The Segmentation is apparently complete, the ovum appearing to divide into ectoderm and endoderm cells.
The so-called endoderm cells are at first without a distinct nucleus, they do not get a nucleus until just before the gastrula stage.
All the cells of the ovum, ectodermal as well as endodermal, are connected together by a fine protoplasmic reticulum, which is placed, as are also the cells, immediately beneath the egg membrane, and therefore around a central space.
Each ectoderm cell consists of a central nucleus around which is a close protoplasmic spongework, which, at the outer parts of the so-called cell, becomes of a gradually looser nature until it runs into the spongework of the surrounding cells.
Each endoderm mass consists of a central denser spongework which gradually becomes looser towards the periphery of the mass until it is continued into a fine reticulum. The endoderm masses are far apart from each other and are connected by this reticulum.
The continuity of the various cells of the segmenting ovum is primary and not secondary, i.e. in the cleavage the segments do not completely separate from one another. But are we justified in speaking of cells at all in this case? The fully segmented ovum is a syncytium, and there are not and have not been at any stage cell limits. I think the cleavage should be rather described not as segmentation, but a multiplication of the nucleus or centre of force which causes a corresponding readjustment in the density of the network at different parts of the ovum, but no break in continuity.
The Gastrula arises by a process of epibole and is at first solid.
The endoderm masses at first have no nuclei. Nuclei first appear in them during the progress of the epibole by which the gastrula is formed. I have not been able to determine the origin of these nuclei. They either arise de novo in the endoderm masses or migrate into the latter from the ectoderm. The protoplasmic network at the centre of each endoderm mass is denser than at the periphery, but is without the chromatin granules, so characteristic of a nucleus. But I have described a stage of the nucleus in the fertilised unsegmented ovum in which the chromatin granules are almost entirely absent, and in which the network presents no essential difference from the surrounding network. Again, another in which the nuclear network merges so gradually into the surrounding network, that it is impossible to point to any limit between them. I therefore think it quite possible that this central denser protoplasm in the endoderm masses may give rise to the nucleus which subsequently appears.
The gastrula is a syncytium; the ectodermal nuclei are arranged around the periphery of the ovum, while the endodermal nuclei are within. The latter are characterised by their angular shape, and by never presenting the karyokinetic figures characteristic of the ectodermal nuclei. The protoplasm of this syncytium is much vacuolated throughout, but the vacuoles are largest in the centre. These central vacuoles unite and give rise to the gut cavity, which opens to the exterior through a point on the surface where the ectodermal nuclei have always been absent. This opening is the blastopore. The blastoporej until quite late in development, is traversed by protoplasmic strands, which anastomose with similar strands projecting from the protoplasm lining the large central vacuole or gut.
The gut of Peripatus arises, therefore, as a vacuole in a multinucleated mass of protoplasm, and the gastrula of Peripatus is a multinucleated mass or syncytium, with absolute continuity of the protoplasm of all parts of the ovum.
The Mesoderm.--After the definite formation of the blastopore, an area of protoplasm, placed in the ectodermal layer of the syncytium, and characterised by possessing several nuclei less densely packed together than elsewhere, is distinctly visible in the middle line of the ventral surface just behind the blastopore. This area I have called the polar area. Its nuclei undergo division and give rise to the densely packed mass of nuclei of the primitive streak. A part of it seems to persist for some time in the deeper parts of the primitive streak close to the endoderm.
The nuclei of the primitive streak migrate forwards between the ectodermal and endodermal nuclei, and take up their position in the protoplasm intervening between the latter.
These rows of nuclei are the mesodermal bands. They soon arrange themselves into groups around a central vacuole, and so give rise to the most conspicuous parts of the mesoblastic somites. I leave the ovum for the present at the commencement of the formation of the somites, merely stating that it is still a syncytium.
There are a certain number of facts in the above account which are of general interest and seem to deserve more discussion so far as their relation to processes in other forms are concerned. These are:
1. The connection between the intra- and extra-nuclear reticulum.
2. The segmentation.
3. The origin of the gut as a vacuole.
4. The syncytial nature of the embryo.
5. The origin of the mesoderm.
I propose to consider some of these points at once, and to defer the 5th, to Part 3 of this series.
A. The nucleus of the unsegmented ovum and of the early stages of segmentation of the Cape Peripatus are particularly favorable for study, because of their large size and the rapid changes which they undergo. I have not been, able to make out the sequence of these changes, but I hope with more material, which I expect to obtain this year, to be able to communicate some more facts concerning them in a future paper.
It is a disputed point as to whether or no the nuclear and extra-nuclear reticulum are continuous. Leydig (12), Stricker (16), Klein (9,10,11), and Heitzmann (5), hold that they are. So far as the nucleus of the early segmentation stage, and of the endoderm of Peripatus is concerned, I am able fully to confirm the views of these observers.
