Glutamate‐mediated CA++ influx and nuclear damage in retinal ganglion cells purified by panning: role of glia

Abstract

The visual acuity decline in glaucoma is due to degeneration of retinal ganglion cells. We used cell panning with the antibody Thy 1.1 to isolate retinal ganglion cells (RGCs) from rat retina. Individual RGCs were extremely sensitive to glutamate, demonstrating sustained calcium influx responses to 100 nm glutamate. We tested unoprostone for its ability to elicit neuroprotection. This docosanoid decreases intraocular pressure in glaucoma patients and protects mixed retinal ganglion cell cultures from glutamate excitotoxicity and calcium influx. In the presence of retinal glia, RGCs responded to 100 and 500 nm glutamate with transient calcium influx, demonstrating glial buffering effects. Pure retinal glial cultures required on average at least 1 mm glutamate to trigger calcium influx, which was transient and oscillating. Unoprostone (10 μm) inhibited calcium influx stimulated by 5 mm glutamate in retinal glial cultures, while DMSO did not. Unoprostone at 1 μm also blocked neuronal calcium responses to 100 nm glutamate in mixed cultures, while DMSO did not. In RGCs isolated by panning, unoprostone blocked calcium responses to 100 nm glutamate in some cells, while DMSO blocked none. Unoprostone also protected RGCs from 20 μm glutamate‐induced toxicity and nuclear damage. We hypothesize that while unoprostone may affect individual RGCs depending on a putative receptor, the presence of retinal glia also plays a key role in neuroprotection and unoprostone action. The use of the cellular model described here and whole retina preparations will be useful to understand the mechanisms of RGC survival.Acknowledgements: Support: Novartis Ophthalmics, Basel, Switzerland.