title: Pharmacological assessment of m1 muscarinic acetylcholine receptor-gq/11 protein coupling in membranes prepared from postmortem human brain tissue.
abstract: Using a selective Galpha(q/11) protein antibody capture guanosine 5'-O-(3-[35S]thio)triphosphate ([35S]GTPgammaS) binding approach, it has been possible to perform a quantitative pharmacological examination of the functional activity of the M(1) muscarinic acetylcholine receptor (mAChR) in membranes prepared from human postmortem cerebral cortex. Oxotremorine-M caused a > or = 2-fold increase in [35S]GTPgammaS-Galpha(q/11) binding with a pEC(50) of 6.06 +/- 0.16 in Brodmann's areas 23 and 25 that was almost completely inhibited by preincubation of membranes with the M(1) mAChR subtype-selective antagonist muscarinic toxin-7. In addition, the orthosteric and allosteric agonists, xanomeline [3(3-hexyloxy-1,2,5-thiadiazol-4-yl)-1,2,5,6-tetrahydro-1-methylpyridine] and AC-42 (4-n-butyl-1-[4-(2-methylphenyl)-4-oxo-1-butyl]-piperidine hydrogen chloride), increased [35S]-GTPgammaS-Galpha(q/11) binding, but with reduced intrinsic activities, inducing maximal responses that were 42 +/- 1 and 44 +/- 2% of the oxotremorine-M-induced response, respectively. These data indicate that the M(1) receptor is the predominant mAChR subtype coupling to the Galpha(q/11) G protein in these brain regions and that it is possible to quantify the potency and intrinsic activity of full and partial M(1) mAChR receptor agonists in postmortem human brain using a selective Galpha(q/11) protein antibody capture [35S]GTPgammaS binding assay.
authors.affiliation: Department of Cell Physiology and Pharmacology, University of Leicester, Henry Wellcome Building, Lancaster Road, Leicester LE1 9HN, United Kingdom.
authors.forename: Hasib
authors.initials: H
authors.lastname: Salah-Uddin
authors.forename: David R
authors.initials: DR
authors.lastname: Thomas
authors.forename: Ceri H
authors.initials: CH
authors.lastname: Davies
authors.forename: Jim J
authors.initials: JJ
authors.lastname: Hagan
authors.forename: Martyn D
authors.initials: MD
authors.lastname: Wood
authors.forename: Jeannette M
authors.initials: JM
authors.lastname: Watson
authors.forename: R A John
authors.initials: RA
authors.lastname: Challiss
mesh_terms: D000368:Aged; D000818:Animals; D000906:Antibodies / Q000378:metabolism; D001285:Atropine / Q000378:metabolism; D002540:Cerebral Cortex / Q000378:metabolism*; D004546:Elapid Venoms / Q000378:metabolism; D005260:Female; D043802:GTP-Binding Protein alpha Subunits, Gq-G11 / Q000378:metabolism*; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000378:metabolism; D006801:Humans; D008566:Membranes / Q000378:metabolism*; D018721:Muscarinic Agonists / Q000378:metabolism; D018727:Muscarinic Antagonists / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D043643:Receptor, Muscarinic M1 / Q000819:agonists / Q000037:antagonists & inhibitors / Q000378:metabolism*; D013462:Sulfur Radioisotopes
chemical_list: D000906:Antibodies; D004546:Elapid Venoms; D018721:Muscarinic Agonists; D018727:Muscarinic Antagonists; D043643:Receptor, Muscarinic M1; D013462:Sulfur Radioisotopes; C419876:muscarinic toxin 7; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D001285:Atropine; D043802:GTP-Binding Protein alpha Subunits, Gq-G11