Function of Astrocytic GPCR Signaling Cascades in Physiology and Mental Illness

星形胶质细胞 GPCR 信号级联在生理和精神疾病中的功能

• 批准号: 8442109
• 负责人: Ken Douglas McCarthy
• 金额: $ 34.2万
• 依托单位: UNIV OF NORTH CAROLINA CHAPEL HILL
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-03-05 至 2017-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8442109
• 关键词: Affect; Agonist; Anxiety; Applications Grants; Area; Astrocytes; Autonomic nervous system; Behavior; Behavioral; Biological Models; Blood - brain barrier anatomy; Brain; Calcium; Cells; Cholinergic Receptors; Clinical; Communities; Coupled; Designer Drugs; Directed Molecular Evolution; Disease; Drug Receptors; Drug usage; Employee Strikes; Exhibits; Family; G Protein-Coupled Receptor Signaling; G-Protein-Coupled Receptors; Gene Activation; Gene Expression; Genes; Genetic; Genetic Models; Glial Fibrillary Acidic Protein; Goals; Human; Image; Intraperitoneal Injections; Knock-out; Ligands; Measures; Mental disorders; Mus; Neurons; Oxides; Pharmacogenetics; Phenotype; Physiological; Physiology; Population; Research; Role; Signal Transduction; Signaling Molecule; Sorting - Cell Movement; Specificity; Synaptic Transmission; System; Technology; Testing; Therapeutic Agents; Time; Transgenes; Transgenic Mice; behavioral impairment; brain cell; cell type; design; drugged driving; genetic inhibitor; memory retrieval; mouse model; optogenetics; promoter; public health relevance; receptor; receptor function; research study; stem; therapeutic target; tool

DESCRIPTION (provided by applicant): The goal of this grant proposal is to develop genetic tools that can be used to activate or inactivate G-protein coupled receptor (GPCR) signaling cascades in rigorously identified populations of astrocytes using approaches that can be readily used by the research community; we believe the availability of such tools will be transformational with respect to understanding the role of astrocytes in physiology, behavior, and mental illness. While astrocytes make up a large portion of the CNS and express a wide variety of GPCRs that are activated during neuronal activity, the role of astrocytic GPCRs is poorly understood. The lack of information in this area stems from our inability to selectively activate or inactivate astrocytic signaling cascades while measuring physiological and behavioral parameters. Importantly, the majority of therapeutic agents used clinically to treat mental illness effect GPCR signaling cascades. Recently, genetic tools have been developed that can be used to selectively activate or inactivate GPCR signaling cascades in specific cell types. One such pharmacogenetic approach utilizes a receptor family referred to collectively as DREADD receptors (for Designer Receptor Exclusively Activated by Designer Drug) to activate GPCR signaling cascades in specific cell types. DREADD receptors were prepared by directed molecular evolution of M3-ACh receptor DREADD receptors, fail to respond to any known endogenous GPCR ligand, and are activated by clozepine-N-oxide (CNO). Importantly, CNO crosses the blood-brain-barrier enabling, for the first time, the selective activation of GPRC signaling in specific populations of astrocytes via an intraperitoneal (ip) injection of CNO. We plan to use intersectional gene activation which requires two cell- type specific promoters to activate gene expression and greatly increases the cell specificity of gene expression. Intersectional gene activation will be used to express Gq-, Gs-, and Gi-DREADD receptors in rigorously defined populations of astrocytes. A concern with any pharmacological approach is whether the resulting phenotype (e.g., LTP, behavior, etc.) reflects the over stimulation of the system under study. We plan to develop genetic tools that can be used to inactivate GPCR signaling cascades in rigorously defined populations of astrocytes; these genetic tools will also take advantage of intersectional gene activation. The combination of being able to activate or inactivate GPCR signaling cascades in rigorously defined populations of astrocytes will provide a powerful approach to sorting out the role of astrocytes in physiology, behavior, and mental illness; distribution of these mouse models to the research community is likely to be transformational. The genetic tools developed in this proposal will be used to test the hypothesis that the activation of astrocytic GPCR signaling cascades modulates synaptic transmission; this hypothesis is fundamental to our understanding of the role of astrocytes in brain function and is currently extremely controversial within the research community.

描述（由申请人提供）：该赠款提案的目的是开发可用于激活或灭活G蛋白偶联受体（GPCR）信号级联的遗传工具，使用严格识别的星形胶质细胞群体使用研究社区可以轻松使用的星形胶质细胞群体；我们认为，此类工具的可用性将在了解星形胶质细胞在生理，行为和精神疾病中的作用方面具有变革性。尽管星形胶质细胞占中枢神经系统的很大一部分，并表达了在神经元活性期间被激活的多种GPCR，但对星形胶质细胞GPCR的作用知之甚少。该领域缺乏信息源于我们无法在测量生理和行为参数的同时选择性地激活或灭活星形胶质细胞信号级联。重要的是，大多数治疗剂在临床上用于治疗精神疾病效应GPCR信号级联。最近，已经开发出可用于在特定细胞类型中选择性激活或灭活GPCR信号级联的遗传工具。一种这样的药物遗传学方法利用一种被称为Dreadd受体的受体家族（对于设计器药物专门激活的设计器受体）来激活特定细胞类型的GPCR信号级联。 Dreadd受体是通过M3-ACh受体Dreadd受体的定向分子演化制备的，无法对任何已知的内源性GPCR配体反应，并被氯脂激素-N-氧化物（CNO）激活。重要的是，CNO首次通过腹膜内注射（IP）注射CNO的特定星形胶质细胞中GPRC信号的选择性激活GPRC信号传导。我们计划使用截面基因激活，该基因激活需要两个细胞类型的特异性启动子激活基因表达并大大提高基因表达的细胞特异性。截面基因激活将用于在严格定义的星形胶质细胞种群中表达GQ-，GS-和GI-DreadD受体。任何药理学方法的关注点是所产生的表型（例如LTP，行为等）是否反映了正在研究的系统的过度刺激。我们计划开发可用于在严格定义的星形胶质细胞种群中灭活GPCR信号级联的遗传工具；这些遗传工具还将利用交叉基因激活。在严格定义的星形胶质细胞种群中激活或灭活GPCR信号级联的结合将提供一种有力的方法来整理星形胶质细胞在生理，行为和精神疾病中的作用；这些小鼠模型向研究界的分布可能是变换的。该提案中开发的遗传工具将用于检验以下假设：星形胶质细胞GPCR信号级联反应调节突触传播的假设；该假设是我们对星形胶质细胞在大脑功能中的作用的理解至关重要的，目前在研究界非常有争议。