Fig. 7. Axon guidance factors are relatively unchanged in the optic chiasm of Foxd1 (BF-2) deficient embryos. (A) In situ hybridization for ephrin-B2 in frontal cryosections of E15.5 Foxd1^+/+ (a) and Foxd1 mutant (b) embryos, revealing similar expression pattern in cell bodies at the floor of the third ventricle, thought to belong to the midline radial glia. (B) In situ hybridization for Slit2 in horizontal cryosections at E13.5 (a-d) and E15.5 (e-h). Comparison of Foxd1^+/+ (left panels) and Foxd1^lacZ/lacZ (right panels) chiasms at E13.5 show that although Slit2 expression appears normal in more dorsal sections (a,b), Slit2 levels are increased in the region ventrocaudal (c,d) to the chiasm in Foxd1 deficient embryos (red arrows, d). At E15.5, the same pattern is maintained (e-h). Green dashed lines indicate the position of retinal axons in each section. (C) Immunostaining with anti-CD44/SSEA antibodies (red) and anti-neurofilament (green) in horizontal cryosections at E13.5 (a,b) and E15.5 (c,d). Comparison of Foxd1^+/+ (left panels) and Foxd1^lacZ/lacZ (right panels) chiasms indicates that although CD44/SSEA expression appears normal in Foxd1 deficient embryos, retinal axons enter the CD44/SSEA-positive area (d), in contrast to the wild type in which axons never transgress the CD44/SSEA zone (c). Green indicates retinal axons labeled with neurofilament antibodies. Red arrows indicate that retinal axons invade the CD44/SSEA expression zone. Scale bars: 200 µm. (D) Schematic indicating that in the chiasm, the Foxg1 territory expands in the absence of Foxd1, as in retina, and retinal axons aberrantly course through the CD44/SSEA zone. Islet1 and Zic2 expression territories are reduced around the chiasmatic midline, and Slit2 expression is expanded ventrocaudally. Ephrin-B2 and CD44/SSEA expression appear unaltered in the Foxd1 deficient chiasm; however, in contrast to wild-type axons, Foxd1 deficient RGCs trespass the CD44/SSEA domain.