A possible pivotal role of mitochondrial free calcium in neurotoxicity mediated by N-methyl-D-aspartate receptors in cultured rat hippocampal neurons
A possible pivotal role of mitochondrial free calcium in neurotoxicity mediated by N-methyl-D-aspartate receptors in cultured rat hippocampal neurons
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DOI：
10.1016/j.neuint.2011.03.018
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发表时间：
2011-08-01
影响因子：
4.2
通讯作者：
Yoneda, Yukio
中科院分区：
文献类型：
--
作者：
Kambe, Yuki;Nakamichi, Noritaka;Yoneda, Yukio
我们之前的研究表明，线粒体膜电位破坏参与了大鼠皮层和海马制备的原代培养神经元对n -甲基- d -天冬氨酸（NMDA）受体介导的兴奋毒性差异脆弱性的机制。为了进一步阐明NMDA受体激活后线粒体在兴奋毒性中的作用，我们在神经元细胞质和细胞器中加载可扩散的荧光染料钙黄蛋白，测定不同线粒体分子渗漏的线粒体通透性过渡孔（mPTP）的活性。在两种培养的神经元中，CoCl2的加入同样淬灭了除线粒体外的细胞内荧光，而进一步添加NMDA导致染料渗漏到海马神经元的细胞质中。mPTP抑制剂阻止了nmda诱导的海马神经元活力丧失，而mPTP激活剂诱导皮层和海马神经元活力丧失同样有效。虽然NMDA在增加罗丹明-2荧光作为海马线粒体钙指标方面比皮质神经元更有效，但线粒体钙单转运抑制剂可显著防止NMDA诱导的海马神经元活力丧失。假设的单转运蛋白-2在海马中的mRNA表达明显高于皮质神经元。这些结果表明，通过促进海马神经元mPTP协调的机制，线粒体钙单转运体至少在一定程度上负责NMDA神经毒性。(C) 2011 Elsevier B.V.版权所有
We have previously shown that mitochondrial membrane potential disruption is involved in mechanisms underlying differential vulnerabilities to the excitotoxicity mediated by N-methyl-D-aspartate (NMDA) receptors between primary cultured neurons prepared from rat cortex and hippocampus. To further elucidate the role of mitochondria in the excitotoxicity after activation of NMDA receptors, neurons were loaded with the fluorescent dye calcein diffusible in the cytoplasm and organelles for determination of the activity of mitochondrial permeability transition pore (mPTP) responsible for the leakage of different mitochondrial molecules. The addition of CoCl2 similarly quenched the intracellular fluorescence except mitochondria in both cultured neurons, while further addition of NMDA led to a leakage of the dye into the cytoplasm in hippocampal neurons only. An mPTP inhibitor prevented the NMDA-induced loss of viability in hippocampal neurons, while an activator of mPTP induced a similarly potent loss of viability in cortical and hippocampal neurons. Although NMDA was more effective in increasing rhodamine-2 fluorescence as a mitochondrial calcium indicator in hippocampal than cortical neurons, a mitochondrial calcium uniporter inhibitor significantly prevented the NMDA-induced loss of viability in hippocampal neurons. Expression of mRNA was significantly higher for the putative uniporter uncoupling protein-2 in hippocampal than cortical neurons. These results suggest that mitochondrial calcium uniporter would be at least in part responsible for the NMDA neurotoxicity through a mechanism relevant to promotion of mPTP orchestration in hippocampal neurons. (C) 2011 Elsevier B.V. All rights reserved.