Stimulation of P2X7 receptors elevates Ca2+ and kills retinal ganglion cells

PURPOSE. Retinal ganglion cells are known to express ionotropic P2X(7) receptors for ATP. Stimulation of these receptors in other cells can elevate Ca2+ and sometimes lead to cell death. This study asked whether P2X(7) receptor stimulation alters the Ca2+ levels and viability of retinal ganglion cells. METHODS. P2X(7) agonists were applied to retinal ganglion cells from neonatal rats loaded with fura-2 to examine their effect on intracellular Ca2+ levels. The effect of P2X(7) receptor stimulation on cell viability was examined in rat retinal ganglion cells back-labeled with aminostilbamidine. RESULTS. The P2X(7) agonist benzoylbenzoyl adenosine triphosphate (BzATP) led to a large, sustained increase in Ca2+. BzATP was >100-fold more effective than ATP at raising intracellular Ca2+, when both agonists were applied at 10 mu M. The response to BzATP was enhanced threefold by removal of extracellular Mg2+, was dependent on extracellular Ca2+, and was prevented by brilliant blue G (BBG). BzATP led to a concentration-dependent reduction in the number of cells with a median lethal dose (LD50) of 35 mu M. Cell death was prevented by the P2X(7) antagonists BBG and oxidized ATP, but not by 30 mu M suramin, consistent with the actions of the P2X(7) receptor. BzATP activated caspases in ganglion cells, but did not lead to membrane blebbing or increased permeability to Yo-Pro-1. The L-type Ca2+ channel blocker nifedipine attenuated cell death, suggesting excessive Ca2+ influx contributes to the lethal effects of BzATP. CONCLUSION. Short-term stimulation of the P2X(7) receptor can raise Ca2+ in rat retinal ganglion cells, whereas sustained stimulation of the receptor can kill them.