13C and 15N MRS Study of glutamate control in epilepsy

描述（由申请人提供）：谷氨酸兴奋性毒性与癫痫发作有关。我们的目标是：1）阐明控制完整哺乳动物脑中谷氨酸和谷氨酰胺代谢通量和室间转运的机制; 2）确定谷氨酸兴奋性毒性如何引起癫痫发作。细胞外液中GLU的神经胶质摄取受损（GLUECF）可导致GLU受体的过度刺激和神经元变性（GLU兴奋性毒性）。神经递质GLU从细胞外液摄取到星形胶质细胞中并随后代谢成GLN的速率已经通过以下方式测量：a）通过同位素追踪选择性13 C富集神经递质GLU，B）通过微透析收集细胞外GLU并通过气相色谱/质谱分析其13 C富集，和c）观察[5- 13 C，现在，具体目标#1，GLN/GLU循环中的“缺失环节”，将使用体内15 N确定GLN从星形胶质细胞转运到神经元的动力学和机制，1H-15 N HMQC和13 C MRS。结果将阐明最近鉴定的GLN转运蛋白在完整正常脑中的作用，并确定胶质GLN流出速率（假设#1）或神经元GLN摄取速率（假设#2）是否限制体内GLN/GLU循环速率。具体目的#2将使用慢性红藻氨酸诱导的癫痫（KA）大鼠模型研究颞叶癫痫中GLU释放和摄取的动力学。将在损伤和对侧海马体中测量体内通量率，并将其与电生理和行为癫痫发作相关联，以确定癫痫发作和传播是否是（假设#3）异常GLU释放、（假设#4）胶质转运蛋白的受损GLU清除或（假设#5）这些过程的组合的结果。这些研究将有助于更好地了解KA中GLN/GLU循环的调节事件，并增加替代颞叶癫痫手术治疗的可能性。

DESCRIPTION (provided by applicant): Glutamate excitotoxicity has been implicated in epileptic seizures. Our objectives are 1) to clarify the mechanisms that control the metabolic flux and intercompartmental transport of glutamate and glutamine in the intact mammalian brain and 2) determine how glutamate excitotoxicity may cause epileptic seizures. Impaired glial uptake of GLU in the extracellular fluid (GLUECF) can lead to excessive stimulation of the GLU receptors and to neuronal degeneration (GLU excitotoxicity). The rate of uptake of neurotransmitter GLU from the extracellular fluid into astrocytes and subsequent metabolism to GLN has been measured through a) selective 13C enrichment of neurotransmitter GLU by isotope chase, b) collection of extracellular GLU by microdialysis and analysis of its 13C enrichment by gas-chromatography/mass spectrometry and c) observation of [5-13C, 5-15N] GLN formed in astrocytes using localized in vivo 13C and 15N MRS. Now, Specific Aim #1, a 'missing-link' in the GLN/GLU cycle, the kinetics and mechanism of transport of GLN from astrocytes to neurons will be determined using in vivo 15N, 1H-15N HMQC and 13C MRS. The results will clarify the roles of recently identified GLN transporters in the intact normal brain and determine whether the rate of glial GLN efflux (hypothesis #1) or of neuronal GLN uptake (hypothesis #2) limits the GLN/GLU cycle rate in vivo. Specific Aim #2 Kinetics of GLU release and uptake in temporal lobe epilepsy will be studied using the chronic kainate-induced epileptic (KA) rats model. In vivo flux rates will be measured in the lesioned and contralateral hippocampus and correlated with electrophysiological and behavioral seizures, to determine whether the onset and propogation of seizures is the result of (hypothesis #3) abnormal GLU release, (hypothesis #4) impaired GLU clearance by glial transporters, or (hypothesis #5) a combination of these processes. Together these studies will contribute to a better understanding of the regulatory events of the GLN/GLU cycle in KA and increase potential to replace surgical management of temporal lobe epilepsy.