Figure 2

Isthmic derived Wnt signals are required at HH8 for maintenance of the early ophthalmic placode identity. In situ hybridization for Pax3 (blue) (B-E, G-J). GFP expression is shown in green (A, F and G). A dominant negative form of Wnt1 (DN Wnt1) was targeted to the right-hand side of the neural tube in all cases. Embryos were co-electroporated with GFP at HH7 (A) or HH10 (F, G) and analyzed 16 and 12 hours later until they reached approximately HH13. When Wnt ligands are perturbed at HH8, expression of Pax3 is reduced in the placodal ectoderm (C, black arrow head; E). The white arrowhead (B) indicates the normally developing placode on the un-injected side. When DN Wnt1 is injected at HH10 Pax3 expression in the placodes is not altered when analyzed at HH13 (G, H, J). The lines in (H) mark the extent through which Pax3 is expressed in the ophthalmic placodes on both sides. Embryos injected with dominant negative Wnt1 at HH8 (I) or HH10 (J) were sectioned to reveal the localization of Pax3 expression. Less Pax3 positive cells were observed in general, and within the mesenchyme, when dominant negative Wnt1 was injected at HH8 (I, right hand side). No obvious differences were observed when dominant negative Wnt1 was electroporated after HH10 (J). Asterisks delineate midbrain (G, H).