Characterisation of recombinant rat TRPM2 and a TRPM2-like conductance in cultured rat striatal neurones
Characterisation of recombinant rat TRPM2 and a TRPM2-like conductance in cultured rat striatal neurones
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DOI：
10.1016/j.neuropharm.2005.08.021
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发表时间：
2006-01-01
影响因子：
4.7
通讯作者：
Randall, AD
中科院分区：
文献类型：
--
作者：
Hill, K;Tigue, NJ;Randall, AD
TRPM2是TRP离子通道家族的一员，在单核细胞谱系的大脑和免疫细胞中均有表达。在功能上，它在细胞内adp核糖和氧化和亚硝化应激下的激活是独特的。迄今为止，该通道的研究主要集中在人重组通道和啮齿动物的天然制剂上。这为基于重组通道表型的本地组织观察的解释提供了跨物种并发症的可能性。因此，我们在HEK293细胞中克隆并异源表达了大鼠TRPM2 （rTRPM2）。我们发现，像hTRPM2一样，它以依赖于细胞外Ca2+的方式响应细胞内adp核糖。在单通道水平，rTRPM2是一个缓慢的门控，大电导（84 pS）通道，在孤立的膜斑块中迅速下降。在药理学上，rTRPM2被氯霉唑（10 μ M）快速不可逆阻断，因此类似于hTRPM2，而不是大鼠源性胰岛素瘤CRI-G1的trpm2样电流，表现出可逆性抑制。我们发现，培养的大鼠纹状体神经元在全细胞和单通道水平上都表现出adp核糖激活的传导。从药理学上讲，这种神经元电流可以被克霉唑不可逆地抑制。它对细胞外Ca2+的去除也很敏感，这表明它是由含有trpm2的通道介导的。这些数据提供了异源表达rTRPM2的功能特征，并证明，除了先前在免疫细胞、小胶质细胞和胰岛素瘤中的描述外，可以在来自啮齿动物中枢神经系统的神经元中发现trpm2样电导。(c) 2005 Elsevier Ltd版权所有。
TRPM2, a member of the TRP ion channel family, is expressed both in the brain and immune cells of the monocyte lineage. Functionally, it is unique in its activation by intracellular ADP-ribose and both oxidative and nitrosative stress. To date studies of this channel have concentrated on human recombinant channels and rodent native preparations. This provides the potential for cross-species complications in the interpretation of native tissue observations based on recombinant channel phenotype. Consequently, we have cloned and heterologously expressed rat TRPM2 (rTRPM2) in HEK293 cells. We find that, like hTRPM2, it responds to intracellular ADP-ribose in a manner dependent on extracellular Ca2+. At the single channel level rTRPM2 is a slow gating, large conductance (84 pS) channel that rapidly runs down in isolated membrane patches. Pharmacologically, rTRPM2 is rapidly and irreversibly blocked by clotrimazole (10 mu M), thus resembling hTRPM2 but not the TRPM2-like current of the rat-derived insulinoma CRI-G1, which exhibits reversible inhibition by this agent. We show that cultured rat striatal neurones exhibit an ADP-ribose-activated conductance at both the whole cell and single channel level. Pharmacologically this neuronal current can be irreversibly inhibited by clotrimazole. It is also sensitive to removal of extracellular Ca2+, suggesting that it is mediated by TRPM2-containing channels. These data provide a functional characterisation of heterologously expressed rTRPM2 and demonstrate that, in addition to the previous descriptions in immune cells, microglia and insulinomas, a TRPM2-like conductance can be found in neurones derived from the rodent CNS. (c) 2005 Elsevier Ltd. All rights reserved.