title: Specific Gq protein involvement in muscarinic M3 receptor-induced phosphatidylinositol hydrolysis and Ca2+ release in mouse duodenal myocytes.
abstract: 1. Cytosolic Ca2+ concentration ([Ca2+]i) during exposure to acetylcholine or caffeine was measured in mouse duodenal myocytes loaded with fura-2. Acetylcholine evoked a transient increase in [Ca2+]i followed by a sustained rise which was rapidly terminated after drug removal. Although L-type Ca2+ currents participated in the global Ca2+ response induced by acetylcholine, the initial peak in [Ca2+]i was mainly due to release of Ca2+ from intracellular stores. 2. Atropine, 4-diphenylacetoxy-N-methylpiperidine (4-DAMP, a muscarinic M3 antagonist), pirenzepine (a muscarinic M1 antagonist), methoctramine and gallamine (muscarinic M2 antagonists) inhibited the acetylcholine-induced Ca2+ release, with a high affinity for 4-DAMP and atropine and a low affinity for the other antagonists. Selective protection of muscarinic M2 receptors with methoctramine during 4-DAMP mustard alkylation of muscarinic M3 receptors provided no evidence for muscarinic M2 receptor-activated [Ca2+]i increase. 3. Acetylcholine-induced Ca2+ release was blocked by intracellular dialysis with a patch pipette containing either heparin or an anti-phosphatidylinositol antibody and by external application of U73122 (a phospholipase C inhibitor). 4. Acetylcholine-induced Ca2+ release was insensitive to external pretreatment with pertussis toxin, but concentration-dependently inhibited by intracellular dialysis with a patch pipette solution containing an anti-alpha q/alpha 11 antibody. An antisense oligonucleotide approach revealed that only the Gq protein was involved in acetylcholine-induced Ca2+ release. 5. Intracellular applications of either an anti-beta com antibody or a peptide corresponding to the G beta gamma binding domain of the beta-adrenoceptor kinase 1 had no effect on acetylcholine-induced Ca2+ release. 6. Our results show that, in mouse duodenal myocytes, acetylcholine-induced release of Ca2+ from intracellular stores is mediated through activation of muscarinic M3 receptors which couple with a Gq protein to activate a phosphatidylinositol-specific phospholipase C.
authors.affiliation: Laboratoire de Physiologie Cellulaire et Pharmacologie Moléculaire, CNRS ESA 5017, Université de Bordeaux II, France.
authors.forename: J L
authors.initials: JL
authors.lastname: Morel
authors.forename: N
authors.initials: N
authors.lastname: Macrez
authors.forename: J
authors.initials: J
authors.lastname: Mironneau
mesh_terms: D000109:Acetylcholine / Q000494:pharmacology; D000818:Animals; D002110:Caffeine / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000502:physiology; D004386:Duodenum / Q000378:metabolism*; D019204:GTP-Binding Proteins / Q000502:physiology*; D006493:Heparin / Q000494:pharmacology; D006868:Hydrolysis; D051379:Mice; D009130:Muscle, Smooth / Q000378:metabolism*; D037342:Pertussis Toxin; D010716:Phosphatidylinositols / Q000378:metabolism*; D043587:Receptor, Muscarinic M3; D011976:Receptors, Muscarinic / Q000502:physiology*; D010738:Type C Phospholipases / Q000502:physiology; D010566:Virulence Factors, Bordetella / Q000494:pharmacology; D051552:beta-Adrenergic Receptor Kinases
chemical_list: D010716:Phosphatidylinositols; D043587:Receptor, Muscarinic M3; D011976:Receptors, Muscarinic; D010566:Virulence Factors, Bordetella; D002110:Caffeine; D006493:Heparin; D037342:Pertussis Toxin; D017868:Cyclic AMP-Dependent Protein Kinases; D051552:beta-Adrenergic Receptor Kinases; D010738:Type C Phospholipases; D019204:GTP-Binding Proteins; D000109:Acetylcholine; D002118:Calcium