The general views I hold with regard to the nucleus are stated on p. 189 and I need not repeat them here. I only desire to point out that the opposite view, viz. that the nucleus is isolated, so far as continuity of protoplasm is concerned, is, from a physiological point of view, very difficult to accept; and I think that the burden of proof rests with him who maintains it. The peculiar lobed structure (PI. XII, fig. 3) of certain stages of the nucleus has been described before by other observers, notably by Balfour in his "Monograph on the Development of Elasmobranch Fishes," in the early stages of development.
Klein in his communication on this subject, refers (9, p. 175) to and confirms Stricker's (16) observations on the contractility of the nuclear spongework and its continuity with the extranuclear spongework in the colourless blood-corpuscles of the newt and frog. He further confirms Stricker's statement as to the disappearance of the cell membrane, and himself adds: "The nucleus is therefore a part of the cell substance specially differentiated by the presence of a membrane." Presumably Dr. Klein would still speak of a nucleus when the membrane is absent. I am not able to make out Klein's views with regard to this membrane. He says (11, p. 415): "In the convolution and basket of daughter nuclei the membrane is very indistinct and is also here due to the close position of the fibrils." I infer from this that he regards the nuclear membrane as a part of the general reticulum at the junction of the nuclear and extra-nuclear parts of the reticulum, which gets in certain stages of the nucleus a regular arrangement. This at any rate is my view for the Peripatus nucleus.
Klein figures (10 pl. 18) nuclei from the epidermis of the newt in a state of direct division. These figures resemble very closely some of the endodermal nuclei in the gastrula of Peripatus.
Klein is still more explicit as to the continuity of the nuclear and extra-nuclear reticulum in his second communication on this subject (11, p.416).
Unfortunately I have not been able to see the papers of Stricker and Heitzmann.
Leydig in his latest communication (12) regards the spindlefibres as parts of the ordinary reticulum (spongioplasma he calls it) with much elongated meshes (p. 9). He further looks upon the nuclear membrane as merely the outer portions of the nuclear network, and describes it as being porous, and takes the same view as Klein with regard to the continuity of the nuclear and intra-nuclear network.
Leydig also describes some accessory nuclei as occurring in certain cells. These are smaller than the main nucleus and stain less deeply. It is possible that they are structures of the same nature as those described in the endoderm of Peripatus on p. 184 of this memoir.
He refers, in this connection, especially to the small accessory nuclei which are found in many Protozoa, and which, according to Gruber (3) and Jickeli (7), are for the most part derived from the breaking up of the main nucleus. The particles resulting from this fragmentation of the nucleus seem eventually to come together again to form a new main nucleus. One would like to have some more details about this peculiar process in Infusoria, derived if possible from the study of sections. The term "fragmentation," which is applied to it apparently because the chromatic parts of the nucleus become separated from one another and scattered throughout the animal, seems to imply a distinct breaking up into small isolated portions. If this really happens the nucleus of Infusoria must differ from most other nuclei in which the chromatic matter is a part of the nuclear network, which is itself continuous with the extra-nuclear network. I should be inclined to look upon the process as an increase in size or extension of the nucleus, such as seems to have been described by Stricker in certain leucocytes.
Pfitzner (14), on the other hand, strongly maintains the isolation of the nucleus during the whole of its life-history, and he recommends certain reagents to demonstrate this fact. But inasmuch as he himself admits (p. 72) that these reagents produce great changes in the nucleus, his negative conclusions cannot be regarded as having so good a basis as the positive results of Klein and Leydig, whom I can thoroughly confirm in the matter.
I may draw attention in passing to the similarity of the branched endodermal nuclei of Peripatus to the nuclei of leucocytes figured by Pfitzner (14, Pl. V, fig. 21).
I have not been able to distinguish nucleoli in the nuclei of Peripatus as distinct from the chromatic thickenings of the spongework. Flemming (1) says that nucleoli proper participate in forming the chromatic figures in cell division. Flemming in his work on the cell and cell nucleus (1) has not seen the continuity between the strings of the nuclear and intranuclear spongework. He does not deny its existence but holds that it is not proved.
Flemming makes the important statement that the first change observable in a cell whose nucleus is about to divide is in the extra-nuclear protoplasm, the fibres of which arrange themselves radially around two points on opposite sides and at the circumference of the nucleus. Contemporaneously with this the nuclear network begins to change, and almost immediately afterwards the achromatic spindle-fibres appear in the nucleus.
These facts seem to point to the conclusion that the actual centre of force, of which the nucleus is the seat, divides first and is followed by the re-arrangement of the cell and nuclear protoplasm. Flemming considers that the nuclear network consists of an achromatic substance containing granules of chromatin which have the power of moving about in the network. These chromatic granules are fairly uniformly diffused in the resting nucleus, but in a nucleus preparing to divide they aggregate together in certain parts of the network. The parts of the network from which the chromatin has gone become inconspicuous and form the achromatic spindle-fibres, while the parts into which it has gone form the conspicuous deeply-staining rod-like fibres, so characteristic of a dividing nucleus. The achromatic fibres of the spindle which begin to appear at the first sign of the division of the nucleus are, on this view, parts of the nuclear network. With this view I entirely agree. The structure of the various phases of the nucleus of the ovum of Peripatus will bear the same explanation, allowing for this difference, viz. the amount of chromatic substance in the ovum of Peripatus is much smaller--so small, indeed, that even in the resting stage (PI. XII, fig. 8) the chromatin is absent from the greater part of the network, which thus has the pale appearance of the achromatic fibres of Flemming, an appearance which is only found in the dividing nuclei of the salamander. The reason why achromatic fibres are so ittle marked in the resting nuclei of most animal cells is that they are masked by the large amount of chromatic substance they contain.
