Pharmacological characterization of recombinant N-type calcium channel (Cav2.2) mediated calcium mobilization using FLIPR
Pharmacological characterization of recombinant N-type calcium channel (Cav2.2) mediated calcium mobilization using FLIPR
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DOI：
10.1016/j.bcp.2006.06.003
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发表时间：
2006-09-14
影响因子：
5.8
通讯作者：
Ilyin, Victor I.
中科院分区：
文献类型：
--
作者：
Benjamin, Elfrida R.;Pruthi, Farhana;Ilyin, Victor I.
N 型电压门控钙通道 (Ca(v)2.2) 在神经元中发挥调节神经递质释放的作用。它包含慢性疼痛的临床相关目标。我们已经验证了一种钙动员方法，可在两种稳定的 Ca(v)2.2 细胞系中评估 Ca(v)2.2 药理学：α 1(B)、α 2 delta、β(3)-HEK-293 和 α 1(B)、β(3)-HEK-293。通过添加 KCl 和 Ca2+ 打开 Ca(v)2.2 通道，通过荧光成像板读数器 (FLIPR96) 上的 Fluo-4 荧光测量动员情况。 Ca(v)2.2 表达和生物物理学通过膜片钳电生理学 (EP) 得到证实。两种细胞系均以足够的信号背景响应 KC1。来自两种细胞系的信号均被 omega-芋螺毒素 (ctx)-MVIIa 和 omega-芋螺毒素 (ctx)-GVIa 抑制，对于三亚基稳定的 IC50 值分别为 1.8 和 1 nM，对于双亚基稳定的 IC50 值分别为 0.9 和 0.6 nM。其他已知的 Ca(v)2.2 阻滞剂包括镉、氟桂利嗪、fluspirilene 和米贝拉地尔。 IC50 值与文献 EP 得出的值相关。新的 Ca(v)2.2 药理学在具有其他主要药理活性的化合物类别中得到鉴定，包括 Na+ 通道抑制剂和抗抑郁药。在苯氧基苯基吡啶、苯氧基苯基吡唑和其他类别中鉴定出对 Ca(v)2.2 具有高效能的新型 Na+ 通道化合物。已确定 Ca(v)2.2 三环类抗抑郁药中效力最高的是地昔帕明。 (c) 2006 Elsevier Inc. 保留所有权利。
The N-type voltage-gated calcium channel (Ca(v)2.2) functions in neurons to regulate neurotransmitter release. It comprises a clinically relevant target for chronic pain. we have validated a calcium mobilization approach to assessing Ca(v)2.2 pharmacology in two stable Ca(v)2.2 cell lines: alpha 1(B), alpha 2 delta, beta(3)-HEK-293 and alpha 1(B), beta(3)-HEK-293. Ca(v)2.2 channels were opened by addition of KCl and Ca2+, mobilization was measured by Fluo-4 fluorescence on a fluorescence imaging plate reader (FLIPR96). Ca(v)2.2 expression and biophysics were confirmed by patch-clamp electrophysiology (EP). Both cell lines responded to KC1 with adequate signal-to-background. Signals from both cell lines were inhibited by omega-conotoxin (ctx)-MVIIa and omega-conotoxin (ctx)-GVIa with IC50 values of 1.8 and 1 nM, respectively, for the three-subunit stable, and 0.9 and 0.6 nM, respectively, for the two-subunit stable. Other known Ca(v)2.2 blockers were characterized including cadmium, flunarizine, fluspirilene, and mibefradil. IC50 values correlated with literature EP-derived values. Novel Ca(v)2.2 pharmacology was identified in classes of compounds with other primary pharmacological activities, including Na+ channel inhibitors and antidepressants. Novel Na+ channel compounds with high potency at Ca(v)2.2 were identified in the phenoxyphenyl pyridine, phenoxyphenyl pyrazole, and other classes. The highest potency at Ca(v)2.2 tricyclic antidepressant identified was desipramine. (c) 2006 Elsevier Inc. All rights reserved.