Fig. 3. The ventral diencephalic alar plate is sufficient to maintain a program of ZLI differentiation. (A) Strategy to generate stage 13-14 forebrain explants containing the dorsal two-thirds (D2/3) or dorsal half (D1/2) of the diencephalon. Incisions made dorsal to the visible morphological sulcus at the alar-basal plate boundary produced D2/3 and V1/3 explants, whereas D1/2 and V1/2 explants were generated by bisections of the neural tube. ZLI differentiation was monitored by Shh (purple) and Ptc2 (yellow) expression after 48 hours in culture. (B-I) D2/3 explants (D,E), but not D1/2 explants (H,I) can form a ZLI after 48 hours in culture (t=48 hours), as assessed by Shh (D,H) and Ptc2 (E,I) expression. Neither D2/3 explants (B,C) nor D1/2 explants (F,G) contained basal-plate tissue, as assessed by Shh (B,F) and Ptc2 (C,G) expression immediately after dissection (t=0 hours). V1/3 explants, which correspond to the ventral tissue excised from D2/3 explants, expressed the basal-plate markers Shh (J,L) and Ptc2 (K,M) at the time of dissection (J,K) and after 48 hours (L,M). Lfng expression was used to monitor the quality of the explants (red).