MODULATION OF SYNAPTIC EFFICACY & MECHANISMS OF EPILEPSY

描述：（申请人的摘要）本补助金拟审查 兴奋性和抑制性的短期频率依赖性可塑性 突触功能，目的是帮助理解 癫痫患者在癫痫发作过程中发生的事件 病灶和癫痫发作从异常病灶区域向 整个皮质 核心假设是， 抑制性和兴奋性突触的结合负责 以及癫痫活动在大脑皮层的扩散 据了解，根据 在大多数情况下，重复激活会导致抑制减弱， 和兴奋转化为潜能。 为了分析这些过程， 在细胞水平上，开发了一种简化的制备方法， 细胞膜片钳记录突触对中的两个神经元， 应用定量分析来帮助确定机制 潜在的突触可塑性 在海马培养物中，突触 抑制随着重复刺激而显著减少， 由于突触前因素。 兴奋性突触后电流也 在重复动作过程中，也通过突触前机制， 而自发释放的递质却矛盾地被促进。 即使在基线、非刺激条件下， 在抑制性和非抑制性之间的神经递质释放机制中， 和兴奋性神经元，这些可能在差异中发挥作用， 可塑性。 所有观察到的突触可塑性形式都是 依赖于突触末梢中Ca的变化， 假设必须涉及几个钙依赖性过程。 的 钙依赖性的细节将被表征，机制通过 终末内钙差的变化影响 兴奋性和抑制性末端的神经递质将被 测定 短期塑性也将在更成熟的 海马切片培养物中的组织。 在这种制备中，作为切片， 成熟时，兴奋性突触通过重复激活而增强。 使用与细胞培养系统中使用的技术类似的技术， 的增强和发展的机制， 将分析增强机制。 最后，使用新的 该技术允许从单个细胞中定量扩增mRNA， 细胞，确定的兴奋和抑制的mRNA表达谱 神经元，无论是在细胞培养和切片培养将是 考察 不同的配位分子表达， 这两种表型将被分析，重点是突触蛋白， 钙通道，钙依赖性酶，是神经分泌的一部分， 机制和其他钙结合蛋白。 的分子基础 不同形式的短期可塑性， 将确定抑制性神经元的表现。 它是假设 癫痫发作发生在异常兴奋区域， 兴奋性和抑制性神经元的差异调节， 癫痫通过类似的机制扩散到整个大脑。 希望 对这些细胞可塑性本质的进一步理解 两种不同的细胞类型将导致策略的发展， 抑制癫痫发作发展和扩散。

DESCRIPTION: (Applicant's Abstract) This grant proposes to examine short term, frequency-dependent plasticity of excitatory and inhibitory synaptic function with the goal of contributing to an understanding of events which occur during the development of seizures in an epileptic focus and the spread of seizures from a focal area of abnormality to the entire cortex. The central hypothesis is that differential regualtion of inhibitory and excitatory synapses is responsible for the development and spread of seizure activity in the cortex. It is known that under most circumstances, repetitive activation causes inhibition to diminish and excitation to potential. In order to analyze such processes at the cellular level, a simplified preparation was developed to allow whole cell patch clamp recording of both neurons in a synaptic pair and the application of quantal analysis to help determine the mechanisms underlying synaptic plasticity. In the hippocampal cultures, synaptic inhibition profoundly decrements with repeated stimulation and this is due to presynaptic factors. Excitatory postsynaptic currents also decrement during repetitive action, also via a presynaptic mechanisms, whereas spontaneous transmitter released is paradoxically facilitated. Even under baseline, non-stimulated conditions, there are differences in the mechanisms underlying neurotransmitter release between inhibitory and excitatory neurons, and these may play a role in the differences in plasticity. All of the observed forms of synaptic plasticity are dependent on changes in Ca in the synaptic terminals and it is hypothesized that several Ca-dependent processes must be involved. The details of the Ca dependence will be characterized and the mechanisms by which changes in intraterminal Ca differential affect the release of neurotransmitter at excitatory and inhibitory terminals will be determined. Short term plasticity will also be studied in more mature tissue in hippocampal slice cultures. In this preparation, as the slice matures, excitatory synapse potentiate with repetitive activation. Using similar techniques to those employed in the cell culture system, the mechanisms underlying the potentiation and the development of the potentiating mechanisms will be analyzed. Finally, using a new technique which allows quantitative amplification of mRNAs from single cells, mRNA expression profiles of identified excitatory and inhibitory neurons, both in the cell culture ant the slice culture will be examined. The differences in the coordinated molecular expression in these two phenotypes will be analyzed, focusing on synaptic proteins, Ca channels, Ca-dependent enzymes which are part of the neurosecretory mechanisms and other Ca binding proteins. The molecular bases of the different forms of short term plasticity which the excitatory and inhibitory neurons exhibit will be determined. It is hypothesized that seizure develop in areas of abnormal excitability because of a differential regulation of excitatory and inhibitory neurons and that seizures spread throughout the brain by similar mechanisms. It is hoped that an increased understand of the nature of this plasticity in these two distinct cell types will lead to the development of strategies to suppress seizure development and spread.