Mechanisms for Extracellular Glutamate Formation in Brai

据推测，间质中的谷氨酸水平维持在较低水平，以防止兴奋性毒性。这一点至关重要，因为由于兴奋性氨基酸受体的上调，发育中的大脑对兴奋性毒性的敏感性增加。细胞外谷氨酸的升高在围产期缺氧/缺血中仅增加约3倍，但却是死亡和发病率的主要原因。幸存者往往会留下严重的、永久性的神经损伤。然而，我们假设在间质间隙中需要低水平的谷氨酸。谷氨酸的基线水平已被证明与代谢性受体相互作用。该项目将研究正常的围产期和成人脑以及缺氧/缺血后受损的围产期脑中产生细胞外谷氨酸的机制。初步结果表明，谷氨酰胺注入大鼠和小鼠体内 大脑通过微透析显着增加细胞外谷氨酸的浓度，达到兴奋毒性水平，它是在细胞外空间由谷氨酰胺合成的。脑细胞外谷氨酰胺的浓度通常为200-300微米，提供了一种现成的前体。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激活的谷氨酰胺酶(MAG)，这是胞外酶γ-谷氨酰转肽酶(伽马-GTP)的副反应。我们假设第二种机制在创伤中起主导作用。我们认为线粒体内磷酸依赖的谷氨酰胺酶 (PDG)从受损的细胞中释放(或暴露)，将谷氨酰胺转化为谷氨酸，导致兴奋毒性和细胞死亡。这些酶也可能在氨浓度升高等情况下发挥作用。我们将在正常和7日龄的缺氧缺血大鼠脑中、氨中毒条件下的成年大鼠、伽马-GTP酶缺陷的小鼠以及神经细胞培养中确定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，在存在和不存在相关酶的效应物的情况下。预计这些研究将导致在婴儿期经历缺氧或缺血损伤的婴儿的护理方面的新方法。

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.