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NEUROPHARMACOLOGY

Pharmacological Assessment of M₁ Muscarinic Acetylcholine Receptor-G_q/11 Protein Coupling in Membranes Prepared from Postmortem Human Brain Tissue

Department of Cell Physiology and Pharmacology, Henry Wellcome Building, University of Leicester, Leicester, United Kingdom (H.S.-U., R.A.J.C.); and Psychiatry Centre for Excellence in Drug Discovery, GlaxoSmithKline, Harlow, Essex, United Kingdom (D.R.T., C.H.D., J.J.H., M.D.W., J.M.W.)

Using a selective G_q/11 protein antibody capture guanosine 5'-O-(3-[³⁵S]thio)triphosphate ([³⁵S]GTPS) binding approach, it has been possible to perform a quantitative pharmacological examination of the functional activity of the M₁ muscarinic acetylcholine receptor (mAChR) in membranes prepared from human postmortem cerebral cortex. Oxotremorine-M caused a 2-fold increase in [³⁵S]GTPS-G_q/11 binding with a pEC₅₀ of 6.06 ± 0.16 in Brodmann's areas 23 and 25 that was almost completely inhibited by preincubation of membranes with the M₁ 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]-GTPS-G_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 M₁ receptor is the predominant mAChR subtype coupling to the G_q/11 G protein in these brain regions and that it is possible to quantify the potency and intrinsic activity of full and partial M₁ mAChR receptor agonists in postmortem human brain using a selective G_q/11 protein antibody capture [³⁵S]GTPS binding assay.

Address correspondence to: Dr. R. A. John Challiss, Department of Cell Physiology and Pharmacology, University of Leicester, Room 4/04, Henry Wellcome Building, Lancaster Road, Leicester LE1 9HN, United Kingdom. E-mail: jc36{at}leicester.ac.uk