1. At the feeding stage (st. 38), a high percentage (79 %) of Pleurodeles homozygous ac/ac larvae show bent tails after a persistent ascitic blister in the dorsal part of the fin, when embryonic development occurred at 12°C; about only 25 % of them are affected by abdominal and pericardic ascites; about 40 % can feed and survive. The larval phenotype is very different when embryonic development occurred at 23 °C, in which case tail growth appears to be normal, but 95 % larvae die, due to ascitic fluid collection in the abdominal and heart regions, marked anaemia and microcephaly.
2. The exchange of posterior neural plates and dorso-lateral epidermis between normal and mutant neurulae has shown that the localization of the blister in the dorsal fin is not dependent on autonomous properties of the mutant dorsal tissues, but should be considered as resulting from general disturbances in the mutant organism.
3. Experiments were performed, involving a temperature shift from 12 to 23°C or 23 to 12°C, occurring at various developmental stages from the end of gastrulation (stage 13) to the stage of spontaneous embryonic muscle contractions (stage 26). When the temperature shift was applied after the end of neurulation (stage 21), the caudal phenotype was statistically similar to that of larvae which had been bred continuously at the first temperature. Thus temperature-sensitive phases can be characterized between neurula stages 15 and 18 (for a 12–23° shift) or 15 and 21 (for a 23–12° shift). Similarly, abdominal ascites can be induced when embryos are kept at 23 °C till stage 23 (early tail-bud) only, and occurs much less frequently when embryos are kept at 12°C till stage 23 and then transferred to 23°C.
4. It could be concluded from these experiments that the caudal mutant phenotype is already temperature-determined during neurulation, before stage 21.
Nevertheless, double temperature-shift experiments showed that the second shift could modify the results which would be obtained if the first shift only occurred. Paradoxical results were obtained, more than 90 % of the tail phenotypes being of the ‘warm type’ when the embryos were first kept at 12°C, then shifted up to 23 °C between stages 22 and 26, and shifted down again to 12°C. Such a treatment markedly lowers the percentage of bent tails (‘cold type’) from the percentage which would occur if ac/ac embryos were constantly kept at 23 °C after stage 21, but this longer warm treatment is of no effect of itself as compared to the case when the whole development occurs at 12°C (bent tails are predominant in this latter case).
Thus, whereas the early determination of the position of the caudal blister can be considered as a stable phenomenon under given temperature conditions, it is not irreversible.
5. As compared to cold-bred larvae, thrice as many completely anaemic larvae (66 %) were obtained from ac/ac embryos kept at 23 °C between stages 21 and 26; this offers an opportunity for the experimental study of this anaemia.
6. Implications of these results for further analysis of temperature-sensitive mutations in cold-blooded vertebrates are suggested.
Neural-fated self-renewing cells regulated by Sox2 during secondary neurulation in chicken tail bud
