100. Further Studies Upon the Regional Differentiation of the Inductive Capacity of the Organizer

Home , Dorsal lip, Regional differentiation

614 [Vol. 18,

100. Further Studies upon the Regional Differentiation of the Inductive Capacity of the Organizer.

By Yo Kaname OKADA. Tokyo Imperial University. Hirosi TAKAYA. Konan Koto-Gakko. (Comm.by N. YATSU,. M.LA. Oct. 12, 1942.) As stated in the preceding report of this series of experiments'', in the early gastrula of Triturus the dorsal wall of the archonteron possesses the head-organizing capacity, and the uninvaginated dorsal lip,, the trunk- or tail-organizing effect, the difference between the organizing effects of these two regions being quite distinct. Moreover, these effects are not influenced at all by the position of the graft in the host. Accordingly, there is reason to expect the formation of an entire embryo by transplanting pieces from these two regions together. To see if this actually takes place the following experiment was performed. I. As in Spemann's case, a section of invaginated and uninvaginated tissues was cut out in one piece from the region of the dorsal lip of embryos in stages 11, 12, and 13, flattened out, and transplanted. As the increased size of the graft could not be ignored, it was neces- sary to study beforehand its influence, if any, upon the outcome, To this end a test was made of the organizing capacity of separate grafts of approximately the abovee size, obtained respectively from the dorsal wall of the archenteron and from the uninvaginated dorsal lip of the blastopore of embryos of the same age. Regardless of the size of the grafts, the former always evoked 'the formation of a head, whereas the latter was confined to the organization of a trunk or a tail. The only variation in the result, as compared to the case of small-sized transplants, was the relatively greater tendency to produce trunk and tail at the same time. By the simultaneous transplantation of both invaginated and uninvaginated portions of the dorsal lip, whole embryos, provided with head, trunk, and tail, were obtained (Fig. 1 and 2). However, it has already been ascertained by a previous experiment (Okada and Takaya, 1942, Experiment III) that the coexistence of both headT and trunk- organizing regions in such pieces of the blastoporic lip is limited to stage 11 and stage 12. That in earlier stages prior . to the onset of invagination the head-organizing capacity is not yet in evidence, while in stage 13 the trunk- or tail-organizing capacity js manifested by the uninvaginated dorsal lip of course and likewise by the posterior end of the dorsal wall of the archenteron ; and finally, that there exists in stage 13 a regional difference characterized by the head-organizing

1) Okada, Y. K., and H. Takaya; Proc. 18 (1942), 605-513. No. 8.] Further Studies upon the Regional Differentiation, 616

Fig. 1 (left). Induction of an entire embryo by simultaneous trans~ plantation of both invaginated and uninvaginated portions of edge of dorsal lip of blastopore of an embryo of stage 11. Fig. 2 (right). Induction of an entire embryo by joint transplantation of separate pieces obtained from anterior and posterior portion of dorsal wall of archenteron of an embryo of stage 13.

