Depletion of Phosphatidylinositol 4,5-Bisphosphate by Activation of Phospholipase C-coupled Receptors Causes Slow Inhibition but Not Desensitization of G Protein-gated Inward Rectifier K+ Current in Atrial Myocytes

Abstract

G protein-gated inwardly rectifier K+ current in atrial myocytes (I(K(ACh))) upon stimulation with acetylcholine (ACh) shows a fast desensitizing component (t(1/2) approximately 5 s). After washout of ACh, I(K(ACh)) recovers from fast desensitization within < 30 s. A recent hypothesis suggests that fast desensitization is caused by depletion of phosphatidylinositol 4,5-bisphosphate (PtIns(4,5)P(2)), resulting from costimulation of phospholipase C (PLC)-coupled M3 receptors (M3AChR). The effects of stimulating two established PLC-coupled receptors, alpha-adrenergic and endothelin (ET(A)), on I(K(ACh)) were studied in rat atrial myocytes. Stimulation of these receptors caused activation of I(K(ACh)) and inhibition of the M2AChR-activated current. In myocytes loaded with GTPgammaS (guanosine 5'-3-O-(thio)triphosphate), causing stable activation of I(K(ACh)), inhibition via alpha-agonists and ET-1 was studied in isolation. Stimulation of either type of receptor under this condition, via G(q/11), caused a slow inhibition (t(1/2) approximately 50 s) by about 70%. No comparable effect on GTPgammaS-activated I(K(ACh)) was induced by ACh, suggesting that PLC-coupled M3AChRs are not functionally expressed in rat myocytes, which was supported by the finding that M3AChR transcripts were not detected by reverse transcriptase-polymerase chain reaction in identified atrial myocytes. Supplementing the pipette solution with PtIns(4,5)P(2) significantly reduced inhibition of I(K(ACh)) but had no effect on fast desensitization. From these data it is concluded that stimulation of PLC-coupled receptors causes slow inhibition of I(K(ACh)) by depletion of PtIns(4,5)P(2), whereas fast desensitization of I(K(ACh)) is not related to PtIns(4,5)P(2) depletion. As muscarinic stimulation by ACh does not exert inhibition of I(K(ACh)) comparable to stimulation of alpha(1)- and ET(A) receptors, expression of functional PLC-coupled muscarinic receptors in rat atrial myocytes is unlikely.