus, and certainly the appearances which have come under my notice favour this view.1 It is, however, immaterial to the line of inquiry I have adopted, which method of reproduction actually obtains. The genital gland is an irregular tube with numerous diverticula branching out on all sides. The cells lining the main tube and its numerous ramifications are excessively minute columnar bodies (figs. 24, 31, and 35—37), and the whole appearance is far more like that of an Invertebrate intestine than a reproductive gland. Each of the epithelioid cells contains a small peculiar nucleus (fig. 36), whose position in the rod-shaped mass of protoplasm it dominates, varies in concert with all the nuclei of the same diverticulum. The nuclei oscillate backwards and forwards from the extremities of the cells nearest the lumen of the gland to those nearest the basal membrane bounding at its periphery. When in the former position, such protoplasm as remains between them and the actual glandular cavity is seen to be rapidly degenerating into masses of slime (fig. 31) ; and, just as in the case of Branchipus, this slime is ultimately worked up into an ornamental eggcase. When the nuclei have translocated themselves towards the bases of the cells, the slime has broken away in streaks and globules, and many nuclei are seen subdividing themselves into groups, from whose derivatives the nuclei of the future eggs are formed (figs. 25, 27—29, 31).

It is thus obvious, that for some reason or other an economy has been effected in the reproductive apparatus of Apus, and that there is no such permanent differentiation between slime and egg-producing cells as is apparent in Branchipus, but that the same type of nuclei having gone down to the lumen of the gland and, so to speak, performed their dirty work themselves, travel back again to the more peripheral regions, proceeding by a series of extraordinary divisions to instal themselves directly as the nuclei of the eggs. During these migrations the nuclei retain their peculiar character little changed. In all the genital cells, the chromatin is aggregated into one or two nucleoli (figs. 24, 30, 36), constituting a nuclear type which represents the extreme term in a series, whose mean would be represented by the intestinal nuclei of Carcinus, Idotea, and others described by Frenzel ;1 where the chromatic substance is still, to a certain extent, distributed through the mass of the nucleus, although some may be aggregated into massive nucleoli.

In Apus there is no vestige of colouring matter outside the single chromosome which occupies its centre (fig. 36), the substance of which is so dense and refractive, that it appears like a red lens suspended by one or two colourless threads from the hollow sphere of the nuclear membrane. The intervening space is entirely filled by a perfectly clear nuclearplasm.

If such nuclei are at the periphery of the gland, and the egg formation is about to begin, one of these single chromosomes is seen to elongate just as the nucleolus in Frenzel’s “Nucleolöre Kernbulbirung “become constricted in the middle, and finally separate into two halves, the nuclear membrane being but slightly elongated in the direction of the fission (figs. 36—4L). The two derived chromosomes may in turn divide at right angles to the first separation axis, and a nucleus with four chromosomes results (figs. 27, 29, 34, 35), whose membrane is seen to be gradually tucked in at four intermediate points, and at fig. 27 a final cross-shaped differentiation can be made out between the indentations.

Along these lines the four quadrants ultimately separate, giving rise to four nuclei, each with a single chromosome (fig. 29).

Such groups of four nuclei are always associated with the egg formation in Apus, and are seen in all stages bulging out the membrane of the gland into the body-cavity. But it is by no means the rule that these tetrad groups are formed in this manner from one cell. In some cases two nuclei in adjacent cells divide, and the derivatives nearest the periphery divide again to form the group (fig. 25), or the group formation may proceed in a much more irregular fashion out of one or two secondary divisions of adjacent cells (fig. 27).

The dividing chromosomes within the nuclei at times present the very curious appearance represented in figs. 34, 35. It will be here seen, that between the separating, more darkly stained portions stretches a stainless band, which again suggests that the stain only affects particles suspended in a clear fluid, and that this fluid is non-miscible. Moreover it would seem that these particles tend to run together into chromatic drops, leaving a clear fluid (paranuclein).

The initial impulse, whatever it may be, which gives rise to the groups of four chromosomes, and is ultimately concerned in the formation of the egg, is seen also to affect the surrounding nuclei, which divide in the same peculiar manner again and again, until they form a narrow stalk connecting the original group of four with the cavity of the gland (fig. 42). The extreme minuteness to which this subdivision is carried will be seen (fig. 33), where it will be observed that all trace of cell membrane is fast disappearing. The minute, free, nuclear elements then left spread over the surface of the tetrad group as a thin protoplasmic membrane, in which they rest without dividing walls of any kind (fig. 42, a).

