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

Abstract

Using a selective Gα_q/11 protein antibody capture guanosine 5′-O-(3-[³⁵S]thio)triphosphate ([³⁵S]GTPγS) 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]GTPγS-Gα_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]-GTPγS-Gα_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 Gα_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 Gα_q/11 protein antibody capture [³⁵S]GTPγS binding assay.