Figure 3

ZLI formation occurs independently from ventral tissues after stage HH10 and ZLI morphogenesis depends on the integrity of its flanking regions. Diencephalic explants were taken from (A-F) stage HH10 and (G-J) stage HH14 embryos and were analysed by in situ hybridisation for the expression of Shh (a-d, g-j) and Lfng (E, F) at the time of dissection (pre-cult.) or after culture for 48 (B, D, J) or 24 hours (F) (post-cult.). (A, B) Explants comprising pro-Pth, pro-ZLI, pro-Th and basal tissue (red). Note basal expression of Shh at the time of dissection and formation of a ZLI after culture (red arrowhead). (C, D) Pro-Pth + pro-ZLI + pro-Th explants excluding basal tissue. Note absence of Shh expression at the time of dissection and induction of a ZLI after culture. (E, F) Explants as in (C, D). Note formation of a Lfng-free stripe of cells after culture (red arrowhead). (G, H) Pro-Pth + pro-ZLI explants; (I, J) pro-ZLI + pro-Th explants. Note induction of Shh in a patch, rather than a line, after culture (compare to (B, D)).