Lazareno S, Buckley N J, Roberts F F
Department of Neuropharmacology, Glaxo Group Research Ltd., Ware Hertfordshire, United Kingdom.
Mol Pharmacol. 1990 Dec;38(6):805-15.
We have carried out an extensive pharmacological characterization of muscarinic binding sites in rabbit lung and chicken heart in parallel with M1, M2, and M3 sites, [3H]Pirenzepine, a selective antagonist at M1 receptors, bound saturably and reversibly to membranes from chicken heart and rabbit lung. These binding sites were not M1 receptors, however, because the cardioselective antagonist himbacine had 10-fold higher affinity at these sites than at [3H]pirenzepine sites in rat and rabbit cortex (true M1 sites). We measured the inhibitory potency of 28 antagonists at [3H]N-methylscopolamine-labeled sites in chicken heart, rabbit lung, rat heart (M2 sites), and rat submandibular gland (M3 sites) and at M1 sites in rat cortex. The sites in rabbit lung were different from M1, M2, and M3 sites, because they had moderate to high affinity for M1-selective compounds (pirenzepine and telenzepine), M2-selective compounds (himbacine and methoctramine), and M3-selective compounds (hexahydrosiladifenidol and 4-diphenylacetoxy-N-methylpiperidine methiodide). The sites in chicken heart resembled most those in rabbit lung, with similar high affinity for secoverine, but they were not the same because tropicamide, diphenylacetoxybutynyl dimethylamine, and [3H]-N-methylscopolamine were more potent in rabbit lung. In a further series of experiments, we compared the affinity of six of the most discriminating antagonists in membranes from rabbit lung and NG108-15 cells, a neuroblastoma-glioma cell line reported to express the muscarinic m4 receptor gene. The antagonists had very similar affinities in the two tissues, the largest discrepancy being that pirenzepine was twice as potent in rabbit lung as in NG108-15 cells. Northern blots using probes designed to discriminate between five species of muscarinic receptor RNA detected only m4 mRNA in rabbit lung. We conclude that rabbit lung contains a muscarinic M4 binding site with a quite distinctive pharmacology and that chicken heart contains a receptor with similarities to the M4 sites. This is the first report to characterize native M4 binding sites in a nonneuronal mammalian tissue.
我们对兔肺和鸡心中的毒蕈碱结合位点进行了广泛的药理学特性研究，并与M1、M2和M3位点进行了对比。[3H]哌仑西平是一种M1受体选择性拮抗剂，可饱和且可逆地结合于鸡心和兔肺的膜上。然而，这些结合位点并非M1受体，因为心脏选择性拮抗剂辛巴生在这些位点的亲和力比在大鼠和兔皮层（真正的M1位点）的[3H]哌仑西平位点高10倍。我们测定了28种拮抗剂对鸡心、兔肺、大鼠心脏（M2位点）和大鼠下颌下腺（M3位点）中[3H]N - 甲基东莨菪碱标记位点以及大鼠皮层中M1位点的抑制效力。兔肺中的位点不同于M1、M2和M3位点，因为它们对M1选择性化合物（哌仑西平和替仑西平）、M2选择性化合物（辛巴生和甲溴东莨菪碱）以及M3选择性化合物（六氢硅二苯乙啶和4 - 二苯基乙酰氧基 - N - 甲基哌啶甲碘化物）具有中度至高亲和力。鸡心中的位点与兔肺中的位点最为相似，对司可维林具有相似的高亲和力，但它们并不相同，因为托吡卡胺、二苯基乙酰氧基丁炔基二甲胺和[3H] - N - 甲基东莨菪碱在兔肺中的效力更强。在另一系列实验中，我们比较了六种最具区分性的拮抗剂对兔肺膜和NG108 - 15细胞（一种据报道表达毒蕈碱m4受体基因的神经母细胞瘤 - 胶质瘤细胞系）膜的亲和力。拮抗剂在这两种组织中的亲和力非常相似，最大的差异在于哌仑西平在兔肺中的效力是在NG108 - 15细胞中的两倍。使用旨在区分五种毒蕈碱受体RNA种类的探针进行的Northern印迹分析仅在兔肺中检测到m4 mRNA。我们得出结论，兔肺含有具有相当独特药理学特性的毒蕈碱M4结合位点，鸡心含有与M4位点相似的受体。这是首次在非神经元哺乳动物组织中对天然M4结合位点进行表征的报告。
