Figure 4

Nr2e1 ^frc/frc P7 retinas had increased numbers of glycinergic amacrine cells. Transverse retinal sections from P7 Nr2e1 ^+/+ and Nr2e1 ^frc/frc mice were immunostained for the pan-amacrine markers Pax-6 and syntaxin-1A, and the subclass markers Islet-1/2, GABA, calretinin, and glycine transporter 1 (GlyT1). a Retinal section showing amacrine cells immunostained with Pax-6 (red) and syntaxin-1A (green). Nr2e1 ^frc/frc retinas had more Pax-6 and syntaxin-1A positive cells than wild-type retinas, and these cells occupied the majority of the INL. An ectopic plexiform layer (EPL) is also indicated (open arrow). Magnification of the Pax-6 (red) and syntaxin-1A (green) staining is shown for a region of b Nr2e1 ^+/+ or c Nr2e1 ^frc/frc retina (rectangle). d-f Neither (d) Islet-1/2 positive (green) (arrows), (e) GABA positive (red), or (f) calretinin positive amacrine cells appeared increased in Nr2e1-mutant retinas. g In contrast, GlyT1 positive amacrine cells appeared increased in the INL of Nr2e1-mutant retinas. EPL (open arrow). h Amacrine cells were counted in the GCL throughout five sections across the retina of Nr2e1 ^+/+ and Nr2e1 ^frc/frc mice. Numbers were normalized to retinal length and expressed as percentages of Nr2e1 ^+/+ cell numbers. Increased numbers of amacrine cells (assessed as Pax6 positive minus Brn3a positive) were observed in the GCL of Nr2e1-mutant retinas compared to wild type (45 % increase). n = 3 for Nr2e1 ^+/+, n = 3 for Nr2e1 ^frc/frc; *, P ≤ 0.05; error bars represent SEM. EPL, ectopic plexiform layer; GCL, ganglion cell layer; Hoechst, nuclear counterstain (blue); INL, inner nuclear layer; scale bar = 50 μm in all images except for B where it represents 12.5 μm