Fig. 3 (left). Induction of tail alone despite joint transplantation of anterior portion of dorsal wall of archenteron of an embryo of stage 13 (head-organizing region) and uninvaginated portion of dorsal lip of blastopore of an embryo of stage 11. (tail-organizing region). Fig. 4 (right). Independent induction of head hnd of tail according to the respective organizing capacities of head- and tail-organizing tissues, as a result of their separation following joint transplantation. 516 Y. K. OKADAand H. TAKAYA. [Vol.18, anterior half and the trunk-organizing posterior half of the dorsal wall of the archenteron. Therefore, when the graft was secured from stage 13, it should have been impossible to call forth the formation of an entire embryo by means of the aforementioned operation, but it should be obtainable by rising the combined anterior and posterior parts of the dorsal wall of the archenteron. Nevertheless, it so happened that actually the formation of whole embryos occurred very rarely following the joint transplantation of head- and trunk-organizing areas, the percentage of induction being 25 per cent (3/12) for stage 11, 6 per cent (1/15) for stage 12, and 30 per cent (3/10) for stage 13. It must be admitted that this percentage is strikingly low in comparison to the cases of separate transplantation of the head- or the trunk-organjz- ing region. In the light of such results this experiment has shown the possibility of inducing an entire embryo. And again, it should be mentioned that in all cases of non-formation of whole embryos the induced body was comprised of trunk and other posterior structures (Fig. 3). Obviously, these were cases of complete failure of the head- organizer to express itself. As to the cause of this, it calls for further investigation. II. Thereupon, it was deemed advisable to make a careful study of the organizing effect of grafts which were combinations of head- organizing and trunk- or tail-organizing tissues. The three types of union attempted were : 1) the anterior and posterior parts of the dorsal wall of the archenteron of an embryo of stage 13, 2) the invaginated portion of the dorsal lip of stages 11 and 13, and 3) the anterior dorsal wall of the archenteron of stage 13 plus the uninvaginated dorsal lip region of stage 11. Even though the members of a pair may have differed as to the stage of their development, on the basis of previous results it is evident that one constituent always possessed the capacity of inducing a head and the other that of inducing a tail. In reality, however, the inducedd body was always morphologically merely trunk or further posterior parts, and never was it provided with a head as well (Fig. 3).. In addition to the majority of combinations having the members joined in the same plane, the method of placing the one upon the other was also tried with no dissimilarity in the outcome. To be sure, it is true that in the latter case the two pieces, by be- coming separated now and than, set up independent centers of induction, but the same also occurred, though on very rare occasions, in conjunction with the former procedure. Under such circumstances head and trunk or tail were developed according to the characteristic inductive capacity of the transplant, and because of the lack of any organic connection between the two structures, even when they came to be situated adjacent to each other, they could be distinguished at a glance from the induced whole embryos mentioned above (Fig. 4). These results clearly demonstrate that the grafting of a piece of the head-organizing area in union with trunk-organizing tissue causes the organizing effect of the former to be entirely suppressed, and that accordingly the infrequency of induction of whole embryos in the present experiments is not due purely to chance. In the previous report the No. 8.] Further Studies upon the Regional Differentiation. 517 question was raised whether or not the head-organizing effect possessed by the dorsel wall of the archenteron could be considered the same as the trunk- or tail-organizing effect possessed by other regions, and the facts disclosed here tend to suggest that the question was not ground- less. Moreover, when Hall (1937)1' undertook the regional interchange of organizer tissue, a graft of trunk-organizer, placed in the cephalic end, instead of being influenced by its position in the host, displayed its characteristic behavior, not only preventing the development of the host's brain, but also producing a tail-like process ; . in contrast, a piece of head-organizer, transferred to the trunk region, rather than causing the neural plate to enlarge in order to spur the development of a brain, readily adapted itself to its new location and gave rise to a normal neural tube. The loss of its organizing effect by the head- organizing tissues in union with trunk-organizing tissue in the present experiment may be imagined to follow from the reciprocating action of the two areas. If the participants of such a reciprocating action are equipotental, the suppressing effect should be revealed equally ii both regions. The fact is, however, that in these combinations although the head-organizing effect was easily checked, it offered no indication of having any influence upon the trunk-organizing effect. Thus, in the end, the combined effect of the paired organizers was as if it repre- sented the sole action of the trunk-organizer. III. It is a known fact that treatment with heat or alcohol causes the dorsal lip of the blastopore to lose the regional specificity of its organizing effect (Holtf reter, 1934)2). To be exact, although such treatment may not bring about any special modification of the inductive capacity itself, it does produce a very unmistakable change in the organizing effect. Thus, the uninvaginated portion of the dorsal lip of a gastrula, which had been exposed to the action of heat, evoked the formation of head organs only, never of trunk or tail (Figs. 5 and 7). Not only was this true, as a general rule, throughout all the gastrula stages, but also in stage 13 the portion of the dorsal wall of the archenteron near the blastopore, which in the living condition always induces trunk or tail, was made to manifest the head-organizing effect through the action of heat (Fig. 6). In the same way, treatment with alcohol caused the loss of the trunk- or tail-organizing effect, characteristic of the living tissue, and the acquisition of the capacity to form cephalic structures. Furthermore, the change in the organizing effect was complete, the induced body being limited to a head or head organs and never being accompanied by a trunk or a tail as well. Accord- ingly, this change may be considered to resemble very closely the change observed in the organizing effect before and after invagination. Indeed, in view of the transformation of the tail-organizing effect into the head-organizing effect, it may be that there is no difference what- ever between them. At the same time, it can not be said that even

1) Hall, E. K. 1937 Regional differences in the action of the organization center. Roux' Arch., 135, 671-688. 2) Holtf reter, J. 1934 Der Einfluss thermischer, mechanischer and chemischer Ein- griffe auf die Induzierfahigkeit von Triton-Keimteilen. Roux' Arch., 132, 223-306. 518 X. K. OKADAand H. TAKAYA. [Vol. 18,

after such treatment with heat the transplant still induced a morphologically perfect head, like that organized by living tissues of the dorsal wall of the archenteron~ for although the induced body consisted unquestionably of cephalic organs, not only was their arrangement at random in many cases, but also their number was often insufficient for a comnlete head (see Fig. 7). If this fact truly nfTrs p'rnnnds fnr

Fig. 5. (upper). Induction of cephalic organs by the transplantation of heat-killed tissue obtained from dorsal lip of blastopore of an embryo of stage 12. Fig. 6 (middle). Induction of cephalic organs by the transplantation of heat-killed tissue obtained from posterior end of dorsal wall of archenteron of an embryo of stage 13. Fig. 7 (lower). Transverse section through induced body of specimen shown in Fig. 5. Note eye in particular is evident in induced body. making a distinction between the head-organizing effect as revealed by the uninvaginated dorsal lip subjected to high temperatures end the same effect as evinced by living tissue of the dorsal wall of the archenteron, account must be taken of the difference in the organizing process of induction by what might be called dead tissues and of that by living inductors, as found in the dorsal lip of the blastopore. That is No. 8.] Further Studies upon the Regional Differentiation. 519 to say, the latter must possess the added capacity of placing organs in their regular position so that it does not stop simply at inducing organs, but goes further to arrange them properly. IV. The general tendency throughout the above experiments was the conversion of trunk- or tail-organizing effect into head-organizing effect. The possibility or impossibility of the reverse transformation is of even greater uncertainty. A single point in its favor was the occurrence of a filamentous outgrowth, morphologically a tail, which was induced by grafts of the dorsal wall of the archenteron, subjected to extremely high temperatures. However, the very marked reduction in inductive capacity following such treatment (Chung, 1939)1' does not permit of the conclusion that this reverse change is necessarily like the change observed in living tissue. In any case, that the organizing effect of the tissues of the dorsal lip region is not fixed and unchange- able is, at least, an assured fact. Furthermore, there seems to be no doubt that this change occurs within the tissue concerned in the course of development, and also that it can be traced to a material change. It is still a question, nevertheless, whether the different substance pro- duced by the material change determines the character of the trans- formed organizing effect, or the process of transition is in itself of importance. The investigation of this problem is now in progress.

1) Chung, H. 1939 Induktionsleistungen von frischen and gekochten Organteilen (Niere, Leber) nach ihrer Verpflanzung in Explantate and verschiedene Wirtregionen von Tritonkeimen. Roux' Arch., 139, 556-638.