EXCITATORY TRANSMISSION MODULATION BY GLUTAMATE UPTAKE

钠依赖性的谷氨酸高亲和力摄取被认为是 在突触活动的快速终止中起主要作用， 维持非常低的细胞外兴奋性氨基浓度 中枢神经系统中的EAA。然而，很少有研究 直接或系统地解决了高亲和力谷氨酸的作用， 兴奋性突触传递中的摄取。这可能部分是由于 缺乏选择性和有效的抑制剂钠依赖性高， 亲和谷氨酸转运系统。最近开发的选择性， 高效摄取抑制剂L-反式-吡咯烷-2，4-二羧酸酯（L- trans-PDC）将用于所提议的实验中以检验假设 高亲和力谷氨酸转运系统在 调节培养海马的兴奋性突触传递 神经元EAA神经传递参与快速信息传递 传递、突触可塑性和神经毒性的研究使得 调节正常兴奋性突触传递， 与理解与气候变化相关的 神经病理学，如帕金森氏病和阿尔茨海默氏病， 衰老 电生理方法将被用来阐明的影响， 高亲和力谷氨酸转运蛋白的药理学抑制 L-trans-PDC对培养的大鼠海马神经元自发性和诱发性突触事件的影响 海马神经元微小兴奋性突触后电流 反映了神经递质的自发量子释放， 突触前末梢全细胞膜片钳记录方法允许 这些电流将以足够高的分辨率进行解析， 分析其频率、振幅分布和时间 课程检查mEPSC的这些参数将提供有用的 突触后EAA敏感性的任何变化的信息 受体和/或抑制引起的突触前释放的变化 高亲和力的谷氨酸摄取。诱发兴奋性突触后电流 （EPSC）也可以通过记录分离的成对的 单突触耦合海马神经元维持在非常低的水平， 密度培养。各种突触前刺激范式，包括单一 刺激、成对脉冲刺激和高频强直电刺激 刺激将用于确定谷氨酸摄取的影响 抑制剂L-trans-PDC对EPSC的NMDA和非NMDA组分的影响。的 假设高亲和力谷氨酸转运抑制将 导致环境中谷氨酸盐浓度的增加， 培养海马神经元细胞外间隙;实验 旨在确定高水平或长时间的 暴露于谷氨酸对非NMDA受体脱敏，时间 的过程和幅度较慢的NMDA成分的EPSC，和 突触前代谢型受体的参与。

Sodium-dependent high-affinity uptake of glutamate is thought to play a major role in the rapid termination of synaptic activity and the maintenance of very low extracellular concentrations of excitatory amino acids (EAA) in the central nervous system. However, few studies have directly or systematically addressed the role of high-affinity glutamate uptake in excitatory synaptic transmission. This may be due, in part, to the lack of selective and potent inhibitors of the sodium-dependent high- affinity glutamate transport system. The recently developed selective, highly potent uptake inhibitor L-trans-pyrrolidine-2,4-dicarboxylate (L- trans-PDC) will be used in the proposed experiments to test the hypothesis that the high-affinity glutamate transport system plays a major role in modulating excitatory synaptic transmission in cultured hippocampal neurons. The involvement of EAA neurotransmission in rapid information transfer, synaptic plasticity and neurotoxicity make the study of the modulation of normal excitatory synaptic transmission particularly relevant for understanding mechanisms underlying changes associated with neuropathologies such as Parkinson's and Alzheimer's diseases and brain aging. Electrophysiologic approaches will be used to elucidate the effects of pharmacological inhibition of the high-affinity glutamate transporter by L-trans-PDC on both spontaneous and evoked synaptic events in cultured hippocampal neurons. Miniature excitatory postsynaptic currents (mEPSCs) reflect the spontaneous quantal release of neurotransmitter at single presynaptic terminals. Whole-cell patch clamp recording methods allow these currents to be resolved with sufficiently high resolution to allow analysis of their frequencies, distribution of amplitudes and time courses. Examining these parameters of the mEPSCs will provide useful information of any changes in the sensitivity of the postsynaptic EAA receptors and/or changes in presynaptic release resulting from inhibition of high-affinity glutamate uptake. Evoked excitatory postsynaptic currents (EPSCs) may also be examined by recording from isolated pairs of monosynaptically coupled hippocampal neurons maintained in very low density cultures. Various presynaptic stimulus paradigms including single stimulation, paired-pulse stimulation and high-frequency tetanic stimulation will be used to determine the effects of the glutamate uptake inhibitor L-trans-PDC on NMDA and non-NMDA components of the EPSC. The hypothesis is that inhibition of high-affinity glutamate transport will result in an increase in the ambient concentration of glutamate in the extracellular space of the cultured hippocampal neurons; the experiments are designed to determine the effects of the elevated levels or prolonged exposure to glutamate on non-NMDA receptor desensitization, the time course and amplitude of the slower NMDA component of the EPSC , and the involvement of presynaptic metabotropic receptors.