Regulation of Gene Expression in Striatal Neurons

纹状体神经元基因表达的调控

• 批准号: 6819072
• 负责人: QIANG WANG
• 金额: $ 29.77万
• 依托单位: UNIVERSITY OF MISSOURI KANSAS CITY
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-04-01 至 2009-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6819072
• 关键词: antisense nucleic acid; biological signal transduction; cAMP response element binding protein; calcium ion; calmodulin dependent protein kinase; corpus striatum; endogenous opioid; gel mobility shift assay; gene induction /repression; genetically modified animals; glutamate receptor; immunocytochemistry; in situ hybridization; laboratory mouse; laboratory rat; mitogen activated protein kinase; neural plasticity; neurogenetics; neuropharmacology; phosphorylation; western blottings

DESCRIPTION (provided by applicant): Receptor-mediated gene expression in matured CNS neurons remains a hot topic in elucidating molecular mechanisms for the development of neuroplasticity. As a key structure of the basal ganglia, the striatum is among the major central sites for investigating such gene expression aimed to unravel striatal mechanisms underlying a variety of mental illnesses (psychiatric and cognitive disorders). Metabotropic glutamate receptors (mGluRs) are densely expressed in the striatal projection neurons. From studies conducted in the last period, we have established that Ca2+ -Iinked group I mGluRs play a profound role in regulating opioid peptide gene expression (prodynorphin and proenkephalin) in the rat striatum. In the effort of identifying intracellular signaling pathways transducing group I mGluR signals to the gene expression, we recently found that activation of group I receptors activates mitogen-activated protein kinases (MAPKs). As an information superhighway between the surface membrane and the nucleus, the MAPK pathway therefore likely couples group I mGluRs to gene expression. In this continuation proposal, a series of experiments was proposed to evaluate an overarching hypothesis that group I mGluRs activate MAPK cascades to facilitate opioid gene expression in striatal neurons. Using multidisciplinary approaches, this hypothesis will be tested both in vivo and in vitro n striata neurons in four general experiments: (1) define the role of group I receptors in the regulation of MAPKs by characterizing group I I mGluR-regulated phosphorylation of a major MAPK subclass, extracellular signal-regulated kinase 1 and 2 (ERKI/2), in a I well-characterized in vivo rat model, (2) differentiate the relative importance of the two group I subtypes, mGluR1 and I mGluR5, in this event using the subtype-selective agonists/antagonists, antisense oligos and mutant mice (mGluR1 or 5l knockouts) in vivo, (3) identify signaling pathways transducing group I mGluR signals to MAPK/ERK by evaluating roles of Ca 2+ signals and protein kinases (CaMKII, PI3-kinase, PKC, and tyrosine kinases) in our primary striatal neuronal culture model with selective inhibitors, antisense oligos and silencer siRNAs, and (4) define physiological roles of group I mGluRactivated MAPK/ERK cascades in the regulation of striatal gene expression via evaluating the importance of the two transcription factor targets of MAPK/ERK, Elk-1 and CREB, for group I/ERK-sensitive prodynorphin and proenkephalin gene expression in vivo and in vitro. Accomplishment of this project will provide a new interpretation of receptor and signal transduction mechanisms underlying inducible gene expression in matured CNS neurons. Since inducible gene expression is conceived to be an important component of the development of neuroplasticity, data from this project can ultimately contribute to the development of novel pharmacotherapies, by targeting group ImGluRs and MAPK cascades, for the treatment of various mental illnesses resulted from striatal dysfunctions.

描述（由申请人提供）： 成熟中枢神经系统神经元中受体介导的基因表达仍然是阐明神经可塑性发展分子机制的热门话题。作为基底神经节的关键结构，纹状体是研究此类基因表达的主要中心部位之一，旨在揭示各种精神疾病（精神疾病和认知障碍）背后的纹状体机制。代谢型谷氨酸受体（mGluR）在纹状体投射神经元中密集表达。根据上一时期进行的研究，我们已经确定 Ca2+ 连接的 I 组 mGluR 在调节大鼠纹状体中阿片肽基因（强啡肽原和脑啡肽原）的表达中发挥着深远的作用。在鉴定将 I 类 mGluR 信号转导至基因表达的细胞内信号通路的过程中，我们最近发现 I 类受体的激活会激活丝裂原激活蛋白激酶 (MAPK)。作为表面膜和细胞核之间的信息高速公路，MAPK 途径可能将 I 组 mGluR 与基因表达结合起来。在这个延续提案中，提出了一系列实验来评估一个总体假设，即 I 组 mGluR 激活 MAPK 级联以促进纹状体神经元中阿片类药物基因的表达。使用多学科方法，该假设将在四个一般实验中在体内和体外对纹状体神经元进行测试：（1）通过表征主要 MAPK 亚类、细胞外信号调节激酶 1 和 2 (ERKI/2) 的 I 组 I mGluR 调节磷酸化，定义 I 组受体在 MAPK 调节中的作用，在 I 中已充分表征。 体内大鼠模型，(2) 区分两种 I 组亚型 mGluR1 和 I mGluR5 的相对重要性，在这种情况下使用体内亚型选择性激动剂/拮抗剂、反义寡核苷酸和突变小鼠（mGluR1 或 5l 敲除），(3) 通过评估作用来确定将 I 组 mGluR 信号转导至 MAPK/ERK 的信号传导途径 Ca 2+ 信号和蛋白激酶（CaMKII、PI3 激酶、PKC 和酪氨酸激酶）在我们的原代纹状体神经元培养模型中的选择性抑制剂、反义寡核苷酸和沉默子 siRNA 的影响，以及 (4) 通过评估重要性来定义 I 组 mGluR 激活的 MAPK/ERK 级联在纹状体基因表达调节中的生理作用 MAPK/ERK、Elk-1 和 CREB 的两个转录因子靶标，用于体内和体外 I/ERK 敏感的强啡肽原和脑啡肽原基因表达。该项目的完成将为成熟中枢神经系统神经元诱导基因表达的受体和信号转导机制提供新的解释。由于诱导基因表达被认为是神经可塑性发展的重要组成部分，因此该项目的数据最终可以通过靶向 ImGluR 和 MAPK 级联组来促进新型药物疗法的开发，用于治疗纹状体功能障碍引起的各种精神疾病。