G protein coupling and signaling pathway activation by M1 muscarinic acetylcholine receptor orthosteric and allosteric agonists

Rachel L Thomas, Rajendra Mistry, Christopher J Langmead, Martyn D Wood, R A John Challiss

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44 Citations (Scopus)


The M1 muscarinic acetylcholine (mACh) receptor is among a growing number of G protein-coupled receptors that are able to activate multiple signaling cascades. AC-42 (4-n-butyl-1-[4-(2-methylphenyl)-4-oxo-1-butyl] piperidine) is an allosteric agonist that can selectively activate the M 1 mACh receptor in the absence of an orthosteric ligand. Allosteric agonists have the potential to stabilize unique receptor conformations, which may in turn cause differential activation of signal transduction pathways. In the present study, we have investigated the signaling pathways activated by AC-42, its analog 77-LH-28-1 (1-[3-(4-butyl-1-piperidinyl)propyl]-3,4-dihydro- 2(1H)-quinolinone), and a range of orthosteric muscarinic agonists [oxotremorine-M (oxo-M), arecoline, and pilocarpine] in Chinese hamster ovary cells recombinantly expressing the human M1 mACh receptor. Each agonist was able to activate Gaq/11-dependent signaling, as demonstrated by an increase in guanosine 5 -O-(3-thiotriphosphate) ([ 35S]GTP?S) binding to Gaq/11 proteins and total [3H]inositol phosphate accumulation assays in intact cells. All three orthosteric agonists caused significant enhancements in [ 35S]GTP?S binding to Gai1/2 subunits over basal; however, neither allosteric ligand produced a significant response. In contrast, both orthosteric and allosteric agonists are able to couple to the Gas/cAMP pathway, enhancing forskolin-stimulated cAMP accumulation. These data provide support for the concept that allosteric and orthosteric mACh receptor agonists both stabilize receptor conformations associated with Gaq/11-and Gas-dependent signaling; however, AC-42 and 77-LH-28-1, unlike oxo-M, arecoline, and pilocarpine, do not seem to promote M1 mACh receptor-Gai1/2 coupling, suggesting that allosteric agonists have the potential to activate distinct subsets of downstream effectors.
Original languageEnglish
Pages (from-to)365 - 374
Number of pages10
JournalJournal of Pharmacology and Experimental Therapeutics
Issue number2
Publication statusPublished - 2008
Externally publishedYes

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