REGULATION OF BRAIN NEUROTRANSMITTER SYNTHESIS

脑神经递质合成的调节

• 批准号: 6540087
• 负责人: SUSAN M HUTSON
• 金额: $ 33.37万
• 依托单位: WAKE FOREST UNIVERSITY HEALTH SCIENCES
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-06-05 至 2004-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6540087
• 关键词: aminoacid metabolism; anticonvulsants; brain metabolism; chemical kinetics; glutamates; glutamine; isozymes; laboratory rat; neurotransmitter metabolism; neurotransmitter transport; nuclear magnetic resonance spectroscopy; pharmacokinetics; pyruvate carboxylase; radiotracer; retina; tissue /cell culture

Glutamate is the most widely used excitatory neurotransmitter in the nervous system. Certain diseases and neurodegenerative disorders are thought to involve disturbances of the glutamatergic systems of the brain or retina. The goal of this proposal is to understand the mechanisms that regulate glutamate levels in the nervous system. Considerable evidence indicates that glial cells rapidly internalize glutamate released by neurons during neurotransmission converting it to glutamine before releasing it back to neurons in the glutamate/glutamine cycle. However, it is now clear that possibly as much as 50 percent of the glutamate taken up by the glia is actually converted to pyruvate and lactate. Therefore, the nervous system must regenerate the lost carbon by carboxylation of pyruvate catalyzed by the anaplerotic enzyme pyruvate carboxylase (PC) which is found is astroglia. Second, it now appears that rather than ammonia the essential branched chain amino acids (BCAA) are needed to provide sufficient alpha-amino nitrogen for optimal rates of de novo glutamate synthesis. Indeed, it is our hypothesis that the BCAA and branched chain aminotransferase isoenzymes (cytosolic BCATc, mitochondrial BCATm) participate in a nitrogen shuttle whereby the glial BCATm and neuron specific BCATc act in series to transfer nitrogen from neurons to astrocytes. The aims of this proposal are designed to understand the mechanisms that regulate glutamate levels in the nervous system and test the BCAA shuttle hypothesis. 1) We will develop methods to simultaneously measure the rate of de novo glutamate synthesis and the rate of glutamate glutamine cyclin in the ex vivo retina and in brain in vivo and correlate the influence of neuronal activity with flux. Both H14CO3 and 13C NMR spectroscopy will be used in vivo for the kinetic analysis. 2) The function of the BCAT isoenzymes in regulating glutamate synthesis and brain neurotransmitter pool sizes will be quantified in vivo in brain and ex vivo in the retina. The hypothesis that the neuroactive drug gabapentin acts via specific inhibition of cytosolic BCATc will be tested. 3) The degree of control exerted by the anaplerotic enzyme pyruvate carboxylase (PC) relative to that of the BCAT isoenzymes will be assessed. Antisense technology will be used to vary enzyme levels in vivo. Data will be analyzed using control strength theory to determine the relative control strength of the BCAT and PC on the pathways of de novo glutamate synthesis. Finally, understanding the mechanisms that regulate glutamate synthesis will increase our knowledge of certain disease processes and may allow for future generation of novel therapeutic compounds.

谷氨酸是神经系统中使用最广泛的兴奋性神经递质。 某些疾病和神经退行性疾病被认为涉及大脑或视网膜谷氨酸能系统的干扰。 该提案的目的是了解调节神经系统中谷氨酸水平的机制。 大量证据表明，神经元在神经传递期间释放的谷氨酸迅速内化，然后将其转化为谷氨酰胺，然后将其释放回谷氨酸/谷氨酰胺循环中的神经元。 但是，现在很明显，被神经胶质吸收的谷氨酸的50％实际上被转化为丙酮酸和乳酸。 因此，神经系统必须通过丙酮酸的羧化催化丙酮酸羧化酶羧化酶（PC）催化的碳酸盐（PC）来再生碳的损失。 其次，现在似乎需要提供基本的分支链氨基酸（BCAA）而不是氨，以提供足够的α-氨基氮，以实现从新手谷氨酸合成的最佳速率。 的确，我们的假设是BCAA和分支链氨基转移酶同工酶（胞质BCATC，线粒体BCATM）参与氮班车，从而使神经胶质BCATM和NERURON特异性BCATC在系列中以将氮从神经元转移到神经元中。 该提案的目的旨在了解调节神经系统中谷氨酸水平的机制，并测试BCAA穿梭假设。 1）我们将开发方法，以同时测量新谷氨酸合成的速率和谷氨酸谷氨酰胺细胞周期蛋白在离体视网膜中和体内大脑中的速率，并将神经元活性与磁通的影响相关。 H14CO3和13C NMR光谱都将在体内用于动力学分析。 2）BCAT同工酶在调节谷氨酸合成和脑神经递质池大小中的功能将在视网膜中的脑和Ex Vivo中进行定量。 将测试神经活性药物Gabapentin通过特异性抑制胞质BCATC起作用的假设。 3）将评估丙酮酸丙酮酸羧化酶（PC）相对于BCAT同工酶的控制程度。反义技术将用于在体内改变酶水平。将使用控制强度理论对数据进行分析，以确定BCAT和PC在从头谷氨酸合成途径上的相对控制强度。 最后，了解调节谷氨酸合成的机制将增加我们对某些疾病过程的了解，并可能允许未来的新型治疗化合物。