Mitochondria and calcium flux as targets of neuroprotection caused by minocycline in cerebellar granule cells
Mitochondria and calcium flux as targets of neuroprotection caused by minocycline in cerebellar granule cells
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DOI：
10.1016/j.bcp.2009.07.028
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发表时间：
2010-01-15
影响因子：
5.8
通讯作者：
Jordan, Joaquin
中科院分区：
文献类型：
--
作者：
Maria Garcia-Martinez, Eva;Sanz-Blasco, Sara;Jordan, Joaquin
米诺环素是四环素家族的一种抗生素，因其在神经退行性疾病中的理论治疗应用而引起了相当大的兴趣。然而，其作用机制仍然难以捉摸。在这里，我们研究了米诺环素的兴奋性毒性条件下的影响。荧光和生物发光成像研究在大鼠小脑颗粒神经元培养物中使用fura 2/AM和靶向水母发光蛋白显示，米诺环素，在浓度高于那些显示阻止炎症和炎症诱导的神经元死亡，抑制NMDA诱导的细胞溶质和线粒体Ca 2+浓度的上升，在一个可逆的方式。此外，二甲胺四环素在NMDA刺激过程中加入降低细胞内Ca 2+的水平，但不诱导去极化与高K+介质。我们还发现，米诺环素，在相同的浓度，部分去极化线粒体约5-30 mV，防止线粒体Ca 2 +-摄取的条件下的环境压力，并废除NMDA诱导的活性氧（ROS）的形成。因此，米诺环素也消除了75 μ M Ca 2+在分离的脑线粒体中诱导的ROS升高。在线粒体去极化的机制研究中，我们发现米诺环素明显抑制大鼠脑线粒体的状态3呼吸，但明显增加状态4的氧摄取。米诺环素抑制NADH-细胞色素c还原酶和细胞色素c氧化酶的活性，而琥珀酸-细胞色素c还原酶的活性没有改变，这表明复合物I和IV的选择性抑制。最后，米诺环素影响的电压依赖性阴离子通道（VDAC）的活动，在重建系统中确定。总之，我们的研究结果表明，线粒体是米诺环素介导的神经保护的关键因素。(C)2009 Elsevier Inc. All rights reserved.
Minocycline, an antibiotic of the tetracycline family, has attracted considerable interest for its theoretical therapeutic applications in neurodegenerative diseases. However, the mechanism of action underlying its effect remains elusive. Here we have studied the effect of minocycline under excitotoxic conditions. Fluorescence and bioluminescence imaging studies in rat cerebellar granular neuron cultures using fura2/AM and mitochondria-targeted aequorin revealed that minocycline, at concentrations higher than those shown to block inflammation and inflammation-induced neuronal death, inhibited NMDA-induced cytosolic and mitochondrial rises in Ca2+ concentrations in a reversible manner. Moreover, minocycline added in the course of NMDA stimulation decreased Ca2+ intracellular levels, but not when induced by depolarization with a high K+ medium. We also found that minocycline, at the same concentrations, partially depolarized mitochondria by about 5-30 mV, prevented mitochondrial Ca2+-uptake under conditions of environmental stress, and abrogated NMDA-induced reactive oxygen species (ROS) formation. Consistently, minocycline also abrogates the rise in ROS induced by 75 mu M Ca2+ in isolated brain mitochondria. In search for the mechanism of mitochondrial depolarization, we found that minocycline markedly inhibited state 3 respiration of rat brain mitochondria, although distinctly increased oxygen uptake in state 4. Minocycline inhibited NADH-cytochrome c reductase and cytochrome c oxidase activities, whereas the activity of succinate-cytochrome c reductase was not modified, suggesting selective inhibition of complexes I and IV. Finally, minocycline affected activity of voltage-dependent anion channel (VDAC) as determined in the reconstituted system. Taken together, our results indicate that mitochondria are a critical factor in minocycline-mediated neuroprotection. (C) 2009 Elsevier Inc. All rights reserved.