This view of the spindle-fibres is not at all opposed to Strasburger's contention (15, fig. 44) that part of them are derived from the extra-nuclear spongework; for the nuclear and extra-nuclear spongework are, as I have already maintained, continuous with each other; in other words, part of the same system.
I have seen nothing of any process corresponding to the splitting of the fibres; but this is not to be wondered at considering that I have only twice found the spindle stage of the nucleus.
B. It is becoming more and more clear every day that the cells composing the tissues of animals are not isolated units, but that they are connected with one another. I need only refer to the connection known to exist between connective tissue cells, cartilage cells, epithelial cells, &c. And not onlymay the cells of one tissue be continuous with each other, but they may also be continuous with the cells of other tissues. For instance, I may refer to Fraipont's (2) work on the nervous system of the Archiannelida. He describes an intermuscular nervous plexus which is continuous with the muscle-cells and with the surface epithelial cells (2, Pl. 13, figs. 11, 16).
Instances of this kind might be multiplied from recorded observations, and are being multiplied day by day by histological observers to such an extent, that we are almost, if not quite, justified in regarding the body of an adult animal as a syncytium. It is true that the cells of the blood and lymph, and the ripe generative cells, are completely isolated. But the former, in their first stages of growth, form part of the syncytium; as in all probability do the latter also.
This continuity, which for a priori reasons we should expect, has hitherto been regarded as a fact of little morphological importance and relegated to the category of secondary features. The ovum, it is said, segments into completely isolated cells; and the connection between these is a secondary feature acquired late in development. It has always been considered that the first stage in the evolution of the Metazoa was a colonial Protozoon, i. e. a mass of perfectly isolated unicellular organisms derived by complete division from a single cell.
Now, while I do not wish to exalt the facts of the cleavage and early development of Peripatus above recorded to a position of undue importance, or to maintain that of themselves they are sufficient to destroy this conception of the origin and structure of a Metazoon, I think I am justified in pointing out that if they are found to have a general application, our ideas on these subjects and others connected with them will have to undergo a considerable modification.
The ancestral Metazoon will no longer be looked upon as a colonial Protozoon, but rather as having the nature of a multi-nucleated Infusorian with a mouth leading into a central vacuolated mass of protoplasm.
The continuity between the various cells of the adult--the connections between the nerves and muscles and sensory epithelial cells, receive an adequate morphological explanation; being due to a primitive continuity which has never been broken.
Herbert Spencer's view of the origin of the nervous system may perhaps not be so far from the mark as at first sight appeared. In any case the efforts to find out how the connection is established between the nervous and muscular tails of the ectoderm and endodenn of the lower animals should be transferred to the earliest phase of the embryo, i. e. to the segmentation stages.
Finally, if the protoplasm of the body is primitively a syncytium and the ovum until maturity a part of that syncytium, the separation of the generative products does not differ essentially from the internal gemmation of a Protozoon, and the inheritance by the offspring of peculiarities first appearing in the parent, though not explained, is rendered less mysterious; for the protoplasm of the whole body being continuous, change in the molecular constitution of any part of it would naturally be expected to spread, in time, through the whole mass.
In short, if these facts are generally applicable, embryonic development can no longer be looked upon as being essentially the formation by fission of a number of units from a single primitive unit, and the co-ordination and modification of these units into an harmonious whole. But it must rather be regarded as a multiplication of nuclei and specialisation of tracts and vacuoles in a continuous mass of vacuolated protoplasm.
At any rate I may safely say that, so far as the individual embryonic development of Peripatus is concerned, the connection of cell with cell is not a secondary feature acquired late in development, but is primary, dating from the very beginning of development.
Since making these observations on the syncytial nature of the cleavage and gastrula stage of Peripatus capensis, I have examined other segmenting ova to see if the fact was one of general application, with negative results.
The cells of segmenting ova are generally so closely applied together and the protoplasmic strands so hidden by food-yolk, that it is difficult to be certain of the point either way. But with ova in which the segments are slightly separated from one another--and I believe there are such though I have never seen them--the observation ought to present no special difficulty.
Indeed it is a well-known fact that an incomplete separation of the cells is found in the early stages of the segmentation of centrolecithal eggs; but it has always been assumed that this was a temporary phase, and that the segments eventually separated. We now know, thanks to the researches of Heathcote (4), that this separation does not occur in the centrolecithal egg of the Myrapod, Julus; and it seems to me extremely probable that his results for this form will be found on careful examination applicable to other similar ova.