Distinct Agonist Regulation of Muscarinic Acetylcholine M(2)-M(3) Heteromers and Their Corresponding Homomers

Each subtype of the muscarinic receptor family of G protein-coupled receptors is activated by similar concentrations of the neurotransmitter acetylcholine or closely related synthetic analogs such as carbachol. However, pharmacological selectivity can be generated by the introduction of a pair of mutations to produce Receptor Activated Solely by Synthetic Ligand (RASSL) forms of muscarinic receptors. These display loss of potency for acetylcholine/carbachol alongside a concurrent gain in potency for the ligand clozapine N-oxide. Co-expression of a form of wild type human M(2) and a RASSL variant of the human M(3) receptor resulted in concurrent detection of each of M(2)-M(2) and M(3)-M(3) homomers alongside M(2)-M(3) heteromers at the surface of stably transfected Flp-In(TM) T-REx(TM) 293 cells. In this setting occupancy of the receptors with a muscarinic antagonist was without detectable effect on any of the muscarinic oligomers. However, selective agonist occupancy of the M(2) receptor resulted in enhanced M(2)-M(2) homomer interactions but decreased M(2)-M(3) heteromer interactions. By contrast, selective activation of the M(3) RASSL receptor did not significantly alter either M(3)-M(3) homomer or M(2)-M(3) heteromer interactions. Selectively targeting closely related receptor oligomers may provide novel therapeutic opportunities.