Exploring nucleocytoplasmic IEG mRNA export in striatal neuron subpopulations

探索纹状体神经元亚群中核细胞质 IEG mRNA 输出

• 批准号: 9005845
• 负责人: KRISTEN A KEEFE
• 金额: $ 18.44万
• 依托单位: UNIVERSITY OF UTAH
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-04-01 至 2018-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9005845
• 关键词: Adult; Affect; American; Antibodies; Basal Ganglia; Behavior; Behavioral Paradigm; Brain; Cell Nucleus; Cell physiology; Centrifugation; Complex; Confocal Microscopy; Corpus striatum structure; Coupled; Cytoplasm; Data; Detection; Development; Dopamine; Dopamine D1 Receptor; Dopamine D2 Receptor; Dorsal; Drug Addiction; Drug Exposure; Drug abuse; Efferent Neurons; Extinction (Psychology); Flow Cytometry; Fluorescent in Situ Hybridization; Genes; Goals; Growth; Health; Immediate-Early Genes; Immunoprecipitation; Individual; Label; Literature; Mediating; Mediator of activation protein; Memory; Messenger RNA; Molecular; Mus; Neurons; Neurotransmitters; Nuclear; Nuclear Export; Outcome; Pathway interactions; Pharmaceutical Preparations; Population; Process; Protein Export Pathway; Proteins; Receptor Activation; Recording of previous events; Regulation; Response to stimulus physiology; Reverse Transcriptase Polymerase Chain Reaction; Ribonucleoproteins; Role; Seminal; Sorting - Cell Movement; Synaptic plasticity; System; Testing; Transcriptional Activation; Transcriptional Regulation; Translations; Treatment outcome; brain circuitry; drug seeking behavior; improved; in vivo; insight; long term memory; mRNA Export; mRNA Expression; new therapeutic target; novel; novel therapeutics; promoter; response; trafficking

 DESCRIPTION (provided by applicant): As of 2012, approximately 22 million Americans required treatment for drug dependence. Drug addiction arises as a consequence of changes in brain circuitry induced by drug exposure and resultant dopamine release. Synaptic plasticity processes underlie numerous aspects of drug abuse and addiction, including the formation of drug-clue, action-outcome, and stimulus-response associations affecting instrumental behavior, as well as extinction of drug-seeking behavior. Synaptic plasticity is heavily dependent on the expression and function of immediate early genes (IEGs), in particular activity-regulated, cytoskeletal-associated (Arc) and early growth response-1 (egr1/zif268) genes. These genes critically mediate consolidation and reconsolidation of long-term memories, including memories implicated in aspects of drug abuse and addiction. Understanding the regulation of these IEGs is therefore significant in its potential to identify novel therapeutic targets to disrupt aberrant plastic changes contributing to drug abuse and addiction. Many studies have focused on understanding processes mediating transcriptional regulation of IEGs, as well as on dendritic trafficking and local translation of Arc mRNA. However, to date there are no studies in the extant literature examining the basic cellular process of nuclear export of mRNAs in general, much less IEG RNAs in particular, in the adult mammalian brain. Our experimental evidence suggests that nucleocytoplasmic export of these IEG mRNAs differs in striatonigral (direct pathway) vs. striatopallidal (indirect pathway) neurons in dorsal striatum. Further, our recent data suggest that the nuclear export of IEG mRNAs in striatonigral efferent neurons may be regulated by dopamine. The overall goal of the proposed studies is thus to test the novel hypothesis that there are differences in nucleocytoplasmic export of IEG mRNAs in striatal efferent neuron subpopulations. In Specific Aim 1, we will determine the phenotypic distribution of known mediators of nucleocytoplasmic mRNA export and their co-localization with IEG mRNAs in striatal efferent neuron subpopulations. In Specific Aim 2, we will differential centrifugation and FACS to separate nuclei from striatonigral and striatopallidal neurons, followed by ribonucleoprotein immunoprecipitation and RT-PCR to biochemically assess the interaction of Arc and zif268 with specific ribonucleoprotein complexes involved in nuclear export. In Specific Aim 3, we will determine the role of dopamine D1 receptor activation in the regulation of nuclear export of IEGs in striatonigral efferent neurons. Given the lack of data regarding the regulation of nuclear export of mRNAs in neurons of the adult mammalian brain and potential regulation of this process by neurotransmitter systems, including DA, successful completion of the proposed studies will provide seminal insight into the regulation of these critical plasticity-related IEGs n striatal circuitry intricately involved in drug abuse and addiction. These findings thus have the potential to reveal novel therapeutic targets to modulate striatal plasticity so as to improve treatment outcomes in individuals with histories of drug abuse and addiction.

 描述（由适用提供）：截至2012年，大约2200万美国人需要治疗药物依赖。药物成瘾是由于药物暴露引起的脑回路变化而产生的，并导致多巴胺释放。突触可塑性过程是药物滥用和成瘾的许多方面的基础，包括影响工具行为的药物凝结，动作结果和刺激 - 反应关联，以及灭绝药物的行为。突触可塑性在很大程度上取决于早期基因（IEG）的表达和功能，特别是活动调节，细胞骨架相关（ARC）和早期生长响应1（EGR1/ZIF268）基因。这些基因对培养基的巩固和长期记忆的重新整合，包括在药物滥用和成瘾方面隐含的记忆。因此，了解这些IEG的调节对于确定新颖的治疗靶标而破坏异常的可能性很重要 塑料变化导致滥用药物和成瘾。许多研究的重点是理解介导IEG转录调控的过程，以及树突贩运和弧mRNA的局部翻译。但是，迄今为止，在现有文献中还没有研究MRNA的核输出基本细胞过程，尤其是IEG RNA，尤其是在成年哺乳动物大脑中。我们的实验证据表明，这些IEG mRNA的核细胞质出口在纹状体（直接途径）与纹状体（间接途径）神经元中的纹状体（直接途径）中有所不同。此外，我们最近的数据表明，纹状体有效神经元中IEG mRNA的核出口可能受多巴胺调节。因此，提出的研究的总体目标是检验新的假设，即在纹状体有效的神经元亚群中，IEG mRNA的核酸性胞质出口存在差异。在特定的目标1中，我们将确定已知的核细胞质mRNA导出介质的表型分布及其与IEG mRNA在纹状体有效神经元亚群中的共定位。在特定目标2中，我们将差异离心和 FACS将核与纹状体和纹状体有效神经元分开，然后是核糖核蛋白免疫沉淀和RT-PCR，以生化评估ARC和ZIF268与参与核Export的特定核糖核蛋白复合物的相互作用。在特定的目标3中，我们将确定多巴胺D1受体激活在纹状体有效神经元中IEG的调节IEG的调节中的作用。鉴于缺乏有关成年哺乳动物大脑神经元核输出mRNA的调节的数据，以及神经递质系统对这一过程的潜在调节，包括DA，成功完成拟议的研究将为这些关键可变性IEG的调节提供第二次洞察，以使这些关键相关的IEG与药物滥用无关紧要。因此，这些发现有可能揭示新的治疗靶标，以调节纹状体可塑性，从而改善药物滥用和成瘾史的个体的治疗结果。