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J. Xia, J. C. Lim, W. Lu, A. M. Laties, C. H. Mitchell; Mechanosensitive Release of ATP via Pannexins Autostimulates P2X7 Receptors on Retinal Ganglion Cells. Invest. Ophthalmol. Vis. Sci. 2010;51(13):3317.
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The cellular stretch induced by intraocular pressure (IOP) elevation may initiate signals that affect retinal ganglion cells (RGCs). In other tissues, pressure and stretch lead to a release of ATP, with the subsequent autostimulation of purine receptors. We have previously demonstrated that swelling RGCs activates a cation current that is inhibited by removal of extracellular ATP. As these RGCs were surrounded by other retinal cell types, the cellular source of this ATP was unclear. The present study asks whether isolated RGCs can autostimulate receptors, examines the mechanism for ATP release and the receptor subtype.
Ionic currents from RGCs of rat neonatal retinas were measured using standard patch clamp protocols. RGCs were isolated with the two-step immunopanning technique. In other experiments, RGCs were identified in mixed culture after injection of fluorescent label into the superior colliculus. Mild membrane swelling was induced by the removal of 30 - 55 mM mannitol from the extracellular solution.
Swelling of isolated RGCs activated a cation current. The P2X7 receptor antagonist A438079 (1 µM) inhibited this current by 74±11% (n=12). This suggested RGCs can themselves release ATP that autostimulates their P2X7 receptors. The conduit of this release was probed by swelling labeled RGCs. These currents were similar to those observed in isolated cells, with A438079 inhibiting the current by 75±10 % (n=9). Currents were also reduced by pannexin hemichannel blockers. Carbenoxolone (10 µM) blocked the current by 57±9% (n=9) while probenecid (1 mM) blocked the current by 60±7% (n=10).
Swelling RGCs induced a current dependent upon extracellular ATP, P2X7 receptors and pannexin channels. The current was activated in the absence of other retinal cell types, implicating ganglion cells themselves are a source of released ATP. These data are consistent with a mechanosensitive release of ATP through pannexin channels which autostimulates P2X7 receptors on RGCs. Whether an analogous autostimulation occurs in response to elevated IOP remains to be determined.
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