Quantitative and topographical microglial cell changes in the ganglion cell layer and inner plexiform layer after optic nerve axotomy

Abstract

Purpose : To analyze the microglial response in the rat retina after optic nerve axotomy.

Methods : Adult Sprague Dawley rat retinal ganglion cells (RGCs) were retrogradely labelled with fluorogold applied to both superior colliculi and one week later the left optic nerve was intraorbitally transected (ONT). Rats were sacrificed at 2, 5, 9 or 14 days or 2 months after ONT (n=8/group). Retinas were dissected as wholemounts and Brn3a and Iba-1 immunodetected to identify RGCs and microglial cells (MC), respectively. Retinas were photographed for each signal. Brn3a⁺RGCs were automatically quantified and their distribution visualized by isodensity maps. The position of each MC (Iba 1⁺) and phagocytic MC (PMC, identified by their transcellular labelling with fluorogold and Iba-1 staining) found in the ganglion cell layer (GCL) and inner plexiform layer (IPL) was dotted on the retinal photomontages. These dots were counted and graphically represented with neighbour maps.

Results : In intact retinas, the mean number of microglial cells (MCs) in the GCL is 14,151±1,186 and of 11,006±989 in the IPL. In both layers, MC distribute homogenously across the retina. Following ONT the population of RGCs decreases linearly, while the number of MCs in the GCL increases linearly. MCs numbers in this layer peak at 14 days (28,328±2018), and most of them are phagocytic (22,092±3514). Afterwards their numbers decrease and at 2 months have reached control values. Thus, at 15 days after ONT there are more MCs in the GCL than in intact retinas. However the number of MCs in the IPL at the same time point is reduced by 53% suggesting that MCs migrate from the IPL to the GCL during the exponential death of RGCs. Topographically, during the first 15 days after ONT, MCs in the GCL relocate to the retinal areas of higher RGC density and, therefore, of higher RGC death. Two months after ONT, they are homogenously distributed, following the topography found in intact retinas.

Conclusions : Microglial cells are distributed homogenously in the ganglion cell layer and inner plexiform layer in intact retinas. The increase of PMC in the GCL and the decrease of MC in the IPL suggest that upon RGC injury, MC migrate between both layers.

This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.