G protein coupling to M1 and M3 muscarinic receptors in sublingual glands.
Rat sublingual gland M2 and M3 muscarinic receptors each directly activate exocrine secretion. To investigate the functional role of coreceptor expression, we determined receptor-G protein coupling. Although membrane proteins of 40 and 41 kDa are ADP-ribosylated by pertussis toxin (PTX), and 44 kDa proteins by cholera toxin (CTX), both carbachol-stimulated high-affinity GTPase activity and the GTP-induced shift in agonist binding are insensitive to CTX or PTX. Carbachol enhances photoaffinity labeling ([alpha-(32)P]GTP-azidoaniline) of only 42-kDa proteins that are subsequently tractable to immunoprecipitation by antibodies specific for Galpha(q) or Galpha(11) but not Galpha(12) or Galpha(13). Carbachol-stimulated photoaffinity labeling as well as phosphatidylinositol 4,5-bisphosphate (PIP2) hydrolysis is reduced 55% and 60%, respectively, by M1 receptor blockade with m1-toxin. Galpha(q/11)-specific antibody blocks carbachol-stimulated PIP2 hydrolysis. We also provide estimates of the molar ratios of receptors to Galpha(q) and Galpha(11). Although simultaneous activation of M1 and M3 receptors is required for a maximal response, both receptor subtypes are coupled to Galpha(q) and Galpha(11) to stimulate exocrine secretion via redundant mechanisms.
Center for Oral Biology, University of Rochester Medical Center, 601 Elmwood Ave., Rochester, NY 14642-8611, USA.
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W
Luo
L R
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Latchney
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DJ
Culp
D000345:Affinity Labels / Q000494:pharmacology; D000818:Animals; D001386:Azides / Q000494:pharmacology; D002217:Carbachol / Q000494:pharmacology; D002772:Cholera Toxin / Q000494:pharmacology; D018679:Cholinergic Agonists / Q000494:pharmacology; D004546:Elapid Venoms / Q000494:pharmacology; D043802:GTP-Binding Protein alpha Subunits, Gq-G11; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000494:pharmacology; D006160:Guanosine Triphosphate / Q000031:analogs & derivatives* / Q000494:pharmacology; D020962:Heterotrimeric GTP-Binding Proteins / Q000378:metabolism*; D006868:Hydrolysis; D066298:In Vitro Techniques; D008297:Male; D037342:Pertussis Toxin; D019269:Phosphatidylinositol 4,5-Diphosphate / Q000378:metabolism; D010761:Phosphorus Radioisotopes; D051381:Rats; D017208:Rats, Wistar; D043643:Receptor, Muscarinic M1; D043587:Receptor, Muscarinic M3; D011976:Receptors, Muscarinic / Q000378:metabolism*; D013047:Specific Pathogen-Free Organisms; D013361:Sublingual Gland / Q000187:drug effects / Q000378:metabolism*; D010566:Virulence Factors, Bordetella / Q000494:pharmacology
D000345:Affinity Labels; D001386:Azides; D018679:Cholinergic Agonists; D004546:Elapid Venoms; D019269:Phosphatidylinositol 4,5-Diphosphate; D010761:Phosphorus Radioisotopes; D043643:Receptor, Muscarinic M1; D043587:Receptor, Muscarinic M3; D011976:Receptors, Muscarinic; D010566:Virulence Factors, Bordetella; C083667:m1-toxin; D016244:Guanosine 5'-O-(3-Thiotriphosphate); C031907:GTP gamma-4-azidoanilide; D006160:Guanosine Triphosphate; D002217:Carbachol; D002772:Cholera Toxin; D037342:Pertussis Toxin; D043802:GTP-Binding Protein alpha Subunits, Gq-G11; D020962:Heterotrimeric GTP-Binding Proteins