EXPRESSION OF FUNCTIONAL MUSCARINIC ACETYLCHOLINE-RECEPTOR SUBTYPES IN HUMAN CORPUS CAVERNOSUM AND IN CULTURED SMOOTH-MUSCLE CELLS

Relaxation of the trabecular smooth muscle, which is necessary for pen ile erection, is controlled locally by neurotransmitters and vasoactiv e agents. The goal of this study was to identify and characterize musc arinic acetylcholine receptor (mAChR) subtypes expressed in cultured h uman corpus cavernosum smooth muscle cells (HCC SMC). Binding analysis with L-[benzilic-4,4'-H-3(N)]quinuclidinyl benzilate ([H-3]QNB) demon strated the expression of specific muscarinic receptor binding sites i n HCC SMC. Analysis of total RNA isolated from whole corpus cavernosum tissue and smooth muscle cells,by RNase protection assays, demonstrat ed the expression of mRNA transcripts for m1, m2, m3, and m4 mAChR sub types in whole tissue and m2 and m4 subtypes in cultured cells. In sit u hybridization with specific m2 and m4 probes further confirmed the e xpression of m2 and m4 mRNA transcripts in cultured cells. Carbachol ( CCh), a nonselective cholinergic agonist, inhibited cAMP synthesis at low concentrations (0.1-1 mu M) and stimulated cAMP synthesis at high concentrations (100 mu M), in cultured HCC SMC. CCh (100 mu M) further augmented forskolin (FSK), isoproterenol(ISO), and prostaglandin E(1) (PGE(1))-induced cAMP synthesis. These observations suggest that, in vivo, in HCC, ACh may activate m3 mAChR subtypes oil endothelial cells or m2 and m4 subtypes on the SMC. Although m2 and m4 are thought to i nhibit adenylate cyclase (AC), the augmentation of cAMP synthesis by h igh concentrations of CCh in SMC suggests an alternative mechanism of coupling to G-proteins that stimulates AC activity. These studies show that HCC tissue expresses different subtypes of mAChR (mi, m2, m3, an d m4), whereas cultured HCC SMC express m2 and m4 subtypes. It is sugg ested that m2 and m4 receptor subtypes may play an important role in m aintaining trabecular smooth muscle tone in vivo. The augmentation of FSK-, ISO, and PGE(1)-induced cAMP synthesis by CCh suggests possible development of a multidrug therapeutic approach to treatment of erecti le dysfunction.