Figure 3.

IO neurons cross the midline in Slit and Robo mutants. A–C, Coronal sections of E13 wild-type (A, B) and Robo1 ^−/− ;Robo2 ^−/− (C) embryos immunolabeled with anti-Brn3.2 and anti-Pax6 antibodies. In both types of embryos, IO neurons migrate in the submarginal stream (arrows) and express only Brn3.2, whereas LRN neurons migrate in the marginal stream (arrowhead) and express only Pax6. Note that Brn3.2-positive cells are observed in the midline of the Robo1 ^−/− ;Robo2 ^−/− embryo (asterisk in C) and not in wild type (A, B). D–K, Coronal sections at the level of the IO of E13 embryos of Slit and Robo compound knock-outs immunolabeled for Brn3.2 (D–K) and counterstained with Hoechst (E–K). Brn3.2-positive IO neurons are found in the floor plate of Slit2 ^−/− (G), Robo2 ^−/− (I), Slit1 ^−/− ; Slit2 ^−/− (J), and Robo1 ^−/− ;Robo2 ^−/− (K) embryos but not in wild type (D, E), Slit1 ^−/− (F), or Robo1 ^−/− (H) mutants. In all cases, LRN neurons cross the midline. L, Quantification of the number of Brn3.2-positive IO neurons in the midline of Slit and Robo mutants (see Materials and Methods). n is the number of animals analyzed for each genotype. NS, Nonsignificant. *p = 0.05; **p = 0.01; ***p < 0.005. Error bars indicate SEM. Scale bars: A–C, 175 μm; D–K, 65 μm.