title: Interaction of agonists and selective antagonists with gastric smooth muscle muscarinic receptors.
abstract: The interaction of cholinergic agonists and antagonists with smooth muscle muscarinic receptors has been investigated by measurement of displacement of the muscarinic antagonist [3H]QNB (quinuclidinyl benzilate) in membranes prepared from toad stomach. The binding of [3H]QNB was saturable, reversible and of high affinity (KD = 423 pM). The muscarinic receptor subtypes present in gastric smooth muscle were classified by determining the relative affinities for the selective antagonists pirenzepine (M1), AF-DX 116 (M2) and 4-DAMP (M3). The results from these studies indicate the presence of a heterogeneous population of muscarinic receptor subtypes, with a majority (88%) exhibiting characteristics of M3 receptors and a much smaller population (12%) exhibiting characteristics of M2 receptors. The binding curve for the displacement of [3H]QNB binding by the agonist oxotremorine was complex and was consistent with presence of two affinity states: 24% of the receptors had a high affinity (KD = 4.7 nM) for oxotremorine and 76% displayed nearly a 1,000-fold lower affinity (KD = 4.4 microM). When oxotremorine displacement of [3H]QNB binding was determined in the presence GTP gamma S, high affinity binding was abolished, indicating that high affinity agonist binding may represent receptors coupled to G proteins. Moreover, pertussis toxin pretreatment of membranes also abolished high affinity agonist binding, indicating that the muscarinic receptors are coupled to pertussis toxin-sensitive G proteins. Reaction of smooth muscle membranes with pertussis toxin in the presence [32P]NAD caused the [32P]-labelling of a 40 dD protein that may represent the alpha subunit(s) of G proteins that are known to by NAD-ribosylated by the toxin.(ABSTRACT TRUNCATED AT 250 WORDS)
authors.affiliation: Department of Physiology, University of Massachusetts, Medical School, Worcester 01655.
authors.forename: P A
authors.initials: PA
authors.lastname: Lucchesi
authors.forename: F D
authors.initials: FD
authors.lastname: Romano
authors.forename: C R
authors.initials: CR
authors.lastname: Scheid
authors.forename: H
authors.initials: H
authors.lastname: Yamaguchi
authors.forename: T W
authors.initials: TW
authors.lastname: Honeyman
mesh_terms: D000218:Adamantane / Q000031:analogs & derivatives / Q000494:pharmacology; D000246:Adenosine Diphosphate Ribose / Q000378:metabolism; D000818:Animals; D002024:Bufo marinus; D005493:Folic Acid Antagonists / Q000494:pharmacology; D006160:Guanosine Triphosphate / Q000494:pharmacology; D066298:In Vitro Techniques; D009130:Muscle, Smooth / Q000187:drug effects*; D010095:Oxotremorine / Q000494:pharmacology; D010276:Parasympatholytics / Q000494:pharmacology*; D010277:Parasympathomimetics / Q000494:pharmacology*; D037342:Pertussis Toxin; D010890:Pirenzepine / Q000031:analogs & derivatives / Q000494:pharmacology; D011813:Quinuclidinyl Benzilate / Q000494:pharmacology; D011976:Receptors, Muscarinic / Q000187:drug effects*; D013270:Stomach / Q000187:drug effects; D010566:Virulence Factors, Bordetella / Q000494:pharmacology
chemical_list: D005493:Folic Acid Antagonists; D010276:Parasympatholytics; D010277:Parasympathomimetics; D011976:Receptors, Muscarinic; D010566:Virulence Factors, Bordetella; D000246:Adenosine Diphosphate Ribose; C016358:2,4-diamino-5-adamantyl-6-methylpyrimidine; D010890:Pirenzepine; D010095:Oxotremorine; D011813:Quinuclidinyl Benzilate; D006160:Guanosine Triphosphate; D037342:Pertussis Toxin; C048593:otenzepad; D000218:Adamantane