Thomas Rachel L, Langmead Christopher J, Wood Martyn D, Challiss R A John
Department of Cell Physiology and Pharmacology, University of Leicester, Room 4/04, Henry Wellcome Building, Lancaster Road, Leicester, LE1 9HN, United Kingdom.
J Pharmacol Exp Ther. 2009 Dec;331(3):1086-95. doi: 10.1124/jpet.109.160242. Epub 2009 Sep 18.
A new class of subtype-selective muscarinic acetylcholine (mACh) receptor agonist that activates the receptor through interaction at a site distinct from the orthosteric acetylcholine binding site has been reported recently. Here, we have compared the effects of orthosteric (oxotremorine-M, arecoline, pilocarpine) and allosteric [4-n-butyl-1-[4-(2-methylphenyl)-4-oxo-1-butyl] piperidine (AC-42); 1-[3-(4-butyl-1-piperidinyl)propyl]-3,4-dihydro-2(1H)-quinolinone (77-LH-28-1)] agonists on M(1) mACh receptor internalization and down-regulation, as well as functional coupling in a Chinese hamster ovary (CHO) cell line. In contrast to full and partial orthosteric agonists, which cause significant receptor internalization and down-regulation, prolonged exposure to AC-42 did not significantly alter either cell-surface or total cellular M(1) mACh receptor expression. 77-LH-28-1, an AC-42 homolog, did cause some receptor internalization, but not down-regulation. The presence of atropine completely prevented the orthosteric agonist-induced adaptive changes in receptor populations; however, in contrast, the copresence of atropine and AC-42 significantly increased both cell-surface receptor and total M(1) mACh receptor expression. Maximal phosphoinositide hydrolysis responses to the partial agonist arecoline were similar in CHO-M(1) cells pretreated for 24 h with either AC-42 or vehicle; in contrast, these responses were markedly reduced when cells were pretreated with oxotremorine-M or pilocarpine. These data indicate that, whereas AC-42 binding to the M(1) mACh receptor can initiate signal transduction, the AC-42-liganded receptor is resistant to the usual mechanisms regulating receptor internalization and down-regulation. In addition, our data suggest unusual interactions between allosteric agonists and orthosteric antagonists to regulate cell-surface and total cellular receptor expression.
最近报道了一类新型的亚型选择性毒蕈碱型乙酰胆碱（mACh）受体激动剂，其通过与正构乙酰胆碱结合位点不同的位点相互作用来激活受体。在此，我们比较了正构（氧化震颤素-M、槟榔碱、毛果芸香碱）和变构[4-正丁基-1-[4-(2-甲基苯基)-4-氧代-1-丁基]哌啶（AC-42）；1-[3-(4-丁基-1-哌啶基)丙基]-3,4-二氢-2(1H)-喹啉酮（77-LH-28-1）]激动剂对M(1) mACh受体内化、下调以及在中国仓鼠卵巢（CHO）细胞系中的功能偶联的影响。与导致显著受体内化和下调的完全和部分正构激动剂不同，长时间暴露于AC-42并未显著改变细胞表面或总细胞M(1) mACh受体表达。77-LH-28-1是AC-42的同系物，确实引起了一些受体内化，但没有下调。阿托品的存在完全阻止了正构激动剂诱导的受体群体适应性变化；然而，相比之下，阿托品和AC-42同时存在显著增加了细胞表面受体和总M(1) mACh受体表达。在用AC-42或载体预处理24小时的CHO-M(1)细胞中，对部分激动剂槟榔碱的最大磷酸肌醇水解反应相似；相比之下，当细胞用氧化震颤素-M或毛果芸香碱预处理时，这些反应明显降低。这些数据表明，虽然AC-42与M(1) mACh受体结合可以启动信号转导，但AC-42配体化的受体对调节受体内化和下调的常见机制具有抗性。此外，我们的数据表明变构激动剂和正构拮抗剂之间存在异常相互作用，以调节细胞表面和总细胞受体表达。
