Fig. 63.—A. After Wetzel. Section through an egg (blastula stage) reversed at two-celled stage. B. After Schultze. Double embryo, from reversed two-celled stage, united ventrally. C, C¹. Two views of another double embryo (united dorsally). C². Cross-section through last. D. After Wetzel. Double embryo united laterally. D¹. Section through same.
Roux interprets Hertwig’s results as due to the sudden partial post-generation of a part of the injured half of the egg. He thinks that a half-embryo had first developed, and then to this there has been quickly added a part of the missing side. This reply fails, however, to meet Hertwig’s description of the method of development of the embryos. Later work, however, has put us in a position to give a more satisfactory account of the differences between the results of Roux and Hertwig. It seemed to me that the two kinds of embryos might be due to the different positions of the eggs after the operation. It had been shown by Schultze (’94) that if a normal egg in the two-celled stage is turned upside down and held in that position two embryos develop from the egg ([Fig. 63], B, C, D). These embryos are united in various ways, and arise presumably one from each of the first two blastomeres. These results have been confirmed by Wetzel, who examined more fully into the early development of the twin embryos. He showed with much probability that the protoplasm rotates in each blastomere, so that in many cases the lighter part flows, or starts to flow, toward the upper hemisphere of the egg. In this way similar protoplasmic regions of the two blastomeres may become separated, and under these circumstances each blastomere gives rise to a whole embryo. A cross-section through one of the segmentation stages of one of these eggs is shown in [Fig. 63], A. The smallest cells are found at the outer side of each half, and the two segmentation cavities lie one in the upper region of each hemisphere. Some of the different kinds of embryos that develop from inverted eggs are shown in [Fig. 63], B, C, D. They are united in [Fig. 63], B, by their ventral surfaces, and in [Fig. 63], C, C¹, C², by their dorsal surfaces, and in [Fig. 63], D, D¹, at the sides. These differences are probably accounted for by the different ways in which the protoplasm of the first two blastomeres rotated before the egg divided.
A consideration of these results led me to carry out the following experiment on eggs operated upon by Roux’s method. After sticking one of the first two blastomeres, some of the eggs were placed so that the uninjured blastomere kept its normal position, i.e. with the black hemisphere upward. Other eggs were turned, so that more or less of the white hemisphere was upward. From the two kinds of eggs two kinds of embryos were obtained. From those with the black hemisphere upward the embryo was a half-embryo like that described by Roux, while from the eggs with the white hemisphere upward embryos developed that were in many respects whole embryos of half size.[112] The explanation of this difference will be obvious from what has been said. When the black hemisphere is uppermost the contents of the uninjured blastomere remain as in the normal egg, and a half-embryo results. When the white hemisphere is uppermost the contents of the uninjured blastomere rotate, so that it generally shifts its relation to the protoplasm in the other injured half, and a whole embryo develops, as in Schultze’s experiment. In one case I obtained a half-embryo from an inverted egg. The result did not appear to be due to a lack of rotation of the protoplasm, because the medullary folds were white, showing that the protoplasm must have changed its position. The result can possibly be explained as due to the protoplasm rotating in each blastomere along the line between the halves, so that it still retains the same relation as that of the normal two-celled stage.
The whole embryos of half size are generally imperfect in certain respects on account of their union with the other half. They resemble in all important points the embryos described by Hertwig, and I see no grounds for interpreting them as embryos of a meroblastic type, but rather as whole embryos of half size, whose development posteriorly and ventrally has been delayed or interfered with by the presence of the other blastomere.
It has not been possible to separate the first two blastomeres of the frog’s egg, for if one is removed the other collapses. In the salamander, that has a mode of development similar to that of the frog,[113] it has been possible to separate the first two blastomeres. Herlitzka, who carried out this experiment, found that each blastomere gives rise to a perfect, whole embryo of half size. We cannot doubt, I think, that the same power of producing a whole embryo is also present in each of the first two blastomeres of the frog’s egg. When the two remain in contact in their normal relation to each other, each produces only a half; when like regions of the two blastomeres are separated, each produces a whole embryo. Thus we see that whatever the factors may be that determine the development of a single embryo from the egg, still each half, and perhaps each fourth also, has the power of producing a whole embryo.
In later papers Roux has stated that he had also, even in his earlier experiments, found other kinds of embryos than the half-embryos that he described. Some of these were whole embryos that had developed from the uninjured blastomere without the injured one taking any part or only a very small share in their formation. He found, he states, all stages between those embryos that had used up all the yolk material of the injured side (though post-generated) and those that had not used any part of it. The latter kind of embryo he does not recognize as a whole embryo of half size in the sense that a single blastomere has developed directly into a smaller whole embryo, but he believes that there must have been formed at first a half-blastula, half-gastrula, half-embryo, and that the last stage completed itself laterally without using any material from the injured half. That the uninjured blastomere may at first segment as a half is not improbable, but that whole embryos are formed only by the formation of new material at the side of a half-embryo is, I think, hardly possible, since the results of Schultze, Wetzel, Hertwig, and myself show that a whole embryo may develop directly out of the material of a single blastomere.
Spemann (1900) has carried out some novel experiments on the eggs of triton, and has shown how in another way double structures may be produced. If a ligature is tied loosely around the egg at the first cleavage exactly along the division plane between the first two blastomeres, it will be found later that the long axis of the single embryo lies, in the great majority of cases, across the ligature, and only in a small percentage of cases does the median plane correspond with that of the ligature, and, therefore, with the first cleavage plane.
If one of the latter eggs is allowed to develop to the blastula stage, and the ligature is then drawn tighter, so that the blastula is completely constricted, an embryo develops from each half.
If one of the former eggs is allowed to develop to a stage when the medullary plate is laid down, but is not yet sharply marked off, and the ligature is then tightened, there will be formed (the plane of constriction being across the medullary plate) from the anterior part a normal head with eyes, nasal pits, ears, and a piece of the notochord, and from the posterior part there will be formed, at its anterior end, another new head just behind the ligature. Ear-vesicles develop in this part at the typical distance from the anterior end. The brain that develops has a typical cervical curvature, and eye evaginations appear at the anterior end. The chorda, that extended at first to the anterior end of this region, is partially absorbed.
If the ligature is drawn tighter at a later stage, when, for instance, the medullary plate is plainly visible but is still wide open, a different result is obtained. The posterior part no longer forms a new head at its anterior end, but develops into those structures that it would form normally. In some cases it was found that the region from which the ear develops had been pinched in two, and in consequence a small vesicle appears in front of the constriction and another behind it.