Differential Terminal Expression of GAD67/GAD65 ? Relevance to Schizophrenia

描述（由申请人提供）：GABA神经传递的突触前强度部分取决于可释放的末端GABA的量。末端GABA由GAD 67和GAD 65蛋白局部合成。GAD 67（GABA的主要合成酶）转录本表达减少可能是精神分裂症中最常见的病理性障碍。GAD 67 mRNA表达的减少可能导致GABA合成减少，导致GABA神经传递减弱，从而导致精神分裂症中的振荡损伤。然而，目前尚不清楚GAD 67 mRNA的缺陷是否伴随着GAD 67蛋白的类似减少，特别是在主要功能位点（例如末端）。此外，依赖于中间神经元，特别是那些GAD 67 mRNA的减少，提出发生在精神分裂症，对GAD 67与GAD 65的末端GABA合成是未知的。因此，我们已经开发了一种新的高通量荧光成像方法，允许准确定量的荧光标记的斑点（推定的终端），在同一终端的不同标签的共定位，并在这些相同的结构中的荧光强度的定量。使用这种方法，我们将进行研究，这将是第一个比较GAD 67和GAD 65蛋白的水平在不同的中间神经元的终端，与精神分裂症在非人类和人类（精神分裂症和匹配的对照组）灵长类动物背外侧前额叶皮层（DLPFC）。在完成拟定研究时，我们将了解：1）在非人灵长类动物DLPFC中是否存在终末GAD 67/GAD 65比值的中间神经元亚群特异性差异; 2）在非人和人灵长类动物之间观察到的差异是否保守; 3）在精神分裂症受试者DLPFC中GABA终末亚群中GAD 67是否减少。这些获得的知识将使我们能够：1）制定关于精神分裂症中GAD 67 mRNA减少的细胞类型特异性后果的假设; 2）在非人灵长类动物存在的丰富的生理学，药理学和解剖学数据的背景下解释发现; 3）提供必要的数据，以优化未来死后人体组织研究的设计。公共卫生相关性：与精神分裂症相关的GABA神经传递缺陷被认为有助于某些认知功能的损害，这些认知功能是疾病的核心特征。在大多数情况下，目前精神分裂症的药物治疗在改善认知功能方面无效。这些研究将提供有关GABA神经传递所涉及的关键细胞、潜在药理学靶点以及它们在精神分裂症中如何受到影响的急需信息。

DESCRIPTION (provided by applicant): The presynaptic strength of GABA neurotransmission is partially determined by the amount of terminal GABA available for release. Terminal GABA is synthesized locally by GAD67 and GAD65 protein. Reduced expression of the transcript for GAD67, the principal synthesizing enzyme for GABA, is perhaps the most replicated pathological disturbance in schizophrenia. Reductions in GAD67 mRNA expression may lead to reduced GABA synthesis, resulting in weaker GABA neurotransmission, and hence oscillatory impairment, in schizophrenia. However, it is unknown if the deficit in GAD67 mRNA is accompanied by a comparable decrease in GAD67 protein, particularly at the major site of function (e.g. the terminal). In addition, the dependency of interneurons, particularly those in which reductions of GAD67 mRNA are proposed to occur in schizophrenia, on GAD67 versus GAD65 for terminal GABA synthesis is unknown. Therefore, we have developed a novel high-throughput fluorescence imaging methodology that allows for the accurate quantification of fluorescently-labeled puncta (putative terminals), the colocalization of different labels in the same terminal, and the quantification of fluorescence intensity in these same structures. Using this methodology, we will perform studies that will be the first to compare the level of GAD67 and GAD65 protein in the terminals of different interneurons that are relevant to schizophrenia in the non-human and human (schizophrenia and matched control subjects) primate dorsolateral prefrontal cortex (DLPFC). At the completion of the proposed studies we will know: 1) if interneuron subpopulation specific differences in terminal GAD67/GAD65 ratios are present in the non-human primate DLPFC; 2) whether observed differences are conserved between non-human and human primates; 3) if GAD67 is reduced in subpopulations of GABA terminals in the DLPFC of subjects with schizophrenia. This acquired knowledge will allow us to: 1) formulate hypotheses about the cell type specific consequences of the GAD67 mRNA reduction in schizophrenia; 2) interpret findings in the context of the rich body of physiologic, pharmacologic, and anatomic data that exists for non-human primate; 3) provide data necessary to optimize the design of future studies in postmortem human tissue. PUBLIC HEALTH RELEVANCE: Deficits in GABA neurotransmission associated with schizophrenia are believed to contribute to the impairments in certain cognitive functions that are core features of the illness. For the most part, current pharmacological treatments for schizophrenia are ineffective at improving cognitive function. These studies will provide much needed information about the key cells, potential pharmacological targets, involved in GABA neurotransmission and how they are affected in schizophrenia.