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Microglial activation and interaction at synaptic sites in experimental glaucoma.

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Posterboard#: B0150

Abstract Number: 672 - B0150

AuthorBlock: Alfred K. Yu1, Kelly Mai1, Esther Choe1, Alan Tran1, Luca Della Santina1, Yvonne Ou1
1Ophthalmology, University of California, San Francisco, San Francisco, California, United States;

DisclosureBlock: Alfred K. Yu, None; Kelly Mai, None; Esther Choe, None; Alan Tran, None; Luca Della Santina, None; Yvonne Ou, None;

Purpose
While the mechanisms of retinal ganglion cell (RGC) degeneration in glaucoma are complex and incompletely understood, evidence suggests an early loss of synaptic components on RGC dendrites following injury. We tested the hypothesis that microglial activation is playing a role in synaptic pruning of RGC dendrites using an experimental model of glaucoma.

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 in the retina by ballistic delivery of fluorescent dextran dye. Immunolabeling of PSD95 was used to detect excitatory postsynaptic sites. Z-stacks were acquired using confocal microscopy, and individual RGCs and microglia were skeletonized to produce binary masks. PSD95 puncta were identified automatically using ObjectFinder (https://lucadellasantina.github.io/ObjectFinder/). Individual alpha RGCs were analyzed in each condition and statistics were performed using the two-tailed Student’s t-test.

Results
As a result of LIOH, there was an increase in microglia volume (307%; p=0.01), the volume overlap between RGCs and microglia (200% increase; p=0.015), and the percentage of RGC colocalized with microglia (291%; p=0.01), compared to contralateral control. On average, there was a trend of increased rate of microglial colocalization with PSD95 sites on the RGC dendrites following LIOH (1300% increase; p=0.06).

Conclusions
Microglial activation is occurring in the inner plexiform layer of the retina following LIOH as evident by an increase in microglia volume. The increase in overlap between microglia and RGC dendrites and the increase in rate of microglia colocalization with PSD95 sites on RGC dendrites suggest that microglia are possibly pruning synaptic sites following glaucomatous injury.

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