The microglia response to electrical overstimulation of the retina imaged under a transparent stimulus electrode

Abstract

Objective. We investigated using the morphological response of retinal microglia as indicators of tissue damage from electrical overstimulation by imaging them through an optically transparent stimulus electrode. Approach. To track the microglia, we used a transgenic mouse where the microglia expressed a water soluble green fluorescent protein. The clear stimulus electrode was placed epiretinally on the inner limiting membrane and the microglia layers were imaged using time-lapse confocal microscopy. We examined how the microglia responded both temporally and spatially to local overstimulation of the retinal tissue. Using confocal microscope vertical image stacks, the microglia under the electrode were imaged at 2.5 min intervals. The retina was overstimulated for a 5 min period using 1 ms 749 μC cm−2 ph−1 biphasic current pulses and changes in the microglia morphology were followed for 1 h post stimulation. After the imaging period, a label for cellular damage was applied to the retina. Main results. The microglia response to overstimulation depended on their spatial location relative to the electrode lumen and could result in three different morphological responses. Some microglia were severely injured and became a series of immotile ball-like fluorescent processes. Other microglia survived, and reacted rapidly to the injury by extending filopodia oriented toward the damage zone. This response was seen in inner retinal microglia outside the stimulus electrode edge. A third effect, seen with the deeper outer microglia under the electrode, was a fading of their fluorescent image which appeared to be due to optical scatter caused by overstimulation-induced retinal edema. Significance. The microglial morphological responses to electrical overstimulation injury occur rapidly and can show both direct and indirect effects of the stimulus electrode injury. The microglia injury pattern closely follows models of the electric field distribution under thinly insulated disc electrodes.

Effect of epiretinal electrical stimulation on the glial cells in a rabbit retinal eyecup model

“…Real-time imaging studies of transgenic mice microglia labeled with green fluorescent protein (GFP) showed focused laser injury in CNS and retinal tissue caused their processes to become oriented centripetally and extended into small local lesions (Davalos et al, 2005;Nimmerjahn et al, 2005;Eter et al, 2008;Lee et al, 2008). Although we were unable to label microglia stably using lectins in the rabbit retina a priori for reliable real-time tracking post stimulation, the local orienting movements of these microglia cell processes around the stimulated zone in the rabbit retina is very similar to our previous results using real-time imaging in the CX3CR1 GFP transgenic mouse retina using a smaller transparent stimulation tube electrode and the same high charge density stimulation (Yohannes et al, 2021). Here we also found, imaging up to 90 min.…”

“…In primates, microglial cells are also distributed throughout the central retina (except in the foveola) but at lower densities than Müller cells, like their distribution in the rabbit ( Ashwell, 1989 ; Singaravelu et al, 2017 ). This cellular distribution presumably would result in local injury responses to overstimulation similar to those described here and in Yohannes et al (2021) . In contrast, the fine processes of inner retinal astrocytes are largely absent from the avascular rabbit retinal model, but are present at low densities in the nerve fiber and ganglion cell layer of the vascular retinas of primates and humans ( Schnitzer and Karschin, 1986 ; Ramirez et al, 1994 ).…”