Ca2+ release and activation of K+ and Cl- currents by extracellular ATP in distal nephron epithelial cells

We have measured ionic currents and changes in intracellular Ca2+ concentration ([Ca2+]i) induced by extracellular ATP in single epithelial cells of the distal nephron from toad (A6 cells). ATP increased [Ca2+]i and concomitantly activated ionic currents. The ATP concentration for half-maximal increase in [Ca2+]i was approximately 10 microM. Current activation and elevation of [Ca2+]i also occurred in Ca(2+)-free bath solutions but were abolished by loading the cells via the patch pipette with 10 mM 1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetic acid (BAPTA) or by preincubating the cells with 10 microM BAPTA-acetoxymethyl ester for 120 min. ATP-activated currents reversed at -53.9 +/- 1.9 mV (n = 22). Tetraethylammonium (TEA, 25 mM), a K+ channel blocker, partially blocked this current but did not affect the Ca2+ transients. The TEA-insensitive component of the current reversed close to Cl- equilibrium potential. 5-Nitro-2-(3-phenylpropylamino) benzoic acid, a putative Cl- channel blocker (100 microM), abolished nearly completely the ATP-activated current. Suramin (100 microM), a P2-purinergic receptor antagonist, strongly attenuated both Ca2+ transients and currents. In cell-attached patches, single channel currents activated by ATP could be observed, i.e., an inwardly rectifying K+ channel with a slope conductance for inward currents of approximately 32 pS and an ohmic Cl- channel with a conductance of 34 pS. It is concluded that ATP activates both Cl- and K+ channels in distal nephron epithelial cells by a Ca(2+)-dependent mechanism.