Sampling method for the counting of RGCs in Group 1 wholemounts (A) and Group 2 tissue sections (B).

RGC counts in retinal wholemounts were made in sampling boxes (0.260 mm2) along quadrant radii at 1.5 mm, 2.5 mm and 3.5 mm from the optic nerve head (white boxes; A). RGCs were counted in sections over 250 µm length regions at a 200–300 µm distance from the centre of the optic nerve in radial retinal sections (white lines; B). For panel A, Brn3a (red) was used to stain the retinal wholemount and RGCs (scale bar: 1 mm). For panel B, Brn3a (green) was used to stain RGCs, GFAP (red) to stain glia and DAPI (blue) was used as a nucleus stain (scale bar: 250 µm).