CONTROL OF GABAERGIC NEURON DIFFERENTIATION

哺乳动物的神经系统由大量的神经元组成。 类型。有理由认为这个曲目是由一个 基因控制的复杂网络。然而，到目前为止，人们对此知之甚少 关于这个网络。我们建议分析一个非常基本的发展 神经元做出的选择：是兴奋性还是抑制性。这个 决定抑制性通常涉及选择性地表达 谷氨酸脱羧酶(GAD)基因我们建议分析 导致GAD67基因在正常小鼠体内表达的机制 哺乳动物大脑中的神经元。这应该会让我们洞察到更大的 基因组如何指导如此多的独立生物的发育 神经元类型。初步数据有力地表明，有几个 GAD67基因转录起始点。DNA周围的片段 转录起始点将被克隆并导入细胞 并用于创造转基因小鼠的品系。这样我们就会 确定给出特定细胞类型所需的最小DNA单位 GAD67基因的表达。一旦确定了这一点，删除分析 将执行，以确定GAD67内及其邻近的监管地点 吉恩。对这些地点的位置和顺序的了解将使 有可能系统地寻找控制 GAD基因的表达。可诱导小鼠P19畸胎瘤细胞 分化为神经元样和神经胶质样细胞 维甲酸。诱导后GAD67 RNA转录本水平升高 戏剧性的。然而，细胞的功能特性是 表达GAD的基因还不清楚。这些细胞产生功能的能力 GAD酶和GABA能突触前终末将被研究。自.以来 有两个基因编码GAD，了解它们的亲缘关系是很重要的 每一个人的贡献。作为朝着这个方向迈出的一步，我们将创建老鼠 其中GAD67基因的两个等位基因都被打断。到时候我们会的 检测功能性GAD67基因缺失对血管紧张素转换酶活性的影响 发展。综上所述，这些研究将为 导致脑神经元的一个主要子集出现的机制。 这项工作将与由或涉及到的人类疾病相关 大脑正常发育和基因表达的紊乱。最近的GAD 已被证明是青少年糖尿病的主要自身抗原， 僵硬男人综合症。这项工作将提供有关以下方面的附加基本数据 一组与疾病相关的重要基因。

The mammalian nervous system is comprised of a vast repertoire of neuronal types. Its reasonable to assume that this repertoire is generated by a complex network of genetic control. However, to date very little is known about this network. We propose to analyze a very fundamental developmental choice made by neurons: whether to be excitatory or inhibitory. The decision to be inhibitory usually involves the selective expression of the genes for glutamic acid decarboxylase (GAD). We propose to analyze the mechanisms that lead to the expression of the GAD67 gene in the correct neurons of the mammalian brain. This should give insight into the larger problem of how the genome can guide the development of so many separate neuronal types. Preliminary data strongly suggests that there are several transcriptional start sites for the GAD67 gene. Segments of DNA surroundin the transcriptional start sites will be cloned and transfected into cell lines and used to create lines of transgenic mice. In this way we will determine the minimal unit of DNA necessary to give cell type specific expression of the GAD67 gene. Once this is determined, deletion analysis will be performed to locate regulatory sites in and adjacent to the GAD67 gene. Knowledge of the location and sequence of these sites will make it possible to systematically search for trans-acting factors that control the expression of the GAD gene. P19 mouse teratocarcinoma cells can be induced to differentiate into neuronal and glial like cells by treatment with retinoic acid. After induction levels of GAD67 RNA transcripts rise dramatically. However the functional properties of the cells which are expressing GAD are not known. The ability of these cells to make functiona GAD enzyme and GABAergic presynaptic terminals will be investigated. Since there are 2 genes coding for GAD it will be important to know the relative contribution of each one. As a step in that direction we will create mice in which both alleles of the GAD67 gene are interrupted. We will then determine the effect of eliminating the functional GAD67 gene on development. Taken together these studies will provide basic information o mechanisms that lead to the emergence of a major subset of brain neurons. The work will have relevance to human diseases that are caused by or involv disturbances of normal brain development and gene expression. Recently GAD has been shown to be a dominant autoantigen in juvenile diabetes and the stiff man syndrome. This work will provide additional basic data about thi important disease-related set of genes.