Differential Terminal Expression of GAD67/GAD65 ? Relevance to Schizophrenia

GAD67/GAD65 的差异末端表达？

• 批准号: 7569786
• 负责人: KENNETH N FISH
• 金额: $ 15.15万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-04-10 至 2011-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7569786
• 关键词: Acute; Affect; Anabolism; Anatomy; Antibodies; Autopsy; Axon; Brain region; Calcium-Binding Proteins; Cannabinoids; Cell Nucleus; Cells; Cholecystokinin; Code; Confocal Microscopy; Corpus striatum structure; Data; Dendrites; Dependency; Detection; Enzymes; Fire - disasters; Fluorescence; Future; Genes; Hippocampus (Brain); Human; Impairment; Interneurons; Knowledge; Label; Lead; Link; Membrane Transport Proteins; Memory impairment; Messenger RNA; Methodology; Monkeys; Muscarinic M2 Receptor; Neurons; Parvalbumins; Pattern; Pharmacological Treatment; Physiological; Prefrontal Cortex; Presynaptic Terminals; Primates; Probability; Process; Property; Protein Isoforms; Proteins; Pyramidal Cells; Regulation; Relative (related person); Reporting; Schizophrenia; Short-Term Memory; Site; Staining method; Stains; Structure; Surface; Synapses; System; Techniques; Testing; Thalamic structure; Tissues; Transcript; Translations; cell type; cognitive function; density; design; flexibility; fluorescence imaging; gamma-Aminobutyric Acid; human tissue; immunocytochemistry; improved; insight; mRNA Expression; neuronal cell body; neurotransmission; nonhuman primate; novel; postsynaptic; presynaptic; public health relevance; receptor; response; trafficking

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.

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的数量。末端GABA由GAD67和GAD65蛋白在局部合成。GABA的主要合成酶GAD67转录物的表达减少可能是精神分裂症中最常见的病理障碍。GAD67 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的振荡损伤。然而，GAD67 mRNA的缺失是否伴随着GAD67蛋白的相应减少，特别是在主要功能位点（如末端），目前尚不清楚。此外，中间神经元，特别是那些GAD67 mRNA减少被认为在精神分裂症中发生的中间神经元，对GAD67和GAD65在GABA末端合成方面的依赖性尚不清楚。因此，我们开发了一种新的高通量荧光成像方法，可以精确定量荧光标记的点（假定的末端），在同一末端的不同标记的共定位，以及在这些相同结构中的荧光强度的定量。使用这种方法，我们将进行研究，首次比较非人类和人类（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）中与精神分裂症相关的不同中间神经元末端的GAD67和GAD65蛋白水平。在完成拟议的研究后，我们将知道：1)在非人灵长类动物DLPFC中，终端GAD67/GAD65比率是否存在中间神经元亚群特异性差异；2)观察到的非人类和人类灵长类之间的差异是否保守；3)精神分裂症患者DLPFC GABA末端亚群中GAD67是否减少。获得的知识将使我们能够：1)制定关于精神分裂症中GAD67 mRNA减少的细胞类型特异性后果的假设；2)在非人类灵长类动物丰富的生理、药理学和解剖学数据背景下解释研究结果；3)为优化未来人体死后组织研究设计提供必要的数据。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会导致某些认知功能的损害，而这些功能是该疾病的核心特征。在很大程度上，目前精神分裂症的药物治疗在改善认知功能方面是无效的。这些研究将提供关于GABA神经传递的关键细胞、潜在的药理学靶点以及它们如何在精神分裂症中受到影响的急需信息。