Fig. 5. Dorsoventral patterning is established properly in Shh^-/-;Gli3^-/- and Smo^-/-;Gli3^-/- double mutants. (A-L) Coronal sections of wild-type and Shh^-/-;Gli3^-/- E10.5 embryos. The telencephalic morphology appears relatively normal in the mutants, except for the loss of dorsal midline structures (arrows). (A-L) Analysis of various homeodomain genes expression. Dlx2 expression assayed by RNA in situ hybridization (A,B) and Gsh2 expression assayed by immunofluorescence (C,D) show that normal ventrolateral patterning is established in the double homozygous mutants. Arrowheads delineate the boundary between lateral and dorsal regions in both wild-type and mutant embryos. Nkx2.1 expression also appears normal (E,F). Overlay of Nkx2.1 (red) and Dlx2 (green) RNA in situ hybridization on adjacent sections using Adobe Photoshop 4 (G,H) shows a similar nested pattern of expression for these two genes in wild-type and Shh^-/-;Gli3^-/- double homozygous embryos. This reveals the existence of ventral and lateral structures similar to the MGE and LGE in the wild-type embryos (asterisk and arrowhead, respectively). Gli1 expression is absent in Shh^-/-;Gli3^-/- double mutants (I,J), showing that the Shh pathway is not active. Brackets in I indicate Gli1 expression. (K,L) Ptch is expressed at low levels in Shh^-/-;Gli3^-/- mutants, with higher expression in ventral areas (bracket), similar to wild-type embryos. (M,N) Analysis of ventral genes expression in Smo^-/-;Gli3^-/- E10.5 embryos. As a result of exencephaly, the dorsal telencephalic structures are disrupted. Despite this, the ventral structures can still be analyzed. Nkx2.1 and Dlx2 are expressed in ventral areas of the telencephalon. Despite the exencephalic morphology of the mutants ventral pattern appears unperturbed, as evidenced by the presence of Nkx2.1 expression nested within the broader Dlx2 domain. This suggests that MGE and LGE cell fates are specified properly in Smo^-/-;Gli3^-/- double mutants.