Estrogen Receptor Gene Transfer in Mouse Brains

小鼠大脑中雌激素受体基因转移

• 批准号: 6805792
• 负责人: SONOKO OGAWA
• 金额: $ 8.43万
• 依托单位: ROCKEFELLER UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-09-30 至 2006-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6805792
• 关键词: adeno associated virus group; aggression; behavioral genetics; estrogen receptors; gene expression; gene targeting; gene therapy; genetic regulation; genetic transduction; genetically modified animals; hormone regulation /control mechanism; in situ hybridization; laboratory mouse; microinjections; neuroendocrine system; oxytocin; polymerase chain reaction; receptor expression; tranquilizer; transfection /expression vector

DESCRIPTION (provided by applicant): The Research objective of this proposal is to test whether brain site-specific gene transduction with the use of adeno-associated virus (AAV) can focally restore expression and functions of a disrupted gene in knockout mouse brains. We have been investigating the differential roles of two types of estrogen receptors (ERs), alpha, and beta, in estrogenic regulation of neuroendocrine and behavioral functions, using knockout mice for ER-alpha (alphaRKO) and ER-beta (betaERKO) genes. Knockout mice are great tools to study specific gene function in many traits and correlate them all, but have the limitation that targeted genes are disrupted globally and permanently. Here, we propose to re-introduce a disrupted gene to knockout mice in a specific brain region depending on the endpoint of specific studies. Our preliminary studies demonstrated that we could site-specifically re-introduce a disrupted gene, ER-alpha or ER-beta, to respective knockout mouse brains. It was also found that transduced ER gene was functional, since progesterone receptor protein, the most well characterized down stream product of ligand-dependent transcriptional activity of ER, was induced by estrogen in the transduced cells. In the proposed studies, we will test the functional consequences of site-specific AAV mediated gene transfer in adult knockout mouse brains. Specifically, we will re-introduce ER-6 gene by injecting AAV vectors containing ER-beta Cdna (AAV.ER-beta) into the hypothalamic paraventricular nucleus of betaERKO male mice and determine whether disrupted neuroendocine and behavioral functions can be restored. In the first part of the proposed studies, the effects on estrogen-inducible up-regulation of oxytocin gene expression will be examined with in situ hybridization and quantitative PCR assays (Aim I). Given a positive outcome, behavioral effects of ER-13 gene transduction will be further examined in terms of ER-beta mediated anxiolytic action of estrogen as well as estrogen-inducible aggression in male betaERKO mice (Aim II). The outcome of the proposed studies will provide valuable information about mechanisms of action of two types of ERs in the brain.

描述（由申请人提供）：本提案的研究目的是测试使用腺相关病毒（AAV）进行脑部位特异性基因转导是否可以局部恢复敲除小鼠大脑中被破坏基因的表达和功能。我们利用敲除er - α (alphaRKO)和er - β (betaERKO)基因的小鼠，研究了两种类型的雌激素受体（er）， α和β在雌激素调节神经内分泌和行为功能中的差异作用。基因敲除小鼠是研究许多性状中特定基因功能及其相互关联的重要工具，但其局限性在于目标基因在全局和永久性地被破坏。在这里，我们建议根据特定研究的终点，在特定大脑区域重新引入被破坏的基因。我们的初步研究表明，我们可以定位特异性地将被破坏的er - α或er - β基因重新引入到各自的敲除小鼠大脑中。我们还发现，转导的内质网基因是功能性的，因为内质网配体依赖性转录活性的下游产物黄体酮受体蛋白在转导的细胞中被雌激素诱导。在拟议的研究中，我们将测试位点特异性AAV介导的基因转移在成年敲除小鼠大脑中的功能后果。具体而言，我们将通过将含有er - β Cdna的AAV载体（AAV. er - β）注射到betaERKO雄性小鼠下丘脑室旁核中重新引入ER-6基因，并观察是否可以恢复被破坏的神经内分泌和行为功能。在第一部分的研究中，我们将通过原位杂交和定量PCR检测雌激素诱导的催产素基因表达上调的影响（Aim I）。鉴于阳性结果，ER-13基因转导的行为效应将进一步研究er - β介导的雌激素的抗焦虑作用以及雌性诱导的雄性betaERKO小鼠的攻击行为（Aim II）。提出的研究结果将为两种类型的内质网在大脑中的作用机制提供有价值的信息。