An Ex Vivo Explant Model for Studying Glial Interactions in the Mouse Retina

The role of glia in glaucoma is an increasingly prominent research topic, but much remains unknown about how populations of these support cells - namely astrocytes and microglia - influence retinal ganglion cell survival. While in vivo and in vitro models provide a degree of insight, both approaches have significant limitations, such as the impact of peripheral immune response in the former and changes to physiological function induced by cell isolation and culture in the case of the latter. To minimize these confounding factors, we have developed an ex vivo retinal explant system in which astrocytes, microglia, and other retinal cell types can be maintained in situ for periods of at least 3 days, enabling targeted investigation at a higher throughput than is typically feasible with in vivo models. Crucially, this approach is highly amenable to methodologies that would be challenging or unfeasible in a living animal yet remains compatible with common downstream assays of intact nervous tissue. Here, we present a protocol suitable for the isolation and culturing of intact retinal explants, along with representative results of immunofluorescence microscopy documenting the changes undergone by glia and retinal ganglion cells in the ex vivo retina.