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Extracellular ATP raises cytosolic calcium and activates basolateral chloride conductance in Necturus proximal tubule

1 Extracellular nucleotides modulate ionic transport mechanisms in various epithelia. In the present study, we investigated the effects of extracellular ATP on the intracellular free Ca2+ concentration ([Ca2+]i) and electrophysiological properties of Necturus maculosus proximal convoluted tubule (PCT). 2 ATP raised [Ca2+]i in microdissected fura‐2‐loaded PCTs (half‐maximal effect, ≈15 μmol l−1 ATP). The initial ATP‐induced changes in [Ca2+]i were not blunted by the removal of external Ca2+ nor by the presence of Ca2+ channel blockers, but were abolished by thapsigargin and suramin. The sequence for the potency of various agonists on [Ca2+]i was 2‐methylthioATP (2MeSATP) = ADP = ATP ≫ UTP, 2′,3′‐O‐(4‐benzoilbenzoil) ATP (BzATP), α,β‐methylene ATP (AMPCPP), adenosine. 3 In vivo electrophysiological measurements showed that 100 μmol l−1 peritubular ATP added to a Ringer solution reduced the basolateral cell membrane potential (Vm) and increased the cell membrane input conductance. In a low Cl− solution, this ATP‐induced depolarization was enhanced. These effects were inhibited by 1 mmol l−1SITS, consistent with the activation of a basolateral Cl− conductance. 4 The ATP‐induced change in Vm was reproduced by ADP but not by UTP or adenosine, and was prevented by suramin. 5 The ATP‐induced membrane depolarization was not influenced by thapsigargin, BAPTA AM, or staurosporin and was not reproduced by manoeuvres increasing [Ca2+]i or intracellular cAMP content. 6 We conclude that, in Necturus PCT, a P2y receptor mobilizes Ca2+ mainly from intracellular pools and increases a basolateral Cl− conductance, GCl. The activation of GCl occurs by a mechanism which is not related either to an increase in [Ca2+]i or cAMP content, or to PKC activation.