During all this multiplication of the nuclei, the character of their division remains precisely the same. In every division the single chromatic ball passes through the metamorphosis represented in figs. 36—41, the nuclear membrane contracting until two precisely similar nuclei are left in the place of one.

It will be obvious that this method of procedure, though on the face of it approaching akinetic or direct division, is in reality very different from the process as it appears in Branchipus, or in the other forms in which it has been described. In all these the resting reticulate nucleus never passes out of that condition, but is constricted into two portions, each retaining its original character. It does, however, as above stated, bear considerable likeness to the “Nucleoläre Kernbulbirung” described by Frenzel. This latter mode of division is also normal to the intestinal cells of Apus. It will, moreover, be admitted that it bears a superficial resemblance to the fragmentation seen in leucocytes. But the division in these elements is certainly merely a shortened-up karyokinesis, accompanied by centrosomes and an archoplasmic metamorphosis, while no such structures are apparent in either intestinal or genital cells of Apus.

In the sense that no spindle or related parts are apparent iu these cells, their division approaches akinesis. In the sense that all the chromatin is gathered into a single chromosome it approaches karyokinesis. Monomeric (or division by single chromosomes) is the best term I can devise for this method of nuclear fragmentation, although of its actual affinities I am still in considerable doubt.

It is well known that in many plant forms, such as the Myxomycètes, the karyokinesis, although not absent, is passed through in a reduced condition, and is apparently, exclusively related to spore formation ; and within the last few days Professor Farmer has drawn my attention to a very remarkable mode of spore formation, which he has found in certain liverworts of Ceylon, in which it is with the utmost difficulty that the apparently akinetic formation of the tetrads can be shown to be in reality a quadripolar karyokinesis ; and, further, it seems generally agreed that such simplification is a reduction from, and not an antecedent of, the more complex karyokinetic division phenomena.

Consider also the nuclear division in the Protozoa themselves. It is now known that karyokinesis of a more complex order, accompanied by an enormous number of chromatic bands, is normal to some Rhizopods, while in the more specialised and less primitive Ciliates, this phenomenon is restricted to the micronuclear elements, being in them so much reduced, that it is not without difficulty that it can be recognised at all.

Such evidence seems to me to favour the conclusion that the monomeric division in the genital cells of Apus is the most extreme term known in a progressive modification of the more primitive karyokinesis through such forms as Frenzel’s “Nucleoläre Kernbulbirung,” in which the spindle, and the breaking up of the chromatin into bands or globes, as well as the resting reticulum, have long since disappeared.

There seems some probability in the assumption, then, that owing to the introduction of a peculiar method of reproduction in Apus (parthenogenesis or hermaphroditism), the divisional phenomenon has exhibited a corresponding change, that the cells of the genital gland are all alike, and can function both in slime or egg formation as opportunity arises.

The egg nuclei take origin from nuclei containing a single chromosome, but they ultimately develop a coarse chromatic reticulum with an external attraction sphere or archoplasm (fig. 42, b)> and appear much like an enlarged somatic cell.

How such a modification of the original type has arisen it is not, perhaps, very difficult to see. In sexually produced species, the nuclei intended for fusion must, so to speak, balance one another; and if karyokinesis is the original method of procedure, any tendency in an individual to infringe this rule in the origin of its reproductive cells would quickly tend to be eradicated from the race on account of the wide abnormalities it produced. But a parthenogenetic or hermaphrodite species might please itself as to the manner in which it evolved its reproductive elements, so long as these contained the premises necessary to the proper development of the individual.

Illustrating Mr. J. E. S. Moore’s paper on “Some Points in the Origin of the Reproductive Elements in Apus and Branchipus.”


a = Pseudosomes. c = Centrosomes, d = Dictyosomes. da = Stages intermediate between dictyosomes and pseudosomes.

FIGS. 1—5.—Stages in first division, from the resting cells to the formation of chromosomes. Flemming’s fluid and gentian violet, a. Pseudosomes.

FIGS. 6 and 7.Formation of the chromosomes.

FIG. 8.—Cell showing the breaking down of the nuclear membrane.

FIGS. 9 and 10.—Two spermatocytes, with nearly the same stage of early spindle figure, da. Bodies intermediate between pseudosomes and dictyosomes.

FIG. 11.—Transverse optical section of equatorial plate; fine strands of protoplasm connecting the chromosomes to the exterior network.

FIG. 12.—Cell a little later, in which the centrosomes have appeared.

FIG. 13.—Older cell, in which the spindle figure seen in section is breaking down, and in which numerous bodies have appeared on the surface dictyosomes.

FIG. 14.—Final division of same.

FIG. 16.—Secondary cell.

