An alkylating derivative of oxotremorine interacts irreversibly with the muscarinic receptor

A 2-chloroethylamine derivative of oxotremorine was studied in pharmacological experiments and muscarinic receptor binding assays. The compound, N-[4-(2-chloroethylmethylamino)-2-butyny1]-2-pyrrolidone (BM 123), forms an aziridinium ion in aqueous solution at neutral pH that stimulates contractions of the guinea pig elium with a potency similar to that of oxotremorine. Following the initial stimulation, there is a long lasting period of lack of sensitivity of the guinea pig ileum to muscarinic agonits. BM 123 also produces muscarinic effects in vivo . When homogenates of the rat cerebral cortex were incubated with BM 123 and assayed subsequently in muscarinic receptor binding assays, a loss of binding capacity for the muscarinic antagonist, [ 3H]N-methylscopolamine ([ 3H]NMS), was noted without a change in affinity. Similar observations were made in [ 3H]--3-quinuclidinyl benzilate ([ 3H]1-QNB) binding assays on the forebrains of mice that had been injected with BM 123 24 hr earlier. The loss in receptor capacity for both [ 3H]NMS and [ 3H]1-QNB was prevented by atropine treatment. Kinetic studies of the interaction of BM 123 with homogenates of the rat cerebral cortex in vitro showed that the half-time for the loss of [ 3H]1-QNB binding sites increased from 10 to 45 min as the concentration of BM 123 decreased from 10 to 1 μM. In contrast to the aziridinium ion, the parent 2-chloroethylamine compound and the alcoholic hydrolysis product were largely devoid of pharmacological and binding activity.