Retina ganglion cell subtype specific cell death following optic nerve crush in mice

Purpose: Glaucoma is an optic neuropathy that causes cupping of the optic disc, retina ganglion cell (RGC) loss, and characteristic visual field defects. Published literature suggests differential RGC susceptibility to damage. However, the mechanisms by which the optic nerve and RGCs become more susceptible to injury and damage are largely unknown. We investigated individual RGC subtypes’ susceptibility to damage after optic nerve crush (ONC). Methods: We utilized two mouse strains that selectively express GFP in individual RGC subtypes: CB2-GFP strain (selectively expresses GFP in transient OFF-alpha RGCs) and TRHR-GFP strain (selectively expresses GFP in On-Off direction selective RGCs). ONC was performed unilaterally, with the contralateral eye serving as a control. RGC subtype specific damage was evaluated at 0, 1, 3, 7, and 14 days post ONC. RGC damage was assessed by immunofluorescence of labeled retinal flat mounts using the GFP biomarker for the specific RGC subtypes and NeuN for total RGCs. Results: Throughout the 14 day time course, GFP positive RGCs in the CB2-GFP strain died faster than the GFP positive RGCs in the TRHR-GFP strain, with similar rates of total RGC death in each strain. The half-life (T1/2) of GFP positive cells in the TRHR-GFP strain was T1/2=7.11 days with total RGC death T1/2=9.65 days. The half-life of GFP positive cells in the CB2-GFP strain was T1/2=4.19 days with total RGC death T1/2=10.77 days. There was a significant difference in percent cell survival of each individual RGC subtype at 3 days (TRHR-GFP, 61.3 +/- 7.4%; CB2-GFP, 38.2 +/- 14.3%; n=4, p=0.029), 7 days (TRHR-GFP, 63.1 +/- 26.4%; CB2-GFP, 22.2 +/- 5.3%; n=4-5, p=0.011) and 10 days (TRHR-GFP, 38.0 +/- 2.9%; CB2-GFP, 3.5 +/- 3.5%; n=4-5, p<0.001) post-crush. There was no significant difference in percent cell survival of each individual RGC subtype at 1 day (TRHR-GFP, 76.2 +/- 20.9%; CB2-GFP, 70.3 +/- 10.0%; n=4-5, p=0.621) and 14 days (TRHR-GFP, 5.0 +/- 6.5%; CB2-GFP, 2.6 +/- 3.3%; n=3-4, p=0.556) post-crush. Conclusions: These studies demonstrate differences in individual RGC subtype susceptibilities to ONC. These data provide valuable information to develop new targets to slow and/or prevent the progression of RGC damage and new methods to detect early damage in diseases such as glaucoma.