Pandya Jignesh D, Pauly James R, Sullivan Patrick G
Spinal Cord and Brain Injury Research Center, University of Kentucky, Lexington, 40536, USA.
Exp Neurol. 2009 Aug;218(2):381-9. doi: 10.1016/j.expneurol.2009.05.023. Epub 2009 May 27.
Experimental traumatic brain injury (TBI) leads to a rapid and extensive necrosis at the primary site of injury that appears to be driven in part by significant mitochondrial dysfunction. The present study is based on the hypothesis that TBI-induced, aberrant glutamate release increases mitochondrial Ca(2+) cycling/overload ultimately leading to mitochondrial damage. Previous work from our laboratory demonstrates that mitochondrial uncoupling during the acute phases of TBI-induced excitotoxicity can reduce mitochondrial Ca(2+) uptake (cycling), ROS production and mitochondrial damage resulting in neuroprotection and improved behavioral outcome. The current study was designed to determine the optimal dosage and therapeutic window of opportunity for the potent mitochondrial uncoupler FCCP following moderate TBI. For this study, we used young adult male Sprague-Dawley rats (300-350 g); either sham-operated or moderately (1.5 mm) injured using the controlled cortical impactor (CCI) model of TBI. In the first set of studies animals were injected with either vehicle (100% DMSO) or different concentrations of FCCP (0.5, 1, 2.5 and 5 mg/kg in 100% DMSO) intraperitoneally at 5 min post-injury; tested behaviorally at 10 days and cortical sparing assessed at 18 days post-injury. The results demonstrate that of all the dosages tested, 2.5 mg/kg rendered the maximum improvement in behavioral outcomes and tissue spared. Using this optimal dose (2.5 mg/kg) and time point for intervention (5 min post-injury), we assessed mitochondrial bioenergetics and mitochondrial structural integrity 24 h post-injury. Furthermore, using this dosage we assessed mitochondrial bioenergetics and Ca(2+) loading at 3 and 6 h post-injury to further verify our target mechanism and establish these assessments as a valid endpoint to use as a means to determine the therapeutic window of FCCP. To begin to address the window of opportunity for maintaining mitochondrial homeostasis, the optimal dose of FCCP was then administered at 5 min, 3, 6, or 24 h post-injury and several parameters of mitochondrial function were used as outcome measures. The results demonstrate that a prolonged window of opportunity exists for targeting mitochondrial dysfunction using uncouplers following TBI and give insight into the cellular pathology associated with TBI.
实验性创伤性脑损伤（TBI）会在损伤的原发部位导致快速且广泛的坏死，这似乎部分是由显著的线粒体功能障碍所驱动。本研究基于这样的假设：TBI诱导的异常谷氨酸释放会增加线粒体钙（Ca²⁺）循环/过载，最终导致线粒体损伤。我们实验室之前的工作表明，在TBI诱导的兴奋性毒性急性期，线粒体解偶联可以减少线粒体钙（Ca²⁺）摄取（循环）、活性氧生成和线粒体损伤，从而产生神经保护作用并改善行为结果。本研究旨在确定中度TBI后强效线粒体解偶联剂羰基氰化物4-(三氟甲氧基)苯腙（FCCP）的最佳剂量和治疗时机窗。在本研究中，我们使用了年轻成年雄性Sprague-Dawley大鼠（300 - 350克）；通过TBI的控制皮质撞击（CCI）模型进行假手术或中度（1.5毫米）损伤。在第一组研究中，动物在受伤后5分钟腹腔注射溶剂（100%二甲亚砜）或不同浓度的FCCP（在100%二甲亚砜中为0.5、1、2.5和5毫克/千克）；在10天时进行行为测试，并在受伤后18天评估皮质保留情况。结果表明，在所有测试剂量中，2.5毫克/千克使行为结果和保留的组织有最大程度的改善。使用这个最佳剂量（2.5毫克/千克）和干预时间点（受伤后5分钟），我们在受伤后24小时评估线粒体生物能量学和线粒体结构完整性。此外，使用这个剂量，我们在受伤后3小时和6小时评估线粒体生物能量学和钙（Ca²⁺）负载，以进一步验证我们的目标机制，并将这些评估确立为一个有效的终点，作为确定FCCP治疗时机窗的一种手段。为了开始探讨维持线粒体稳态的时机窗，在受伤后5分钟、3小时、6小时或24小时给予FCCP的最佳剂量，并使用几个线粒体功能参数作为结果指标。结果表明，TBI后使用解偶联剂靶向线粒体功能障碍存在延长的时机窗，并深入了解了与TBI相关的细胞病理学。
