非损伤微测技术用于神经毒性机制的研究 谷氨酸毒性研究：单个神经元O2消耗量、细胞内Ca2+浓度和线粒体膜电位的同时记录 神经元是可兴奋性细胞，在突触活动及产生动作电位后，神经元要产生大量的ATP驱动离子泵以提高胞内的Na+和Ca2+水平。谷氨酸（Glu）是重要的神经递质，负责快速突触传递及突触传递强度的长期变化，并参与认知和记忆等过程；但如果过度激活谷氨酸受体，谷氨酸会导致离子平衡破坏、细胞死亡等毒性反应。 本文为明确谷氨酸神经毒性的机制，将非损伤微测技术与激光共聚焦技术结合，以大鼠幼崽大脑皮层的神经元为被测样品，用非损伤微测技术（a self-referencing O2 electrode）检测单个神经元O2消耗量（即O2内流），而用激光共聚焦技术检测其胞内Ca2+浓度和线粒体膜电位。 单个神经元细胞在Glu作用下O2消耗量、胞内Ca2+浓度 和线粒体膜电位的变化过程 研究发现，在谷氨酸作用下，细胞内Ca2+浓度上升，随后O2消耗量增加，这期间线粒体膜电位也发生相应改变。该结论直接证实了下述谷氨酸毒性机理模型：谷氨酸受体被激活后能引起Ca2+内流，导致细胞内ATP损耗。 将非损伤微测技术与激光共聚焦等技术相结合，检测生物样品内部和外部离子分子或其他信息的变化情况，已经成为揭示生命过程机理机制的重要手段。 关键词：非损伤微测技术(a self-referencing O2 electrode), 氧气消耗量(oxygen consumption), 谷氨酸(glutamate); 参考文献：Gleichmann M, et al. J Neurochem, 2009,109: 644-655 全文下载：http://www.xuyue.net/xylt/viewthread.php?tid=535&extra=page%3D1

Abstract：
In order to determine the sequence of cellular processes in glutamate toxicity, we simultaneously recorded O₂ consumption,cytosolic Ca²⁺ concentration ([Ca²⁺]i), and mitochondrial membrane potential (mDw) in single cortical neurons. Oxygen consumption was measured using an amperometric selfreferencing platinum electrode adjacent to neurons in which [Ca²⁺]i and mDw were monitored with Fluo-4 and TMRE+,respectively, using a spinning disk laser confocal microscope.Excitotoxic doses of glutamate caused an elevation of [Ca²⁺]i followed seconds afterwards by an increase in O₂ consumption which reached a maximum level within 1–5 min. A modest increase in mDw occurred during this time period, and then, shortly before maximalO₂ consumption was reached, themDw, as indicated by TMRE+ fluorescence, dissipated. Maximal O₂ consumption lasted up to 5 min and then declined together with mDw and ATP levels, while [Ca²⁺]i further increased. mDw and [Ca²⁺]i returned to baseline levels when neurons were treated
with an NMDA receptor antagonist shortly after the [Ca²⁺]i increased. Our unprecedented spatial and time resolution revealed that this sequence of events is identical in all neurons, albeit with considerable variability in magnitude and kinetics of changes in O₂ consumption, [Ca²⁺]i, and mDw. The data obtained using this new method are consistent with a model where Ca²⁺ influx causes ATP depletion, despite maximal mitochondrial respiration, minutes after glutamate receptor activation.
Keywords: excitotoxicity, glutamate, oxygen consumption.