238 



Embryogenesis: Progressive Differentiation 



formations was laid down principally by 

 way of a disturbance of the gastrulation 

 movements. It is furthermore probable that 

 the absence or reduction of brain and sense 

 organs is due to deficiencies of the inductive 

 entomesoderm rather than to primary defects 

 of the ectoderm. There is no experimental 

 evidence in support of the assumption of 

 earlier teratologists that such gross anomalies 

 as microcephaly or cyclopia may arise from 

 a secondary degeneration of originally full- 

 size organ primordia. More generally stated, 

 these malformations are not a problem of 

 abnormal growth but of cell movements and 

 cytological determination. The embryo may 

 possess the cell material necessary for the 

 formation of all ectodermal organs, yet these 

 organs will never begin to differentiate un- 

 less their primordia have been mapped out 

 and fixed in their fate by the appropriate in- 

 ductors. 



STATE OF ORGANIZATION OF THE 

 EARLY GASTRULA 



The data presented so far have dealt with 

 the problem of embryonic organization in 

 rather generalized outlines. Further analysis 

 of these phenomena requires a more detailed 

 study of the determinative relations between 

 the parts of the early embryo. With this aim 

 in mind, three methods which supplement 

 each other have been employed: isolation, 

 transplantation and defect experiments. Such 

 experiments have been made possible through 

 the introduction of fine glass needles as 

 microsurgical instruments (Spemann, '18, 

 '21b) and through the use of a balanced 

 physiological salt solution (henceforth re- 

 ferred to as "standard solution") for cultur- 

 ing the embryonic fragments (Holtfreter, 

 '31, '43a). 



ISOLATION EXPERIMENTS SHOWING 



THE DIFFERENTIATION CAPACITIES 



OF PARTS OF THE GASTRULA 



Isolation experiments have been carried 

 out on a large scale on the early gastrula of 

 both urodele and anuran species (Holtfreter, 

 '38b, c). We shall confine ourselves to the 

 conditions in urodeles. As the preceding 

 chapter has shown, the gastrula is of particu- 

 lar interest since it is mainly during this 

 stage that the basic pattern of organization 

 is laid down. 



The gastrulae were divided up into small 

 fragments which were then cultured in 

 standard solution. The main types of differ- 

 entiation obtained from hundreds of such 



explants as projected back upon the gastrula 

 are represented in Figure 80b. This distribu- 

 tion pattern is markedly different from the 

 map of the prospective organ areas (Fig. 

 80a), indicating that only few of these areas 

 develop true to their prospective significance 

 when they are explanted. Three kinds of 

 behavior can be distinguished which cor- 

 respond more or less to the regions of the 

 ectoderm, the marginal zone and the vegetal 

 hemisphere, respectively. 



The Ectodermal Region. In accordance with 

 the observations on total exogastrulae, the 

 different isolated regions of the ectodermal 

 layer are incapable of forming typical tissues 

 but form merely unspecialized epidermis 

 cells. This holds for both the prospective 

 epidermal and the prospective medullary 

 areas and applies to the axolotl and to vari- 

 ous European, American and Japanese Tri- 

 turus species (Holtfreter, '33a, '38b, '44b; 

 Waddington, Needham and Brachet, '36; 

 Shen, '39; Gallera, '48; Yamada, '50a). When 

 supported by a solid surface such as glass, 

 a piece of ectoderm may spread to form an 

 epithelium which will, however, disintegrate 

 shortly into single cells. In order to maintain 

 an epithelial continuity, ectodermal cells 

 require the presence of connective tissue 

 and a basal membrane (Holtfreter, '34c, 

 '39b). 



The failure of the explants to form typical 

 ectodermal derivatives cannot be ascribed 

 to inadequacies of the culture medium but 

 is a true expression of their state of deter- 

 mination at the time of isolation. This is 

 evidenced by the fact that when correspond- 

 ing ectoderm pieces are isolated from a 

 neurula, they do develop into different tis- 

 sues whose type conforms more or less to 

 the prospective significance of the areas 

 tested (Holtfreter, '31; Mangold, '33b; Mon- 

 roy, '37; von Aufsess, '41). Thus an explant 

 taken from the medullary plate of a neurula 

 will no longer form epidermis but neural 

 tissue, epidermal differentiations being now 

 confined to explants derived from the pre- 

 sumptive epidermis. Obviously, the invagi- 

 nated tissues have changed the determinative 

 state of the neurula ectoderm. 



The Dorsal Marginal Zone. Explants taken 

 from the different sectors of the marginal 

 zone of urodele gastrulae are capable of 

 developing into a great variety of tissues, 

 but their histological fate does not conform 

 exactly to the prospective significance of 

 the areas tested. Most illuminating is the 

 behavior of explants from the upper blasto- 

 poral region. They undergo vigorous move- 



