Fig. 3. Zic2 and EphB1 are expressed in the same RGCs in the ventrotemporal (VT) crescent. (A) Spatiotemporal representation of Zic2 (red) and EphB1 (green) in mouse retina (gray). Zic2 expression is first seen in VT retina at E14.5. Expression peaks at E16.5 and then Zic2 is downregulated by E17.5-18.5. EphB1 expression in the VT retina commences at E14.5, lasting until E17.5-18.5. EphB1 expression then expands into the entire retina in retinal cells other than RGCs. EphB1 expression is also observed in the central retina from E13.5-15.5. (Ba-c') Immunostaining against Zic2 in E15.5 retinas (a,a', red) combined with in situ hybridization for EphB1 (b,b', green) in the same section, indicates that Zic2 is expressed in the most-peripheral (younger) cells, whereas older and more-central cells co-express Zic2 and EphB1. (a',b',c') Higher magnification of the boxed area in c. Red line in c' indicates cells positive only for Zic2, which are presumably younger cells. Yellow line indicates the Zic2 and EphB1-positive region, in which more-mature cells are located. Scale bars: in a, 100 µm for a,b,c; in a', 30 µm for a',b',c'.