Extracellular ATP activates calcium signaling, ion, and fluid transport in retinal pigment epithelium

Abstract

The presence of receptors for ATP has not been established in any native preparation of retinal neurons or glia. In the present study, we used conventional electrophysiological and [Ca2+](in) fluorescence imaging techniques to investigate the effects of ATP added to Ringer's solution perfusing the retinal-facing (apical) membrane of freshly isolated monolayers of bovine retinal pigment epithelium (RPE). ATP (or UTP) produced large, biphasic voltage and resistance changes with a K(d) of ~5 μM for ATP and ~1 μM for UTP. Electrical and pharmacological evidence indicates that the first and second phases of the response are attributable to an increase in basolateral membrane Cl conductance and a decrease in apical membrane K conductance, respectively. The ATP-induced responses were not affected by adenosine, but were reduced by the P2-purinoceptor blocker suramin. ATP also produced a large, transient increase in [Ca2+](in) that was blocked by cyclopiazonic acid, an inhibitor of endoplasmic reticulum Ca2+-ATPases. The calcium buffer BAPTA attenuated the voltage effects of ATP. We also found that apical DIDS significantly inhibited the ATP-evoked [Ca2+](in) and electrical responses, suggesting that DIDS blocked the purinoceptor. Measurements of fluid movement across the RPE using the capacitance probe technique demonstrated a significant increase in fluid absorption by apical UTP. These data indicate the presence of metabotropic P(2Y)/P(2U)- purinoceptors at the RPE apical membrane and implicate extracellular ATP in vivo as a retinal signaling molecule that could help regulate the hydration and chemical composition of the subretinal space.