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

摘要

Background: Muscarinic receptors can form both homo- and hetero-oligomers. Results: Co-expression of M_(2)and M_(3)receptors resulted in concurrent detection of both homomer and heteromer interactions and regulation of M_(2)-containing forms by agonist. Conclusion: Co-existing receptor oligomers display differential regulation. Significance: Oligomers of closely related receptors display distinct properties that may be targeted therapeutically. 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.