Expression and functional characterisation of a human chimeric nicotinic receptor with α6β4 properties
Expression and functional characterisation of a human chimeric nicotinic receptor with α6β4 properties
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DOI：
10.1016/s0014-2999(03)01540-1
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发表时间：
2003-04-11
影响因子：
5
通讯作者：
Broad, LM
中科院分区：
文献类型：
--
作者：
Evans, NM;Bose, S;Broad, LM
尽管几年前就已被克隆，但含有人α 6亚基的功能性尼古丁乙酰胆碱受体在重组哺乳动物细胞系中的表达尚未得到证实。由此产生的选择性配体的缺乏阻碍了含α 6烟碱受体的作用的评价。我们报告称，用嵌合α 6/α 4亚基和β 4尼古丁受体亚基共转染人胚肾（HEK-293）细胞后记录了功能通道。在荧光成像板读数器测定中，它们显示出地棘蛙素的激动剂等级顺序效力远大于1，1-二甲基-4-苯基哌嗪鎓（DMPP），大于或等于野靛碱>乙酰胆碱>尼古丁。尼古丁，野靛碱，DMPP和epibatidine表现出部分激动剂的特性，而α-芋螺毒素MII和methyllycaconitine阻断乙酰胆碱刺激引起的功能反应。α 6/α 4嵌合体与2烟碱受体亚基的共转染没有产生功能性受体。在HEK-293细胞中表达的人α 6/α 4 β 4嵌合烟碱受体可以为亚型特异性配体的产生提供有价值的工具。(C)2003 Elsevier Science B. V.保留所有权利。
Despite being cloned several years ago, the expression of functional nicotinic acetylcholine receptors containing the human alpha6 subunit in recombinant mammalian cell lines has yet to be demonstrated. The resulting lack of selective ligands has hindered the evaluation of the role of alpha6-containing nicotinic receptors. We report that functional channels were recorded following co-transfection of human embryonic kidney (HEK-293) cells with a chimeric alpha6/alpha4 subunit and the beta4 nicotinic receptor subunit. They displayed an agonist rank order potency of epibatidine much greater than 1,1-dimethyl-4-phenylpiperazinium (DMPP) greater than or equal to cytisine>acetylcholine>nicotine measured in a fluorescent imaging plate reader assay. Nicotine, cytisine, DMPP and epibatidine displayed partial agonist properties whilst alpha-conotoxin MII and methyllycaconitine blocked the functional responses elicited by acetylcholine stimulation. Co-transfection of the alpha6/alpha4 chimera with the 2 nicotinic receptor subunit did not result in functional receptors. The human alpha6/alpha4beta4 chimeric nicotinic receptor expressed in HEK-293 cells may provide a valuable tool for the generation of subtype specific ligands. (C) 2003 Elsevier Science B.V. All rights reserved.