Fig. 4. Reducing FGF signaling from the isthmic organizer results in cell death in the anterior mesencephalon. (A-E) Lateral views of embryos of the genotypes indicated, collected at the somite stages denoted and labeled with LysotrackerT (LysoT), which marks regions in which dead cells are abundant. The approximate position of the isthmic constriction (Is) is indicated. (C') The embryo shown in C was fixed after labeling with LysoT and assayed by RNA in situ hybridization for Pax6 expression. The broken yellow line indicates the posterior limit of Pax6 expression, which lies at the border between the prospective forebrain (diencephalon, Di) and midbrain (mesencephalon, Mes). In control embryos (A,D), LysoT labeling is detected where neural crest (NC) is migrating into the first branchial arch (BA1), but there is relatively little labeling in the neuroepithelium. By contrast, in the mutant embryos (B,C,E) there is extensive LysoT labeling in the mes (white arrows). (F-K) Embryos of the genotypes indicated, collected at the somite stages indicated, were assayed in whole mount by RNA in situ hybridization using a probe for Spry1 or Efna2. The extent of the expression domain from the mes/r1 boundary (red circle) towards the forebrain is indicated by a broken red arrow.