CNS L-PROLINE TRANSPORTER

CNS L-脯氨酸转运蛋白

• 批准号: 2270737
• 负责人: ROBERT T FREMEAU
• 金额: $ 16.46万
• 依托单位: DUKE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1993
• 资助国家: 美国
• 起止时间: 1993-12-15 至 1996-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2270737
• 关键词: Xenopus oocyte; aminoacid transport; biological signal transduction; cell cell interaction; chemical binding; chimeric proteins; gene induction /repression; glutamate receptor; glutamates; hippocampus; laboratory mouse; laboratory rabbit; laboratory rat; membrane transport proteins; molecular cloning; molecular site; neural transmission; neuroanatomy; neuronal transport; proline; protein structure function; second messengers; site directed mutagenesis; synapses; tissue /cell culture

L-proline has been postulated to be a neurotransmitter or neuromodulator in the mammalian central nervous system (CNS). Studies to define the role of L-proline in discrete pathways in the mammalian CNS have not been possible due to the inability to block its biosynthesis or high affinity transport in nervous tissue. Our recent cloning of a high affinity, Na+- dependent L-proline transporter expressed in subpopulations of putative glutamatergic neurons provides compelling support for a specific role for L-proline in excitatory transmission. Glutamatergic synapses have been implicated in diverse physiologic processes including the acquisition of associative learning, synapse formation, and neuroendocrine regulation. Abnormalities of glutamatergic transmission have been implicated in the pathophysiology of excitotoxic neurologic disorders. Thus an understanding of the functional role(s) of the L-proline transporter in specific glutamatergic neurons could provide insights into synaptic regulatory mechanisms involved in synaptic plasticity and memory acquisition or their disruption in diseases such as Alzheimer's disease. The long term goal of this research is to elucidate the role(s) of L-proline, and its transporter, in synaptic transmission. Specific Aim l will undertake the first detailed studies of the regional, cellular, and subcellular distribution of the high affinity L-proline transporter protein in rat brain. The precise projection pathways of individual neurons that express L-proline transporter mRNA will be determined by combining retrograde dye tract-tracing with in situ hybridization histochemistry. Knowledge of these pathways will be crucial to the design of physiological studies of the role of L-proline in synaptic transmission. Studies will also examine the possible co-localization of high affinity L-proline and L-glutamate transporters in glutamatergic nerve terminals. Specific Aim 2 will use the L-proline transporter as a model neurotransmitter transporter, to investigate the structure-function properties and regulation of this important class of synaptic proteins using chimeric transporter strategies and site-directed mutagenesis. The final Specific Aim will investigate potential synaptic roles of L-proline. A Xenopus oocyte expression assay will be used to investigate the hypothesis that L-proline modulates signal transduction at a glutamate receptor subtype, analogous to glycine modulation of the NMDA receptor. This assay will also be used to investigate the possibility that distinct L-proline receptors exist in mammalian CNS. Monoclonal antibodies that selectively block L-proline transport will be generated, for use as transport inhibitors, to examine the hypothesis that L-proline modulates the synthesis and/or release of glutamate from nerve terminals. The potential discovery of novel regulatory mechanisms in glutamatergic synapses is the incentive which drives the research described in this proposal.

L-脯氨酸被认为是一种神经递质或神经调质 哺乳动物中枢神经系统（CNS）研究确定的作用 L-脯氨酸在哺乳动物中枢神经系统中的离散途径尚未得到证实。 可能是由于不能阻断其生物合成或高亲和力 在神经组织中运输。我们最近克隆了一种高亲和力的Na+- 依赖性L-脯氨酸转运蛋白在假定的 多巴胺能神经元为神经元的特定作用提供了令人信服的支持， L-脯氨酸在兴奋性传递中的作用谷氨酸能突触 参与各种生理过程，包括获得 联想学习、突触形成和神经内分泌调节。 在过去的几年里， 兴奋毒性神经系统疾病的病理生理学。因此， L-脯氨酸转运蛋白在特定的 突触能神经元可以提供突触调节的见解 参与突触可塑性和记忆获得的机制或其 阿尔茨海默氏症等疾病的破坏。的长期目标 本研究旨在阐明L-脯氨酸的作用， 转运蛋白，在突触传递中。具体目标我将承担 第一次详细研究了区域，细胞和亚细胞 高亲和力L-脯氨酸转运蛋白在大鼠体内的分布 个脑袋单个神经元表达的精确投射途径 L-脯氨酸转运蛋白mRNA将通过结合逆行染料 原位杂交组织化学示踪。知识 这些途径将是至关重要的生理研究的设计， L-脯氨酸在突触传递中的作用研究还将审查 高亲和力L-脯氨酸和L-谷氨酸可能共定位 转运蛋白的作用。 具体目标2将使用 L-脯氨酸转运蛋白作为模型神经递质转运蛋白， 研究其结构-功能特性及其调控 使用嵌合转运蛋白策略的重要类别的突触蛋白 和定点诱变。最终的具体目标将调查 L-脯氨酸的潜在突触作用。非洲爪蟾卵母细胞表达试验 将被用来调查的假设，L-脯氨酸调节信号 谷氨酸受体亚型的转导，类似于甘氨酸 NMDA受体的调节。该试验还将用于 研究不同的L-脯氨酸受体存在于 哺乳动物中枢神经系统选择性阻断L-脯氨酸的单克隆抗体 将产生转运，用作转运抑制剂，以检查 假设L-脯氨酸调节的合成和/或释放 神经末梢中的谷氨酸小说的潜在发现 调节机制的突触是诱因， 推动了本提案中所述的研究。