Mechanisms for Extracellular Glutamate Formation in Brai

据推测，间隙空间中的谷氨酸水平维持在低水平以防止兴奋性毒性。这是至关重要的，因为发育中的大脑由于兴奋性氨基酸受体的上调而增加了对兴奋性毒性的敏感性。围产期缺氧/缺血时细胞外谷氨酸升高仅增加约3倍，但却是死亡率和发病率的主要原因。幸存者往往留下严重的，永久性的神经损伤。然而，我们假设，低水平的谷氨酸是必要的间隙空间。已显示谷氨酸盐的基线水平与代谢型受体相互作用。该项目将研究细胞外谷氨酸在正常和创伤性围产期脑缺氧/缺血发作后在围产期和成人脑中产生的机制。初步结果表明，向大鼠和小鼠输注谷氨酰胺 通过微透析对脑内谷氨酸的清除显著增加了细胞外谷氨酸的浓度至兴奋性毒性水平，并且其在细胞外空间中由谷氨酰胺合成。脑中细胞外谷氨酰胺的浓度通常为200 - 300 μ M，提供现成的前体。我们假设，在正常条件下，细胞外谷氨酸合成的主要机制是马来酸活化的谷氨酸氨基转移酶（MAG），一种胞外酶γ-谷氨酰转肽酶（γ-GTP）的副反应。我们假设第二种机制在创伤中占主导地位。我们认为线粒体内磷酸依赖性转氨酶 (PDG)从受损细胞释放（或暴露）并将谷氨酰胺转化为谷氨酸，导致在初始创伤事件后数天的兴奋性毒性和细胞死亡。这些酶也可能在氨升高等条件下发挥作用。我们将确定在正常和7天大的缺氧缺血性大鼠脑，成年大鼠氨中毒的条件下，在酶γ-GTP缺乏的小鼠，和神经元细胞培养细胞外谷氨酸形成的酶机制。研究将利用微透析和细胞培养在存在和不存在的效应物的酶参与。预计这些研究将为婴儿期缺氧或缺血性损伤的护理提供新的方法。

It has been postulated that glutamate levels in the interstitial space are maintained at low levels to prevent excitotoxicity. This is critical since the developing brain has increased sensitivity to excitotoxicity due to up regulation of excitatory amino acid receptors. Elevated extracellular glutamate in perinatal hypoxia/ischemia increases only about 3 fold, yet is a major cause of mortality and morbidity. Survivors are often left with severe, permanent neurological impairment. However, we postulate that a low level of glutamate is necessary in the interstitial space. Baseline levels of glutamate have been shown to interact with metabotropic receptors. This project will examine mechanisms by which extracellular glutamate is generated in both perinatal and adult brain in the normal and in the traumatized perinatal brain following an episode of hypoxia/ischemia. Preliminary results indicate that infusion of glutamine into the rat and mouse brain by microdialysis markedly increases the concentration of extracellular glutamate to excitotoxic levels and that it is synthesized in the extracellular space from glutamine. The concentration of extracellular glutamine in brain is normally 200 -300 mu M providing a ready precursor. We hypothesize that under normal conditions the primary mechanism for extracellular glutamate synthesis is maleate activated glutaminase (MAG), a side reaction of the ectoenzyme gamma-glutamyl transpeptidase (gamma-GTP). We hypothesize a second mechanism predominates in trauma. We propose that intramitochondrial phosphate dependent glutaminase (PDG) is released (or exposed) from damaged cells and converts glutamine to glutamate resulting in excitotoxicity and cell death days after the initial traumatic event. These enzymes may also play a role in conditions such as elevated ammonia. We will determine the enzymatic mechanism for extracellular glutamate formation in normal and 7-day old hypoxic-ischemic rat brain, adult rats under conditions of ammonia toxicity, mice deficient in the enzyme gamma-GTP, and in neuronal cell cultures. Studies will utilize microdialysis and cell culture in the presence and absence of effectors of the enzymes involved. It is anticipated that these studies will lead to new approaches in the care of infants that have experienced hypoxic or ischemic injury during infancy.