Traish A M, Palmer M S, Goldstein I, Moreland R B
Department of Biochemistry, Boston University School of Medicine, MA 02118, USA.
Receptor. 1995 Fall;5(3):159-76.
Relaxation of the trabecular smooth muscle, which is necessary for penile erection, is controlled locally by neurotransmitters and vasoactive agents. The goal of this study was to identify and characterize muscarinic acetylcholine receptor (mAChR) subtypes expressed in cultured human corpus cavernosum smooth muscle cells (HCC SMC). Binding analysis with L-[benzilic-4,4'-3H(N)]quinuclidinyl benzilate ([3H]QNB) demonstrated the expression of specific muscarinic receptor binding sites in HCC SMC. Analysis of total RNA isolated from whole corpus cavernosum tissue and smooth muscle cells, by RNase protection assays, demonstrated the expression of mRNA transcripts for m1, m2, m3, and m4 mAChR subtypes in whole tissue and m2 and m4 subtypes in cultured cells. In situ hybridization with specific m2 and m4 probes further confirmed the expression of m2 and m4 mRNA transcripts in cultured cells. Carbachol (CCh), a nonselective cholinergic agonist, inhibited cAMP synthesis at low concentrations (0.1-1 microM) and stimulated cAMP synthesis at high concentrations (100 microM), in cultured HCC SMC. CCh (100 microM) further augmented forskolin (FSK), isoproterenol (ISO), and prostaglandin E1 (PGE1)-induced cAMP synthesis. These observations suggest that, in vivo, in HCC, ACh may activate m3 mAChR subtypes on endothelial cells or m2 and m4 subtypes on the SMC. Although m2 and m4 are thought to inhibit adenylate cyclase (AC), the augmentation of cAMP synthesis by high 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 (m1, m2, m3, and m4), whereas cultured HCC SMC express m2 and m4 subtypes. It is suggested that m2 and m4 receptor subtypes may play an important role in maintaining trabecular smooth muscle tone in vivo. The augmentation of FSK-, ISO, and PGE1-induced cAMP synthesis by CCh suggests possible development of a multidrug therapeutic approach to treatment of erectile dysfunction.
小梁平滑肌的舒张是阴茎勃起所必需的，它由神经递质和血管活性物质进行局部调控。本研究的目的是鉴定并表征培养的人海绵体平滑肌细胞（HCC SMC）中表达的毒蕈碱型乙酰胆碱受体（mAChR）亚型。用L-[苯甲酰-4,4'-3H(N)]喹核醇基苯甲酸酯（[3H]QNB）进行的结合分析表明HCC SMC中存在特异性毒蕈碱受体结合位点。通过核糖核酸酶保护分析对从整个海绵体组织和平滑肌细胞中分离的总RNA进行分析，结果表明在整个组织中m1、m2、m3和m4 mAChR亚型的mRNA转录本有表达，而在培养细胞中m2和m4亚型有表达。用特异性m2和m4探针进行的原位杂交进一步证实了培养细胞中m2和m4 mRNA转录本的表达。在培养的HCC SMC中，非选择性胆碱能激动剂卡巴胆碱（CCh）在低浓度（0.1 - 1 microM）时抑制cAMP合成，在高浓度（100 microM）时刺激cAMP合成。CCh（100 microM）进一步增强了福斯高林（FSK）、异丙肾上腺素（ISO）和前列腺素E1（PGE1）诱导的cAMP合成。这些观察结果表明，在体内，在HCC中，乙酰胆碱（ACh）可能激活内皮细胞上的m3 mAChR亚型或SMC上的m2和m4亚型。尽管m2和m4被认为可抑制腺苷酸环化酶（AC），但高浓度CCh在SMC中增强cAMP合成表明存在一种与刺激AC活性的G蛋白偶联的替代机制。这些研究表明HCC组织表达不同亚型的mAChR（m1、m2、m3和m4），而培养的HCC SMC表达m2和m4亚型。提示m2和m4受体亚型可能在体内维持小梁平滑肌张力方面发挥重要作用。CCh增强FSK、ISO和PGE1诱导的cAMP合成提示可能开发出一种治疗勃起功能障碍的多药治疗方法。
