Selective excitotoxic degeneration of adult pig retinal ganglion cells in vitro

PURPOSE. Excitotoxicity is proposed to play a prominent role in retinal gan glion cell (RGC) death ensuing from diseases such as glaucoma and ischemia, but cell culture studies have used tissue from newborn rodents, yielding c onflicting data that implicate either N-methyl D-aspartate (NMDA) or non-NM DA glutamate (Glu) receptor-mediated pathways. Excitotoxic RGC death was ex amined in vitro in this study, using adult pigs. a large-animal model for h uman retina. METHODS. Adult pig retina (and for comparative purposes young and adult rat retina) were dissociated and maintained in monolayer culture. Medium was s upplemented with Glu or pharmacologic agonists or antagonists, and survivin g RGCs and other retinal neurons were quantified using specific immunolabel ing methods. Electrophysiological responses to externally applied Glu of RG Cs in culture were recorded using whole-cell patch-clamp techniques. RESULTS. Application of Glu led to selective, dose-dependent losses in larg e RGCs (maximal 37% decrease at 1 mM: median effective dose [ED50], similar to 80 muM) and neurite damage in surviving RGCs. Application of Glu agonis ts and Glu receptor subclass antagonists showed that large RGC death was me diated through both NMDA and non-NMDA receptor pathways. Small RGCs, amacri ne cells, and all other retinal neurons were resistant to Glu-induced death . By comparison, rat retinal cultures displayed heightened RGC vulnerabilit y to Glu, mediated exclusively by non-NMDA receptor-mediated pathways. Amac rine cells were unaffected by NMDA but were very sensitive to kainate appli cation (<90% loss). Other retinal neurons were un:unaffected by any treatme nt. CONCLUSIONS. The molecular pathways underlying excitotoxic RGC death in vit ro (non-NMDA or NMDA-preferring Glu receptors) vary among species and devel opmental stages. The selective e elimination of adult pig large RGCs by NMD A receptor-mediated pathways more closely resembles human and animal glauco ma in vivo than other published culture models, providing a simplified expe rimental system for investigating the pharmacologic and toxicologic bases o f glaucoma-like neuronal death.