Figure 6: The Sema3E receptors (PlexinD1/Neuropilin1/VEGFR2) expression in developing MAP6-KO brain and in cultured subicular neurons.

(a) Coronal brain slices of WT and MAP6-KO E17.5 embryos were hybridized with probes for PlxD1. PlxD1 mRNA is expressed in the subiculum and in the ventrolateral part of the cortex including the piriform cortex and the perirhinal cortex. Scale bar, 100 μm. S=subiculum; CA1=Ammon’s Horn; cp=cerebral peduncle; Rs=retrosplenial cortex; Pir=piriform cortex; PRh=perirhinal cortex. (b) Binding of sema3E along the fornix. Schematic diagram of a sagittal section with major neuronal tracts indicated (colour code in supplementary Fig. 3). The square indicates the position of microscopic field shown below. The atrophic fornix post-commissural part (pf) observed in MAP6-KO embryos binds Sema3E-AP. (c) Immunolabelling of subicular neurons using anti-PlxD1, -Nrp1 and -VEGFR2 antibodies. Histogram shows quantification of the relative immunofluorescence signal. The values represent the mean pixel intensity of the staining (number of neurons≥60). Scale bar, 20 μm. (d) Immunolabelling of growth cones from subicular neurons using anti-phosphotyrosine antibody to Tyr1175 in VEGFR2 after 15 min of treatment with 10 nM of Sema3E. Histogram shows the mean pixel intensity of phospho-VEGFR2 staining relative to CTL conditions (number of growth cones≥20). Scale bar, 10 μm. Error bars, s.e.m. *P<0.05 Student’s t-test.