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

描述(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)阐明在完整的哺乳动物大脑中控制谷氨酸和谷氨酰胺代谢流量和脑室间转运的机制，以及2)确定谷氨酸兴奋毒性如何导致癫痫发作。细胞外液中GLU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从细胞外液到星形胶质细胞的摄取和随后对GLN的代谢的速率已经通过a)同位素追逐选择性地浓缩神经递质GLU，b)微透析收集细胞外GLU并用气相色谱/质谱仪分析其13C浓缩，以及c)利用体内13C和15N MRS观察星形胶质细胞中形成的[5-13C，5-15N]GLN。现在，专门针对#1，在GLN/GLU循环中的一个‘缺失环节’，将利用体内15N来确定GLN从星形胶质细胞到神经元的运输的动力学和机制，1H-15N HMQC和13C MRS这些结果将阐明最近发现的谷氨酰胺转运体在完整的正常脑中的作用，并确定胶质细胞谷氨酰胺外流(假说1)或神经元谷氨酰胺摄取(假说2)的速率是否限制了体内的谷氨酰胺/谷氨酸周期速率。具体目的#2利用慢性海人酸致癫痫(KA)大鼠模型研究颞叶癫痫GLU释放和摄取的动力学。在活体中，将测量受损和对侧海马区的通量速率，并将其与电生理和行为癫痫相关联，以确定癫痫的开始和传播是(假设3)GLU异常释放的结果，(假设4)胶质转运体对GLU的清除受损，还是(假设5)这些过程的组合。总之，这些研究将有助于更好地了解KA中Gln/GLU周期的调节事件，并增加取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.