MICRODIALYSIS OF NAA/NAAG IN FOCAL EPILEPTOGENESIS

NAA/NAAG 在局灶性癫痫发生中的微透析

• 批准号: 6187584
• 负责人: JOHN M SLOPIS
• 金额: $ 10.85万
• 依托单位: UNIVERSITY OF TEXAS HLTH SCI CTR HOUSTON
• 依托单位国家: 美国
• 财政年份: 1996
• 资助国家: 美国
• 起止时间: 1996-04-01 至 2002-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6187584
• 关键词: anticonvulsants; aspartate; enzyme activity; enzyme inhibitors; extracellular; gas chromatography mass spectrometry; glutamates; high performance liquid chromatography; homeostasis; kindling; laboratory rat; microdialysis; neurons; neurotransmitters; partial seizure; pathologic process; peptidyl dipeptidase; valproate

Epilepsy is a major public health problem with estimated incidence of 30.9 to 56.8 per 100,000. The mechanism of epileptogenesis and actions of anticonvulsant drugs are poorly understood. We propose to study the amino acids N-acetyl-aspartate (NAA), N-acetyl-aspartyl-glutamate (NAAG), and the excitotoxin glutamate (GLU) in the rat kindling model of focal epileptogenesis using novel techniques. NAAG is released at some nerve terminals and cleaved by N-alpha-amino-linked-acidic-dipeptidase (NAALADase) to form NAA and GLU. Regional elevation in NAALADase may contribute to epileptogenesis by locally elevating GLU concentrations. The commonly used anticonvulsant valproic acid (VPA) is known to block kindling and to decrease brain levels of NAA, possibly by inhibition of NAALADase which would inhibit GLU production. Regional variation in NAAG synaptic release and NAALDase activity have not been studied in vivo. We developed gas chromatography mass spectrometry (GCMS) to directly quantify these amino acids following local neuronal depolarization in rat brain as sampled by microdialysis, and we demonstrated peptidase activity in situ by infusing labeled substrate (NAAG-D3) then quantifying the cleavage product NAA-D3. Our specific aims are to: (1) characterize extracellular levels of NAAG, NAA, and GLU in each region of the kindling circuit through seizure induction, (2) assess NAALADase activity in situ in each region of the kindling circuit through seizure induction to determine its potential role in homeostasis of the circuit, (3) test the hypothesis that VPA inhibits NAALADase as its anticonvulsant/antikindling mechanism of action, 4) test the hypothesis that a specific inhibitor of NAALADase, B-N-acetyl-glutamate will serve as an anticonvulsant/antikindling agent.

癫痫是一个主要的公共卫生问题，估计发病率为30.9 每10万人中有56.8人死亡。 癫痫的发生机制及药物的作用 抗惊厥药物知之甚少。 我们建议研究氨基 酸N-乙酰基-天冬氨酸（NAA）、N-乙酰基-乙酰基-谷氨酸（NAAG），以及 谷氨酸兴奋毒素在大鼠局灶性点燃模型中的作用 癫痫的治疗方法 NAAG在某些神经处释放 末端并被N-α-氨基连接的酸性二肽酶裂解 （NAALADase）以形成NAA和GLU。 NAALADase的局部升高可能 通过局部升高GLU浓度而导致癫痫发生。 的 已知常用的抗惊厥药丙戊酸（VPA）可阻断点燃 并可能通过抑制NAALADase降低脑内NAA水平 这会抑制葡萄糖的生成 NAAG突触的区域性变异 释放和NAALDase活性尚未在体内研究。 我们开发 气相色谱质谱法（GCMS），直接量化这些 在大鼠脑中局部神经元去极化后的氨基酸， 通过微透析，我们证明了肽酶活性原位， 注入标记底物（NAAG-D3），然后定量切割产物 NAA-D3 我们的具体目标是：（1）表征细胞外水平的 通过癫痫发作点燃回路各区域的NAAG、NAA和GLU 诱导，（2）原位评估NAALADase在每个区域的活性 通过癫痫发作诱导点燃回路，以确定其潜在作用 （3）验证VPA抑制VPA的假设， NAALADase作为其抗惊厥/抗点燃作用机制，4）试验 NAALADase的特异性抑制剂B-N-乙酰谷氨酸 将用作抗惊厥/抗点燃剂。