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Sandeep Kumar, Abram Akopian, Stewart A Bloomfield; Secondary cell death via gap junctions mediates the loss of amacrine cells in a mouse model of experimental glaucoma. Invest. Ophthalmol. Vis. Sci. 2015;56(7 ):4253.
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Bystander cell death via gap junctions (GJs) results in the extensive loss of amacrine cells (ACs), secondary to the loss of retinal ganglion cells (RGCs) they are coupled to, under a number of pathological conditions including apoptotic, excitotoxic, and ischemic insult. Here we studied the role of GJs in the loss of ACs in an experimental mouse model of glaucoma.
Experimental glaucoma was induced in WT and connexin KO mice by intracameral injections of polystyrene microbeads. Neurobiotin (NB) retrogradely labeled RGCs as well as ACs to which they are coupled via GJs. RGCs were also labeled with Brn3a antibody. Vertical retinal sections were also counterstained with DAPI to determine total nuclear counts.
A comparison of NB, DAPI, and Brn3a-positive labels was used to differentiate RGCs from coupled and uncoupled d(displaced)ACs in the GCL. RGCs in the GCL of WT retinas accounted for 52% of all cells, with the remainder being dACs; 52% of dACs were coupled to RGCs. Eight weeks after microbead injection the DAPI-positive cells were reduced by ~30%. Of the remaining cells, 50% were dACs of which 27% were coupled to RGCs. This reduction in coupled dACs was accompanied by a 25% increase of uncoupled dACs. This suggests that the loss of dACs is not due totally to the loss of RGCs to which they are coupled, but that ~13-18% of uncoupled dACs are lost as well. Interesting, 50% of all cells in the GCL of Cx45 KO mice are dACs, similar to WT, but only 20% of dACs are coupled to RGCs, 62% less than in WT. However, bead-injected retinas of Cx45 KO mice showed no change from control values in the number of dACs nor in the percentage of cells coupled to RGCs. Following microbead injection, CR- positive ACs were significantly reduced in the INL (23%) and GCL (26%). ChAT-positive ACs, which are generally uncoupled, were also reduced in the INL (26%) and GCL (17%) following microbead injection. Ablation of Cx36 largely prevented the loss of CR-and ChAT-positive ACs in both the INL and GCL of bead-injected retinas.
Experimental glaucoma produces a significant loss of ACs in the INL and GCL. Cx45- and Cx36-expressing GJs couple RGCs to ACs and their ablation significantly reduces AC loss. Our results suggest that AC loss in glaucoma is largely via the bystander effect, secondary to the loss of RGCs that they are coupled to, but uncoupled ACs are lost as well.
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