title: The interaction of 4-DAMP mustard with subtypes of the muscarinic receptor.
abstract: The compound 4-DAMP mustard (N-2-chloroethyl-4-piperidinyl diphenylacetate) is a 2-chloroethylamine derivative of the selective muscarinic antagonist 4-DAMP (N,N-dimethyl-4-piperidinyl diphenylacetate). At neutral pH, 4-DAMP mustard cyclizes spontaneously into an oziridinium ion that binds covalently with muscarinic receptors. Analysis of the kinetics of receptor alkylation showed that the interaction of 4-DAMP mustard with M2 and M3 receptors was consistent with a model in which the aziridinium ion rapidly forms a reversible complex with the receptor which converts to a covalent complex at a relatively slower rate. The rate constant (k2) for alkylation of M2 and M3 receptors was approximately the same (k2 = 0.1 min-1); however, the affinity of the aziridinium ion for the M3 receptor (KD = 7.2 nM) was approximately 6.3-fold greater than that for the M2 receptor (KD = 43 nM). The results of competitive binding experiments on Chinese hamster ovary cells transfected with the M1 - M5 subtypes of the muscarinic receptor showed that the affinity of the aziridinium ion for the M1, M3, M4 and M5 subtypes was approximately the same and about 11-fold greater than that for the M2 receptor. 4-DAMP mustard is a useful tool for selectively inactivating all non-M2 muscarinic receptors, particularly when it is used in the presence of a reversible M2 selective antagonist to protect the M2 receptor from alkylation. The results of studies on isolated smooth muscle preparations that have had their M3 receptors alkylated with 4-DAMP mustard are consistent with the postulate that the M2 receptor can elicit contraction by inhibiting the relaxant effect of isoproterenol and forskolin on histamine induced contractions.
authors.affiliation: Department of Pharmacology, College of Medicine, University of California, Irvine 92717, USA.
authors.forename: F J
authors.initials: FJ
authors.lastname: Ehlert
mesh_terms: D000478:Alkylation; D000818:Animals; D004158:Diphenylacetic Acids / Q000494:pharmacology*; D007700:Kinetics; D018727:Muscarinic Antagonists / Q000494:pharmacology*; D009119:Muscle Contraction / Q000187:drug effects; D009130:Muscle, Smooth / Q000378:metabolism / Q000502:physiology; D010880:Piperidines / Q000494:pharmacology*; D011976:Receptors, Muscarinic / Q000145:classification / Q000378:metabolism
chemical_list: D004158:Diphenylacetic Acids; D018727:Muscarinic Antagonists; D010880:Piperidines; D011976:Receptors, Muscarinic; C074183:N-(2-chloroethyl)-4-piperidinyl diphenylacetate