Abstract
Purpose :
While it is well known that pathological consequences of glaucoma involve degeneration of retinal ganglion cells (RGCs) and their axons, increasing evidence has pointed to selective vulnerability of specific RGC types. We have previously shown that microglia activation increases following glaucomatous injury, but their role in synapse disassembly in the inner plexiform layer (IPL) and specific RGC type vulnerability is unknown. Here we determined whether microglia are positioned to play a role in the selective vulnerability of alpha OFF-transient RGCs.
Methods :
Laser-induced ocular hypertension (LIOH) was performed unilaterally on adult CD-1 mice expressing Cx3cr1-GFP, in which microglia are fluorescently labeled. Mice were sacrificed 7 days-post LIOH and individual RGCs were labeled by ballistic delivery of dextran dye. Immunolabeling was used to detect expression of postsynaptic protein, PSD95. Z-stacks were acquired using confocal microscopy. Microglia contact with individual RGCs was analyzed with VolumeCut (https://lucadellasantina.github.io/VolumeCut/) and colocalization of PSD95 with RGCs and microglia were evaluated throughout the IPL using ObjectFinder (https://lucadellasantina.github.io/ObjectFinder/). Statistics were performed using the Wilcoxon rank-sum test and one-way ANOVA.
Results :
After IOP elevation, contact points made between microglia and individual alpha RGCs increased for OFF-sustained (OFF-S), OFF-transient (OFF-T), ON-sustained (ON-S), and ON-transient (ON-T) RGCs (42.5%, p=0.34; 189%, p=0.03; 264%, p=0.02; 215%, p=0.003, respectively). The percent change was significantly greater in OFF-T compared to OFF-S RGCs (p=0.04). This increase in contact points also resulted in greater colocalization of microglia with PSD95 sites on individual RGCs: for OFF-S (75.4%, p=0.02), OFF-T (298%, p=0.02), ON-S (155%, p=0.06), and ON-T (282%, p=0.007). The percent change was significantly greater in OFF-T compared to OFF-S (p=0.03), and in ON-T compared to ON-S (p=0.04).
Conclusions :
Following IOP elevation, microglia in the IPL increase their contact with individual RGCs. Microglia contact favored OFF-T over OFF-S RGCs and targeted PSD95 sites to a greater degree in the transient vs. sustained RGC types, suggesting that microglia may play a role in the selective vulnerabilities of specific RGC types.
This is a 2021 ARVO Annual Meeting abstract.