GLUTAMATE TRANSPORT AND TRANSPORTERS FOREBRIAN NEURONS

描述：这项研究的长期目标是更好地了解 谷氨酸转运在兴奋性生理和病理生理学中的作用 突触。尽管谷氨酸转运在很大程度上被认为是一种 星形胶质细胞的功能，来自各种方法的证据表明 神经元谷氨酸摄取机制也很重要，对于 兴奋性传递的生理学以及精神分裂症的病理生理学 兴奋性毒性。我们研究了大鼠前脑神经元中谷氨酸的运输。 培养，发现谷氨酸转运的药理学是不同的 与神经元转运蛋白EAAC1的同源性，并与已知的GLT1相似 作为星形胶质细胞的运输者。我们克隆了谷氨酸转运蛋白，形成了一个cDNA 用这些文化建造的图书馆。我们发现，除了EAAC1和GLT1之外，还有一个 GLT1的变种形式，在C末端有11个氨基酸不同于 最初克隆的GLT1。我们现在将最初的形式称为“GLT1a”，而 变异形式克隆自神经元库“GLT1b”。初步的LM和EM 免疫细胞化学研究表明，GLT1b定位于神经元 以及前脑中的星形胶质细胞。此外，GLT1b而不是GLT1a具有 PDZ-结构域相互作用C-末端的一致序列。假说 推动这个项目的是GLT1b是一种重要的神经元转运蛋白，并且 这种变异形式通过它的C末端与其他 具有重要功能但不能被GLT1a利用的蛋白质。这个 该项目的具体目标是： 1.比较GLT1a和GLT1b在神经元培养和脑内的表达。 假设是GLT1b的表达解释了广泛存在的 GLT1基因在大脑神经元中的表达。 2.鉴定在酵母双杂交筛选中发现的与GLT1b的相互作用。这个 假设GLT1b与一个或多个PDZ结构域相互作用，包含 蛋白质。 3.研究蛋白质相互作用对GLT1a&b功能的影响。 假设GLT1的变异形式的功能会受到影响 通过与其他蛋白质的相互作用。

DESCRIPTION: The long-term goal of this research is to better understand the role of glutamate transport in the physiology and pathophysiology of excitatory synapses. Although glutamate transport has been assumed to be largely a function of astrocytes, evidence from a variety of approaches suggests that neuronal glutamate uptake mechanisms are important as well, both for the physiology of excitatory transmission as well as for the pathophysiology of excitotoxicity. We studied glutamate transport in rat forebrain neurons in culture, and found that the pharmacology of glutamate transport was distinct from that of the neuronal transporter EAAC1, and resembled that of GLT1, known as an astrocyte transporter. We cloned glutamate transporters form a cDNA library made from these cultures. We found, in addition to EAAC1 and GLT1, a variant form of GLT1 that differs at the C-terminal 11 amino acids from the originally cloned GLT1. We now call the originally form "GLT1a," and the variant form cloned from the neuronal library "GLT1b." Preliminary LM and EM immunocytochemical studies have shown that GLT1b is localized to neurons as well as astrocytes in the forebrain. In addition, GLT1b, but not GLT1a, has a PDZ-domain interaction consensus sequence at its C-terminal. The hypothesis motivating this project is that GLT1b is an important neuronal transporter, and that this variant form has interactions through its C-terminal with other proteins that are functionally important and not available to GLT1a. The specific aims of this project are to: 1. Compare the expression of GLT1a & b in neuronal cultures and in the brain. The hypothesis is that GLT1b expression accounts for the widespread presence of mRNA for GLT1 in neurons in the brain. 2. Characterize interactors found in a yeast two-hybrid screen with GLT1b. The hypothesis is that GLT1b interacts with one or more PDZ domain containing proteins. 3. Characterize the effect of protein interactors on the function of GLT1a & b. The hypothesis is that the function of variant forms of GLT1 will be influenced by interactions with other proteins.