Selective inhibition by ethanol of mitochondrial calcium influx mediated by uncoupling protein-2 in relation to N-methyl-D-aspartate cytotoxicity in cultured neurons.
Selective inhibition by ethanol of mitochondrial calcium influx mediated by uncoupling protein-2 in relation to N-methyl-D-aspartate cytotoxicity in cultured neurons.
复制标题
乙醇对培养神经元中N-甲基-D-天冬氨酸细胞毒性相关的线粒体钙涌入的选择性抑制作用。
DOI：
10.1371/journal.pone.0069718
复制
发表时间：
2013
期刊：
影响因子：
3.7
通讯作者：
Yoneda Y
中科院分区：
文献类型：
--
作者：
Fukumori R;Takarada T;Nakazato R;Fujikawa K;Kou M;Hinoi E;Yoneda Y
我们发现线粒体解偶联蛋白-2(UCP2)参与了N-甲基-D-天冬氨酸受体(NMDAR)对HEK293细胞的细胞毒作用，其机制与NMDAR通道获得性线粒体钙水平升高有关。在这里，我们评估了乙醇对NMDA诱导的培养的小鼠新皮质神经元线粒体钙水平升高的药理作用。在暴露于谷氨酸或NMDA的神经元中，用Rhod-2法测定，在0.1~100µM浓度范围内，线粒体Ca~(2+)水平显著升高。此外，加入250 mM乙醇可显著抑制谷氨酸和NMDA引起的Rhod-2荧光强度的升高，而50~250 mM乙醇可显著抑制NMDA诱导的细胞内钙离子浓度升高。慢病毒过表达UCP2显著加速了NMDA对神经元Rhod-2荧光的增强，但不影响对细胞内钙离子水平的荧光。在高表达UCP2的神经元中，乙醇对NMDA诱导的线粒体游离钙水平升高的抑制作用显著高于未高表达UCP2的神经元，尽管与UCP2过表达无关，细胞内钙水平也同样有效地升高。在谷氨酸暴露的神经细胞中，UCP2的过表达显著增加了死亡细胞的数量，这种增加可以被乙醇阻止。在含有GluN2B亚基的NMDAR的HEK293细胞中，50和250 mM的乙醇对NMDA诱导的线粒体钙水平升高的抑制作用类似于含有GluN2A亚基的细胞。在含UCP2的免疫沉淀物中，短暂暴露于浓度超过50 mM的乙醇的神经元中，GluN2B亚单位的蛋白水平降低，但GluN2A亚单位的蛋白水平没有下降。乙醇可抑制UCP2和NMDAR通道之间的相互作用，从而阻止神经元NMDAR激活后线粒体钙离子的掺入和细胞死亡。
We have shown the involvement of mitochondrial uncoupling protein-2 (UCP2) in the cytotoxicity by N-methyl-D-aspartate receptor (NMDAR) through a mechanism relevant to the increased mitochondrial Ca2+ levels in HEK293 cells with acquired NMDAR channels. Here, we evaluated pharmacological profiles of ethanol on the NMDA-induced increase in mitochondrial Ca2+ levels in cultured murine neocortical neurons. In neurons exposed to glutamate or NMDA, a significant increase was seen in mitochondrial Ca2+ levels determined by Rhod-2 at concentrations of 0.1 to 100 µM. Further addition of 250 mM ethanol significantly inhibited the increase by glutamate and NMDA in Rhod-2 fluorescence, while similarly potent inhibition of the NMDA-induced increase was seen after exposure to ethanol at 50 to 250 mM in cultured neurons. Lentiviral overexpression of UCP2 significantly accelerated the increase by NMDA in Rhod-2 fluorescence in neurons, without affecting Fluo-3 fluorescence for intracellular Ca2+ levels. In neurons overexpressing UCP2, exposure to ethanol resulted in significantly more effective inhibition of the NMDA-induced increase in mitochondrial free Ca2+ levels than in those without UCP2 overexpression, despite a similarly efficient increase in intracellular Ca2+ levels irrespective of UCP2 overexpression. Overexpression of UCP2 significantly increased the number of dead cells in a manner prevented by ethanol in neurons exposed to glutamate. In HEK293 cells with NMDAR containing GluN2B subunit, more efficient inhibition was similarly induced by ethanol at 50 and 250 mM on the NMDA-induced increase in mitochondrial Ca2+ levels than in those with GluN2A subunit. Decreased protein levels of GluN2B, but not GluN2A, subunit were seen in immunoprecipitates with UCP2 from neurons with brief exposure to ethanol at concentrations over 50 mM. Ethanol could inhibit the interaction between UCP2 and NMDAR channels to prevent the mitochondrial Ca2+ incorporation and cell death after NMDAR activation in neurons.
登录
查看更多内容
影响因子：
4.1
作者：
CIK, M;CHAZOT, PL;STEPHENSON, FA
通讯作者：
STEPHENSON, FA
影响因子：
3.5
作者：
Fukumori, Ryo;Nakamichi, Noritaka;Yoneda, Yukio
通讯作者：
Yoneda, Yukio
影响因子：
4.7
作者：
Kambe, Yuki;Nakamichi, Noritaka;Yoneda, Yukio
通讯作者：
Yoneda, Yukio
内膜的四十千克蛋白是线粒体钙Uniporter。
影响因子：
64.8
作者：
De Stefani, Diego;Raffaello, Anna;Teardo, Enrico;Szabo, Ildiko;Rizzuto, Rosario
通讯作者：
Rizzuto, Rosario
影响因子：
64.8
作者：
Baines, CP;Kaiser, RA;Molkentin, JD
通讯作者：
Molkentin, JD