In control retina, immunohistochemistry for GyP revealed strong staining in the GCL, a scattered population of amacrine cells at the inner aspect of the INL and in a sharply defined lamina within the IPL, assumed to be a synaptic layer (Figs. 3A–F). Weaker staining was present throughout the IPL and the nerve fiber layer (NFL). In diabetic retina, the general pattern of GyP staining was similar to controls; however, more amacrine cell bodies showed strong staining for GyP and the overall intensity of staining in the IPL was greater than in control retina (Figs. 3B, 3C). Notably the GyP-positive amacrine cells in the diabetic retina showed strong staining within cytoplasmic distensions at the side of the nucleus facing the INL (Figs. 3B, 3C); these structures demonstrated identical morphology to the perinuclear glycogen stores observed in
PAS-stained preparations. The additional control eyes preserved with DF showed superior morphology than the formalin-fixed diabetic and control eyes, as well as improved immunostaining for GyP (Figs. 3D–F). Following fixation in DF, a much larger population of amacrine cells were seen to express GyP in the control retina (Fig. 3D), and GyP immunoreactivity in ganglion cell bodies and dendrites, and axons in the NFL was more evident (Figs. 3DF,Supplementary
Fig. S5D).