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

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Abstract
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, similar to 15 mu mol 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-methylthio-ATP (2MeSATP) = ADP = ATP >> UTP, 2',3'-O-(4-benzoilbenzoil) ATP (BzATP), alpha,beta-methylene ATP (AMPCPP), adenosine. 3. In vivo electrophysiological measurements, showed that 100 mu mol l(-1) peritubular ATP added to a Ringer solution reduced the basolateral cell membrane potential (V-m) 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(-1) SITS, consistent with the activation of a basolateral Cl- conductance. 4. The ATP-induced change in V-m 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, G(Cl). The activation of G(Cl) occurs by a mechanism which is not related either to an increase in [Ca2+](i) or cAMP content, or to PKC activation.

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