Characterization of muscarinic receptors in guinea pig ileum longitudinal smooth muscle

E Giraldo, MA Vigano, R Hammer and H Ladinsky

Department of Biochemistry, Istituto De Angeli S.p.A., Milan, Italy.

Heterogeneity in the muscarinic receptor population of guinea pig ileum longitudinal smooth muscle was found in competition binding experiments against N-methyl[3H]scopolamine using either a cardioselective (AF-DX 116) or a smooth muscle-selective (hexahydrosiladifenidol) antimuscarinic compound. AF-DX 116 recognized 65% of the total receptors with high affinity and 35% with low affinity. Hexahydrosiladifenidol distinguished 24% of the total receptors with high affinity and 76% with low affinity. The two affinity binding constants displayed in smooth muscle by the compounds were similar to those of heart and glands, suggesting that the muscarinic receptor population in the smooth muscle is formed of about 30% glandular type and 70% cardiac type of the M2 receptors. In dissociation experiments, the rate of breakdown of the N-methyl[3H]scopolamine receptor complex in the smooth muscle was rapid and similar to the dissociation of N- methyl[3H]scopolamine from muscarinic receptors in cardiac membranes, supporting the evidence for the presence of a large fraction of the cardiac receptor type in smooth muscle. To further characterize the population of the smooth muscle receptors recognized as glandular type, we performed protection experiments with hexahydrosiladifenidol, which binds to glandular M3 receptors with high affinity. Smooth muscle membranes were initially incubated with this compound and then phenoxybenzamine was added to irreversibly alkylate the remaining unprotected receptors. Data from competition and dissociation binding experiments showed that, under these conditions, this protected fraction of the total receptor population in ileum smooth muscle had all the characteristics of the glandular type, i.e., slow N- methyl[3H]scopolamine dissociation and affinity constants for a series of selective and nonselective muscarinic antagonists in the same order of magnitude as those found in the glandular tissue. These findings, together with the known observation that hexahydrosiladifenidol is more potent in inhibiting the functional activation of muscarinic receptors in smooth muscle relative to heart, lead to the hypothesis that smooth muscle contractility is mediated by a muscarinic receptor subtype similar to that found in glandular tissue.

Volume 33, Issue 6, pp. 617-625, 06/01/1988
Copyright © 1988 by American Society for Pharmacology and Experimental Therapeutics

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