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

1 The present study was designed to examine the functional role of membrane phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5) 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. 2 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 mu M) or by addition of PtdIns( 4,5) P-2 (50 mM) 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. 3 When atrial myocytes were exposed to wortmannin (50 mM) following ATP (50 mu M) application to impair the resynthesis of PtdIns(4,5) P-2, the activation of I-K,(ACh) evoked by subsequently applied ATP (50 mM) was greatly reduced. Activation of I-K,(ACh) by adenosine (100 mM) was partially reduced by pretreatment of atrial myocytes with ATP (100 mu M) and was largely abolished by a further addition of wortmannin (50 mu M) in the presence of ATP (100 mu M). 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. 4 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.