Neural crest cells can be induced by an interaction between neural plate and ectoderm. To clarify the timing and nature of these inductive interactions, we have examined the time of competence of the neural plate to become neural crest as well as the time of neural fold specification. The neural plate is competent to respond to inductive interactions with the nonneural ectoderm for a limited period, rapidly losing its responsive ability after stage 10. In contrast, nonneural ectoderm from numerous stages retains the ability to induce neural crest cells from competent neural plate. When neural folds are explanted to test their ability to produce neural crest without further tissue interactions, we find that folds derived from all rostrocaudal levels of the open neural plate are already specified to express the neural crest marker Slug. However, additional signals may be required for maintenance of Slug expression, since the transcript is later down-regulated in vitro in the absence of tissue interactions. Taken together, these results suggest that there are multiple stages of neural crest induction. The earliest induction must have occurred by the end of gastrulation, since the newly formed neural fold population is already specified to form neural crest. However, isolated neural folds eventually down-regulate Slug, suggesting a second phase that maintains neural crest formation. Thus, induction of the neural crest may involve multiple and sustained tissue interactions.
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