title: Inverse agonist activity of pirenzepine at M2 muscarinic acetylcholine receptors.
abstract: 1. The intrinsic properties of muscarinic ligands were studied through their binding properties and their abilities to modulate the GTPase activity of G proteins coupled to muscarinic M2 receptors in pig atrial sarcolemma. 2. Competition binding experiments were performed with [3H]-oxotremorine-M to assess the affinity of receptors coupled to G proteins (R*), with [3H]-N-methylscopolamine ([3H]-NMS) to estimate the affinities of coupled and uncoupled receptors (R*+R) and with [3H]-NMS in the presence of GppNHp to assess the affinity of uncoupled receptors (R). 3. The ranking of Ki values for the agonist carbachol was R*<<R*+R<<R (0.95, 124 and 1017 nM). Ki values for atropine and AF-DX 116 were similar for the three binding conditions (0.34, 0.42, 0.41 and 19, 22, 32 nM). The ranking of Ki values for pirenzepine was R*>R*+R>R (174, 155, 115 nM), suggesting inverse agonism. 4. The Vmax of the basal high affinity GTPase activity of pig atrial sarcolemma was increased by mastoparan and decreased by GPAnt-2 indicating the relevance of this activity to G proteins coupled to receptors (R*). The K(M) value (0.26-0.33 microM) was not modified by mastoparan or GPAnt-2. 5. Carbachol increased the Vmax of GTP hydrolysis (EC50 8.1+/-0.3 microM), whereas atropine and AF-DX 116, up to 1 mM, did not modify it. Pirenzepine decreased the Vmax of GTP hydrolysis (EC50 77.5+/-10.3 microM). This effect was enhanced when KCI was substituted for NaCl (EC50 11.0+/-0.8 microM) and was antagonized by atropine and AF-DX 116 (IC50 0.91+/-0.71 and 197+/-85 nM). 6. Pirenzepine is proposed as an inverse agonist and atropine and AF-DX 116 as neutral antagonists at the muscarinic M2 receptor.
authors: Laboratoire de Neuroimmunopharmacologie, INSERM U 425, Faculté de Pharmacie, Université Strasbourg I, Illkirch, France.
authors: L
authors: L
authors: Daeffler
authors: F
authors: F
authors: Schmidlin
authors: J P
authors: JP
authors: Gies
authors: Y
authors: Y
authors: Landry
mesh_terms: D000818:Animals; D020558:GTP Phosphohydrolases / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000378:metabolism; D036341:Intercellular Signaling Peptides and Proteins; D018727:Muscarinic Antagonists / Q000494:pharmacology*; D010455:Peptides; D010890:Pirenzepine / Q000494:pharmacology*; D043585:Receptor, Muscarinic M2; D011976:Receptors, Muscarinic / Q000187:drug effects / Q000378:metabolism*; D012508:Sarcolemma / Q000187:drug effects* / Q000201:enzymology / Q000378:metabolism; D013552:Swine; D014862:Wasp Venoms / Q000494:pharmacology
chemical_list: D036341:Intercellular Signaling Peptides and Proteins; D018727:Muscarinic Antagonists; D010455:Peptides; D043585:Receptor, Muscarinic M2; D011976:Receptors, Muscarinic; D014862:Wasp Venoms; D010890:Pirenzepine; C037500:mastoparan; D020558:GTP Phosphohydrolases; D019204:GTP-Binding Proteins