FIG. 16.—Spermatozoa.

FIG. 17.—Group of three spermatocytes in division, with numerous (d.) dictyosomes and (c., c., c.) centrosomes.

FIG. 18.—Figures produced by the growth of bubbles in a flue froth.

FIG. 19.—Spermatocytes, showing the breaking down of the nuclear area.

FIG. 20.—Foot-cell in akinetic division.

FIG. 21.—Portion of spermatic epithelium, with various stages of spermatogenesis. F. c. Foot-cells. Sp. Spermatocytes and spermatozoa.

FIG. 22.—Spermatocytes in division, showing the extreme position of the centrosomes.

FIG. 23.—Ditto, showing the early formation of the dictyosomes round a spreading clear space.


FIG. 24.—Portion of epithelium of the female gland of Apus.

FIG. 25.—Single cell with nucleus in partial division.

FIG. 26.—Ditto.

FIG. 27.—Irregular formation of tetrad group.

FIG. 28.—Element with four nuclei.

FIG. 29.—Tetrad formed from a single cell.

FIG. 30.—Three nuclei of a tetrad, showing micronucleoli (a).

FIG. 31.—Two tetrad groups in relation to surrounding cells.

FIG. 32.—Cell with dividing nucleus.

FIG. 33.—Portion of nuclei of tetrad, showing the minute subdivision of the nuclei.

FIGS. 34 and 35.—Method of division occurring in same nucleoli.

FIGS. 36—41.—Stages of division.

FIG. 42.—Tetrad group, the upper cell being the future six “Nebenkern.” a, a, a. Nuclei which have spread over the circumference of the group.

FIG. 43.—Group of cells from the spermatic gland of Branchipus.

FIG. 44.—Ditto, ditto, from the female gland of Branchipus, showing the relation of the akinetically dividing elements to the formation of slime.

All the preparations, except where otherwise stated, were fixed in Flemming’s fluid and stained with gentian violet.


“Vergleich der Ei und Samenbildung bei Nematoden. Eine Grundlage für cellulare Streitfragen,” ‘Archiv f. mikroskop. Anat.,’ Bd. xxxvi, 1890.


‘Studies of Reproductive Elements.’


‘Archiv mikro. Anat.,’ Bd. xl, p. 102.


‘Archiv für mikros. Anat.,’ Bd. xxxiv, Tafel 3.


“Ueber die Befruchtung des Axolotleies,” ‘Anatomiseher Anzeiger,’ vii, pp. 818—821.


The results of a re-examination of the facts of Mammalian spermatogenesis have shown that the centrosomes are incorporated in the spermatozoa in the position of the Mittelstück of Amphibia, while a portion of the archoplasm is applied to the pointed extremity of the head. Field has shown that the archoplasm “Nebenkern “of Echinoderms is incorporated together with the centrosomes as the Mittelstück.


Herbert Spencer suggests that cell division becomes a necessity in virtue of the continual decrease of the absorptive area proportionally to the growth of the cell.


“Nouvelles recherches sur la fécondation et la division mitosique chez l’ascaride mégalocéphale,” ‘Bullet, de l’Académie Royal de Belge,’ 3me sér., c. xix, 1887. “Befruchtung und Theilung des thierischen Eies,” ‘Morph. Jahrb.,’ 1875.


‘Arch. f. mikros. Anat.,’ Bd. xxxvii.


Compare vom Rath and Ishikawa, loc. cit.


Compare vom Rath’s figs., loc. cit.


When sufficiently high powers are used the appearance is almost identical with the coarse vacuolation in the ectosarc of Amoeba and other protozoa.


‘Arch, fiir mikroskop. Anat.,’ Bd. xxxvii, pp. 569—582.


I do not mean to maintain that there are no other controlling factors in the formation of a definite number of chromosomes ; this cannot be the case on account of the wide difference in the size of the nuclei of tissues of the same animal. At the same time we have no knowledge of the reticulum as related to different cellular dimensions.


In connection with this see Valentine Hacker, “Die heterotjpische Kerntheilung im Cyklus der generation Zellen,” * Berichteder Naturforschenden Gesellschaft zu Freiburg,’ Bd. vi, 1892, pp. 160—188.


Professor Farmer has kindly shown me some preparations of Lilium which give exactly the same bundles of fibres related to separate granules, any one of which might be individually considered as a centrosome.


For an account of this see the description of Siebold’s results in the ‘Klassen und Ordnungen des Thier Reichs,’ pp. 960—962. Also H. M. Barnard’s ‘Apodidæ.’


‘Arch, für micro. Anat.,’ Bd. xxxix.