NEUROENDO SIGNAL TRANSDUCTION--REPRODUCTIVE PHYSIOLOGY

神经内信号转导——生殖生理学

• 批准号: 2742894
• 负责人: KAREN Ann GREGERSON
• 金额: $ 27.07万
• 依托单位: UNIVERSITY OF MARYLAND BALTIMORE
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-01-01 至 2002-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2742894
• 关键词: Xenopus oocyte; animal puberty; biological signal transduction; calcium ion; dopamine; estrus; female; fertility; gene expression; gene targeting; genetic models; genetically modified animals; hormone regulation /control mechanism; immunocytochemistry; laboratory mouse; lactation; membrane potentials; model design /development; neuroendocrine system; pituitary gland; potassium channel; prolactin; pseudopregnancy; secretion; voltage /patch clamp

Prolactin (PRL), an anterior pituitary hormone, has profound effects on female fertility. In fact, hypersecretion of PRL is the major neuroendocrine-related pathology associated with female infertility. Unfortunately, the underlying defects which lead to the hypersecretion of PRL remain ill-defined. To provide a rational framework for understanding these defects and treating associated pathologies, it is essential to elucidate the cellular and molecular mechanisms of PRL secretion regulation. Normally PRL secretion is under tonic inhibition by hypothalamic dopamine (DA). Periodic, physiological surges of PRL in females require a withdrawal from this dopaminergic inhibition. Since reduced responsiveness to DA is a hallmark of hyperprolactinemic syndromes, our attention has focused on the cellular and molecular basis of DA action on the lactotrope. We have discovered and extensively characterized a DA-activated inwardly-rectifying potassium channel (KDA) in normal lactotropes. KDA activation by DA leads to hyperpolarization of the lactotrope membrane and cessation of calcium-dependent action potentials, the driving force for tonic PRL secretion. In vitro studies have demonstrated a critical role for this KDA channel in the regulation PRL release by DA. Furthermore, the functional expression of KDA is dependent upon estrogen, which may explain a long-recognized modulatory action of estrogen on this system. In these regards, the KDA channel lies at the top of a hierarchy of events in dopaminergic regulation of PRL secretion. As an initial step in elucidating the molecular basis for the physiological role of KDA in lactotropes, we have cloned three different K channel gene products from female anterior pituitary tissue. Based on functional similarities between the native channel and recombinant channels expressed in Xenopus oocytes, two of the gene products are excellent candidate subunits encoding a heteromultimeric KDA. As a logical extension of these studies, we propose to evaluate the role of this effector K channel within its physiological context- the whole animal-through transgenic technology. To this end, we will design and construct "dominant-negative" mutants, capable of inhibiting wild-type KDA function in lactotropes. We will then create a transgenic mouse model in which expression of this mutant is directed to pituitary lactotropes using the PRL promoter. These studies represent a timely and important extension of the P.I.'s work and will ultimately provide insight into the basis of disorders in PRL secretion and their impact on female fertility.

催乳素（PRL）是一种前垂体激素，对 女性生育能力。 实际上，PRL的过度分泌是主要的 与女性不育症有关的神经内分泌相关病理。 不幸的是，导致过度分泌的潜在缺陷 PRL的定义不明。 提供一个理性的框架 了解这些缺陷并治疗相关的病理，这是 阐明PRL的细胞和分子机制必不可少的 分泌法规。 通常PRL分泌受补品抑制作用 通过下丘脑多巴胺（DA）。 PRL的周期性生理激增 在女性中，需要退出这种多巴胺能抑制。 由于降低对DA的响应能力是多丙泊汀的标志 综合征，我们的注意力集中在细胞和分子基础上 在乳酸上作用的作用。 我们发现并广泛地表征了DA激活 正常乳糖中的内部切除钾通道（KDA）。 KDA DA激活导致乳糖膜超极化 并停止钙依赖性作用电位，驱动力 用于补品PRL分泌。 体外研究表明 DA的调节PRL释放中，该KDA通道的角色。 此外，KDA的功能表达取决于 雌激素可以解释一种长期认可的调节作用 该系统上的雌激素。 在这些方面，KDA频道位于 PRL多巴胺能调节中事件的层次结构的顶部 分泌。 作为阐明分子基础的第一步 KDA在乳糖中的生理作用，我们克隆了三种不同的 k通道基因产物来自雌性前垂体组织。 基于 关于天然通道和重组之间的功能相似性 在爪蟾卵母细胞中表达的通道，两个基因产物是 出色的候选亚基编码异类KDA。 作为 这些研究的逻辑扩展，我们建议评估 该效应子k渠道在其生理环境中 - 整体 动物直接转基因技术。 为此，我们将设计和 构建“显性阴性”突变体，能够抑制野生型 乳糖中的KDA功能。 然后，我们将创建一个转基因鼠标 该突变体表达的模型被引向垂体 使用PRL启动子的乳糖。 这些研究代表了及时 以及P.I.工作的重要扩展，最终将提供 深入了解PRL分泌中疾病及其影响的基础 关于女性生育能力。