Na+/H+ exchanger inhibitor, SM-20220, is protective against excitotoxicity in cultured cortical neurons
Na+/H+ exchanger inhibitor, SM-20220, is protective against excitotoxicity in cultured cortical neurons
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DOI：
10.1161/01.str.0000106910.42815.c2
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发表时间：
2004-01-01
期刊：
影响因子：
8.3
通讯作者：
Umemura, K
中科院分区：
文献类型：
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作者：
Matsumoto, Y;Yamamoto, S;Umemura, K
背景与目的：近年来，有报道称Na+/H+交换剂（NHE）抑制剂在动物模型中显示出对缺血/再灌注脑损伤的保护作用。然而，保护神经元免受缺血性损伤的机制尚不清楚。为了揭示NHE抑制剂对神经元死亡的细胞机制，我们研究了选择性NHE抑制剂SM-20220 （N-[氨基亚甲基]-1-甲基- 1h -吲哚-2-羧酰胺甲磺酸盐）对谷氨酸诱导的大鼠皮质培养神经元死亡的影响。方法：制备1日龄大鼠皮层神经元，在玻璃培养皿中培养。用碘化丙啶染色评估谷氨酸诱导的神经元死亡。用视频增强差示干涉对比显微镜观察神经元形态学变化。用共聚焦激光显微镜，以荧光染料fluo-3/acetoxymethylester （AM）和2',7'-双-2-羧基乙基-5(6)-羧基荧光素(BCECF)/AM为荧光染料，分别测定细胞内钙浓度（[Ca2+](i)）和细胞内pH（pH(i)）。结果：sm -20220 （0.3 ~ 30 nmol/L）剂量依赖性减毒谷氨酸（300 mumol/L）在6小时内诱导神经元死亡，并抑制谷氨酸（500 mumol/L）暴露后的急性细胞肿胀。在谷氨酸（500 μ mol/L）暴露后5分钟和12分钟观察到[Ca2+](i)上升的双峰，随后持续上升。SM-20220抑制持续的[Ca2+](i)升高。SM-20220在谷氨酸（500 μ mol/L）暴露后抑制细胞内酸化。谷氨酸诱导的所有事件也被n -甲基- d -天冬氨酸受体拮抗剂MK-801所抑制，表明死亡过程是兴奋毒性的。结论- nhe抑制剂通过抑制持续的[Ca2+](i)升高和兴奋毒性的酸化而具有神经保护作用。
Background and Purpose-Recently, it has been reported that Na+/H+ exchanger (NHE) inhibitors demonstrated protective effects on ischemia/reperfusion brain injury in animal models. However, the mechanisms by which the neurons were protected against ischemic insult remain unclear. To reveal the cellular mechanism of the NHE inhibitor on the neuronal death, we examined the effects of a selective NHE inhibitor, SM-20220 (N-[aminoiminomethyl]-1-methyl-1H-indole-2-carboxamide methanesulfonate), on glutamate-induced neuronal death in rat cortical culture.Methods-Cortical neurons were prepared from 1-day old rats, and cultured on the glass-based dishes. Glutamate-induced neuronal death was assessed by staining the cells with propidium iodide. Morphological changes in the neurons were observed with a video-enhanced contrast-differential interference contrast microscope. The intracellular calcium concentration ([Ca2+](i)) and the intracellular pH (pH(i)) were measured by fluorescence imaging with a confocal laser microscope using fluo-3/acetoxymethylester (AM) and 2', 7'-bis-2-carboxy-ethyl-5(6)-carboxyfluorescein (BCECF)/AM as a fluorescent dye, respectively.Results-SM-20220 (0.3 to 30 nmol/L) dose-dependently attenuated glutamate (300 mumol/L)-induced neuronal death over a period of 6 hours, and inhibited the acute cellular swelling following glutamate (500 mumol/L) exposure. Dual peaks of [Ca2+](i) rise were observed at 5 and 12 minutes after glutamate (500 mumol/L) exposure, followed by a persistent rise. SM-20220 suppressed the persistent [Ca2+](i) increase. SM-20220 inhibited intracellular acidification following glutamate (500 mumol/L) exposure. All of the events induced by glutamate were also inhibited by the N-methyl-D-aspartate receptor antagonist, MK-801, indicating the death process was excitotoxicity.Conclusions-NHE inhibitor is neuroprotective through inhibition of both persistent [Ca2+](i) increase and acidification in excitotoxicity.