Early Development

Treatment with retinoids at gastrulation stages results in the progressive loss of head structures (11). In this case, stage 10 embryos are treated by taking an aliquot of tRA in DMSO and simply adding it to the medium in which the embryos are cultured to produce a final concentration of 10-5 M or 10-6 M. A typical treatment time would be for 30 min, at the end of which the medium is changed.

Virtually all studies on early development of amphibians have used this technique because of its speed and simplicity. One experiment, however, involved the injection of RA suspended in corn oil (32). Stage 10-11 embryos had their fertilization envelope manually removed, and were injected with a 1-nL droplet between the surface and deep ectodermal layers by means of an air pressure injection system. The droplet also contained a fluorescent dye, Dil, so its presence could be traced throughout further development of the embryo. The additional unique feature of this system was that the effects of RA were asymmetric because only one side of the embryo was injected.

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