title: Novel inhibition of gbetagamma-activated potassium currents induced by M(2) muscarinic receptors via a pertussis toxin-insensitive pathway.
abstract: G(i) protein-coupled receptors such as the M(2) muscarinic acetylcholine receptor (mAChR) and A(1) adenosine receptor have been shown to activate G protein-activated inwardly rectifying K(+) channels (GIRKs) via pertussis toxin-sensitive G proteins in atrial myocytes and in many neuronal cells. Here we show that muscarinic M(2) receptors not only activate but also reversibly inhibit these K(+) currents when stimulated with agonist for up to 2 min. The M(2) mAChR-mediated inhibition of the channel was also observed when the channels were first activated by inclusion of guanosine 5'-O-(thiotriphosphate) in the pipette. Under these conditions the M(2) mAChR-induced inhibition was quasi-irreversible, suggesting a role for G proteins in the inhibitory process. In contrast, when GIRK currents were maximally activated by co-expressing exogenous Gbetagamma, the extent of acetylcholine (ACh)-induced inhibition was significantly reduced, suggesting competition between the receptor-mediated inhibition and the large pool of available Gbetagamma subunits. The signaling pathway that led to the ACh-induced inhibition of GIRK channels was unaffected by pertussis toxin pretreatment. Furthermore, the internalization and agonist-induced phosphorylation of M(2) mAChR was not required because a phosphorylation- and internalization-deficient mutant of the M(2) mAChR was as potent as the wild-type counterpart. Pharmacological agents modulating various protein kinases or phosphatidylinositol 3-kinase did not affect the inhibition of GIRK currents. Furthermore, the signaling pathway that mediates GIRK current inhibition was found to be membrane-delimited because bath application of ACh did not inhibit GIRK channel activity in cell-attached patches. Other G protein-coupled receptors including M(4) mAChR and alpha(1A) adrenergic receptors also caused the inhibition, whereas other G protein-coupled receptors including A(1) and A(3) adenosine receptors and alpha(2A) and alpha(2C) adrenergic receptors could not induce the inhibition. The presented results suggest the existence of a novel signaling pathway that can be activated selectively by M(2) and M(4) mAChR but not by adenosine receptors and that involves non-pertussis toxin-sensitive G proteins leading to an inhibition of Gbetagamma-activated GIRK currents in a membrane-delimited fashion.
authors.affiliation: Department of Molecular Pharmacology and Biological Chemistry, Northwestern University Medical School, Chicago, Illinois 60611, USA.
authors.forename: M
authors.initials: M
authors.lastname: Bünemann
authors.forename: T
authors.initials: T
authors.lastname: Meyer
authors.forename: L
authors.initials: L
authors.lastname: Pott
authors.forename: M
authors.initials: M
authors.lastname: Hosey
mesh_terms: D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D051676:G Protein-Coupled Inwardly-Rectifying Potassium Channels; D019204:GTP-Binding Proteins / Q000378:metabolism; D006801:Humans; D018408:Patch-Clamp Techniques; D037342:Pertussis Toxin; D011188:Potassium / Q000378:metabolism*; D015221:Potassium Channels / Q000378:metabolism; D024661:Potassium Channels, Inwardly Rectifying; D011956:Receptors, Cell Surface / Q000037:antagonists & inhibitors*; D011950:Receptors, Cholinergic / Q000378:metabolism; D011976:Receptors, Muscarinic / Q000378:metabolism*; D018047:Receptors, Purinergic P1 / Q000378:metabolism; D014162:Transfection; D010566:Virulence Factors, Bordetella / Q000378:metabolism*
chemical_list: D051676:G Protein-Coupled Inwardly-Rectifying Potassium Channels; D015221:Potassium Channels; D024661:Potassium Channels, Inwardly Rectifying; D011956:Receptors, Cell Surface; D011950:Receptors, Cholinergic; D011976:Receptors, Muscarinic; D018047:Receptors, Purinergic P1; D010566:Virulence Factors, Bordetella; D037342:Pertussis Toxin; D019204:GTP-Binding Proteins; D011188:Potassium