Cloning and expression of a small-conductance Ca(2+)-activated K+ channel from the mouse cochlea: coexpression with alpha9/alpha10 acetylcholine receptors.
Cloning and expression of a small-conductance Ca(2+)-activated K+ channel from the mouse cochlea: coexpression with alpha9/alpha10 acetylcholine receptors.
复制标题
小鼠耳蜗小电导 Ca(2) 激活 K 通道的克隆和表达：与 alpha9/alpha10 乙酰胆碱受体共表达。
DOI：
10.1152/jn.00630.2003
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发表时间：
2004
影响因子：
2.5
通讯作者：
Vázquez,AnaE
中科院分区：
文献类型：
--
作者：
Nie,Liping;Song,Haitao;Chen,Mei-Fang;Chiamvimonvat,Nipavan;Beisel,KirkW;Yamoah,EbenezerN;Vázquez,AnaE
配体门控离子通道、电压激活离子通道和钙激活离子通道之间的功能相互作用对可兴奋细胞的特性至关重要，从而对神经系统的工作至关重要。哺乳动物耳蜗外毛细胞通过胆碱能神经纤维从脑干接受传出输入，胆碱能神经纤维在其底部形成突触。从这些传出纤维释放的乙酰胆碱激活突触后快速抑制性电流，在一定程度上是由尚未克隆的小电导钙激活的钾通道(SK)介导的。在这里，我们报道了从小鼠耳蜗组织中克隆、鉴定和表达完整的SK2基因。我们还获得了小鼠耳蜗乙酰胆碱受体α-9和α-10的基因片段，并对其进行了测序，并与SK2通道共表达。转导SK2的人培养细胞产生钙敏感钾电流，该电流可被已知的SK通道阻断剂地奎宁和阿帕明阻断。注射SK2体外转录的RNA的非洲爪哇卵母细胞，在只能记录到外向K+电流的条件下，表达一种对EGTA、地奎宁和阿帕明敏感的外向电流。在共转染有耳蜗SK2和α9/α10受体的HEK293细胞中，乙酰胆碱诱导了一种内向电流，随后是一种强劲的外向电流。结果表明，SK2和α9/α10乙酰胆碱受体足以部分概括毛细胞传出突触反应。
Functional interactions between ligand-gated, voltage-, and Ca2+-activated ion channels are essential to the properties of excitable cells and thus to the working of the nervous system. The outer hair cells in the mammalian cochlea receive efferent inputs from the brain stem through cholinergic nerve fibers that form synapses at their base. The acetylcholine released from these efferent fibers activates fast inhibitory postsynaptic currents mediated, to some extent, by small-conductance Ca2+-activated K+channels (SK) that had not been cloned. Here we report the cloning, characterization, and expression of a complete SK2 cDNA from the mouse cochlea. The cDNAs of the mouse cochlea α9 and α10 acetylcholine receptors were also obtained, sequenced, and coexpressed with the SK2 channels. Human cultured cell lines transfected with SK2 yielded Ca2+-sensitive K+current that was blocked by dequalinium chloride and apamin, known blockers of SK channels. Xenopus oocytes injected with SK2 in vitro transcribed RNA, under conditions where only outward K+currents could be recorded, expressed an outward current that was sensitive to EGTA, dequalinium chloride, and apamin. In HEK-293 cells cotransfected with cochlear SK2 plus α9/α10 receptors, acetylcholine induced an inward current followed by a robust outward current. The results indicate that SK2 and the α9/α10 acetylcholine receptors are sufficient to partly recapitulate the native hair cell efferent synaptic response.