Regulation of the muscarinic K + channel by extracellular ATP through membrane phosphatidylinositol 4,5-bisphosphate in guinea-pig atrial myocytes

Abstract

The present study was designed to examine the functional role of membrane phosphatidylinositol 4,5-bisphosphate (PtdIns(4,S)P 2) in the regulation of the muscarinic K + channel (I K.ACh) by extracellular ATP and adenosine in guinea-pig atrial myocytes, using the whole-cell patch-clamp method. Bath application of ATP in micromolar concentrations typically evoked a transient activation of I K.ACh; a rapid activation phase was consistently followed by a progressive decline even to the baseline level despite the continued presence of ATP. This progressive decline of I K.ACh was significantly attenuated either by blockade of phospholipase C (PLC) with compound 48/80 (100 μM) or by addition of PtdIns(4,5)P 2 (50 μM) to the cell inside, suggesting that depletion of membrane PtdIns(4,5)P 2 via PLC activation is mainly, if not totally, responsible for the progressive decline of I K.ACh during the presence of ATP. When atrial myocytes were exposed to wortmannin (50 μM) following ATP (50 μM) application to impair the resynthesis of PtdIns(4,5)P 2, the activation of I K.ACh evoked by subsequently applied ATP (50 μM) was greatly reduced. Activation of I K.ACh by adenosine (100 μM) was partially reduced by pretreatment of atrial myocytes with ATP (100 μM) and was largely abolished by a further addition of wortmannin (50 μM) in the presence of ATP (100 μM). These results support the view that the activation of I K.ACh by ATP and adenosine depends on membrane PtdIns(4,5)P 2 that is subject to reduction by extracellular ATP. The present study thus provides functional evidence to suggest that extracellular ATP activates PLC and thereby depletes membrane PtdIns(4,5)P 2 that is critically involved in the activation process of I K.ACh by its agonists ATP and adenosine in guinea-pig atrial myocytes. © 2005 Nature Publishing Group. All rights reserved.