KNOCK-OUT OF K+ CHANNEL IN PITUITARY--PROLACTIN CONTROL AND FEMALE FERTILITY

垂体K通道的敲除--催乳素控制与女性生育力

• 批准号: 6318365
• 负责人: KAREN Ann GREGERSON
• 金额: $ 13.96万
• 依托单位: UNIVERSITY OF MARYLAND BALTIMORE
• 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 2001-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6318365
• 关键词: Xenopus oocyte; cooperative study; dopamine; estrogens; female; female reproductive system; fertility; gene mutation; gene targeting; genetically modified animals; hormone regulation /control mechanism; immunocytochemistry; laboratory mouse; luteinizing hormone; molecular cloning; pituitary gland; potassium channel; prolactin; reproduction; 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 (K/DA) in normal lactotropes. K/DA 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 D/DA channel in the regulation of PRL release by DA. Furthermore, the functional expression of D/DA is dependent upon estrogen, which may explain a long-recognized modulatory action of estrogen on this system. In these regards, the D/DA 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 K/DA 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 K/DA. 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 K/DA 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仍然不确定。提供理性的理解框架 这些缺陷和治疗相关的病理，必须 阐明PRL分泌的细胞和分子机制 规定。通常，PRL分泌受到滋补的抑制作用 下丘脑多巴胺（DA）。 PRL的周期性生理激增 女性需要退出这种多巴胺能抑制。自从 对DA的响应能力降低是多丙甲术的标志 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。 我们已经发现并广泛地表征了向内的DA激活 正常乳糖中的钾通道（K/DA）。 K/DA激活 通过DA导致乳糖膜的超极化和戒烟 钙依赖性作用电位，这是补品PRL的驱动力 分泌。体外研究证明了这一点至关重要 Da对PRL释放的调节中的D/DA通道。此外， D/DA的功能表达取决于雌激素，这可能 解释雌激素对该系统的长期认可的调节作用。在 在这些方面，D/DA频道位于事件的层次结构的顶部 在PRL分泌的多巴胺能调节中。 作为阐明分子基础的第一步 K/DA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 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k/da。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 - 整个动物 - 通过转基因技术。为此，我们将设计和构建 “显性阴性”突变体，能够抑制野生型K/DA功能 在乳糖中。然后，我们将创建一个转基因鼠标模型 该突变体的表达使用使用 PRL启动子。这些研究代表了及时，重要的扩展 P.I.的工作，最终将提供有关的洞察力 PRL分泌的疾病及其对女性生育的影响。