NEURONS AND GLIA IN VITRO

世纪前，高尔基提出神经胶质细胞为神经元提供营养。当发现星形胶质细胞而不是神经元含有糖原（大脑中主要的能量储备）时，这一想法得到了证实。最近对蜜蜂和兔子视网膜的观察提供了现代实验证据，证明神经胶质细胞可以为神经元提供燃料。然而，糖原的作用仍然是一个谜。本申请中提出的实验将关键性地解决糖原在脑能量代谢期间神经胶质-神经元相互作用中的作用。这些实验将使用中枢神经系统（CNS）白色物质的有利制备物（分离的大鼠视神经）进行。本研究的长期目标是：1）了解更多关于神经胶质细胞的生理学和功能，和2）了解脑卒中、低血糖、缺氧或创伤引起的CNS白色物质损伤的机制，并设计更好的方法来减少这种损伤。概括地说，这项建议的具体目标是了解神经胶质细胞和轴突在中枢神经系统中的相互作用时，剥夺葡萄糖和了解如何葡萄糖剥夺损伤中枢轴突。将检验两个假设：1）在CNS中的低血糖期间，星形胶质细胞以源自糖原的乳酸盐的形式向轴突供应能量底物。轴突的功能和存活在没有葡萄糖的情况下依赖于糖原。2)葡萄糖剥夺引起的轴突损伤是Ca 2+依赖性的，并且是由于通过反向Na+/Ca 2+交换和Ca通道介导的Ca 2+内流。将使用体外大鼠视神经制备、监测视神经功能的定量电生理技术、糖原含量的化学测量以及pH、葡萄糖敏感和乳酸敏感微电极来研究星形胶质细胞在向轴突供应能量底物中的作用。将使用视神经功能的定量测量结合使用Ca 2+敏感微电极测量[Ca 2 + ]o来研究低血糖诱导的轴突损伤的机制。这些研究将提供有关神经胶质细胞和神经元在脑能量代谢背景下相互作用的方式的有用的新信息。他们还将评估星形胶质细胞糖原对低血糖期间和低血糖后恢复神经功能的重要性，并可能提出新的策略来避免这些条件下的神经损伤。

More than a century ago, Golgi suggested that glial cells provide nutrients to neurons. This idea gained credibility when astrocytes, but not neurons, were discovered to contain glycogen, the main energy reserve in the brain. Recent observations on the retinas of the honeybee and rabbit, have provided modem experimental evidence that glial cells can provide fuel to neurons. The role of glycogen, however, remains a mystery. The experiments proposed in this application will critically address the role of glycogen in glial- neuronal interactions during brain energy metabolism. These experiments will be carried out using an advantageous preparation of central nervous system (CNS) white matter, the isolated rat optic nerve. The long term goals of this research are: 1) to learn more about the physiology and function of glial cells, and 2) to understand the mechanisms of CNS white matter injury as occurs with stroke, hypoglycemia, anoxia or trauma, and to devise better ways of minimizing this injury. Broadly stated, the specific aims of this proposal are to learn how glial cells and axons in the CNS interact when deprived of glucose and to understand how glucose deprivation injures central axons. Two hypotheses will be tested: 1) During hypoglycemia in the CNS, astrocytes supply energy substrate to axons in the form of lactate derived from glycogen. Axon function and survival depend on glycogen in the absence of glucose. 2) Axon injury caused by glucose deprivation is Ca2+- dependent and is due to Ca2+ entry mediated by reverse Na+/Ca2+ exchange and Ca channels. The role of astrocytes in supplying energy substrate to axons will be studied using the in vitro rat optic nerve preparation, quantitative electrophysiological techniques to monitor optic nerve function, chemical measurement of glycogen content, and pH, glucose-sensitive, and lactate sensitive microelectrodes. The mechanisms of hypoglycemia-induced axon injury will be studied using quantitative measures of optic nerve function in conjunction with measurement of [Ca2+ ]o using Ca2+-sensitive microelectrodes. These studies will provide useful new information about the manner in which glial cells and neurons interact in the context of brain energy metabolism. They will also assess the importance of astrocytic glycogen for neural function during and for recovery after periods of hypoglycemia, and may suggest novel strategies to stave off neural injury under these conditions.