Functional and molecular identification of TREK-1 channel in myometrium in relati

子宫肌层TREK-1通道的功能及分子鉴定

• 批准号: 7843550
• 负责人: SANG Don KOH
• 金额: $ 17.56万
• 依托单位: UNIVERSITY OF NEVADA RENO
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-05-15 至 2012-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7843550
• 关键词: Action Potentials; Acute; Address; Cell membrane; Cells; Chronic; Clinical; Data; Disease; Estradiol; Estrogens; Family; Female; Hormones; Human; Image Analysis; Intervention; Investigation; Ion Channel; Isometric Exercise; Link; Measurement; Mediating; Membrane; Membrane Potentials; Methods; Microelectrodes; Modeling; Molecular; Monitor; Mus; Muscle Cells; Myometrial; Nitric Oxide; Plasma; Postpartum Period; Potassium Channel; Pregnancy; Premature Birth; Premature Labor; Progesterone; Property; Proteins; Regulation; Relaxation; Reporting; Rest; Role; Signal Transduction; Smooth Muscle; Staging; Stretching; Swelling; TRAAK channel; Techniques; Uterine Smooth Muscle Excitability; Uterus; design; insight; member; myometrium; novel; patch clamp; potassium channel protein TREK-1; pregnant; pressure; public health relevance; research study; response; shear stress; steroid hormone; therapeutic development; uterine smooth muscle cell

DESCRIPTION (provided by applicant): In most species, including humans, plasma levels of 17beta-estradiol and progesterone increase during pregnancy. The elevation of female steroid hormones that accompanies the late stages of pregnancy has been linked to electrical remodeling of myometrial smooth muscle. These observations suggest that female steroid hormones can influence the activity and/or expression of ionic conductances involved in uterine smooth muscle excitability. Although it has been reported that the delivery mechanism is related to a decrease in female hormone levels, the mechanism of labor contraction has not been clearly studied in terms of ion channel expression. During pregnancy, the mass of myometrium increases dramatically, with only minimal increases in intrauterine pressure. To accomplish this, the smooth muscle of the myometrium must remain relaxed during pregnancy. Therefore, K+ conductances activated by stretch of the plasma membrane may contribute an important component of the myogenic response in the uterus. Several K+ channels participate in the regulation of resting membrane potential and repolarization of action potentials in the myometrium. Recently unique K+ channels have been reported that consist of four transmembrane segments and two-pore domains (K2P). Among K2P channels, TREK-1, TREK-2 and TRAAK have unique functional properties and represent the first cloned stretch-activated K+ channels. Previous studies suggest that female steroid hormones influence the transcriptional expression of a number of K+ channels. We will address the following specific aims in this proposal: Aim 1, we will identify SDK channel functional expression in the murine myometrium using patch-clamp methods and will characterize the regulation of these channels by nitric oxide and its intracellular signaling mechanisms. Aim 2, we will characterize stretch-dependent hyperpolarization and relaxation in intact myometrium using conventional microelectrode recordings, isometric force measurements and Ca2+ imaging analysis. Aim 3, we will use molecular and protein techniques to investigate TREK-1 expression in murine myometrium in relation to native SDK channels. Aim 4, we will characterize changes in TREK-1 expression in non-pregnant, pregnant, postpartum and ovariectomized murine models to understand the functional expression of native SDK channels under various levels of female hormones. In particular this investigation will expand the molecular mechanism of estrogen regulation of TREK-1 channels that leads to the different levels of myometrial compliance. Understanding this mechanism may allow for clinical intervention in the modulation of labor and delivery, reducing the number of premature births and subsequent disorders. PUBLIC HEALTH RELEVANCE: The rate of premature birth has grown by more than 30 percent in the last 20 years. In about 40 percent of all cases of preterm birth, the causes of preterm labor are unknown. Experiments outlined in this proposal are designed to evaluate changes in uterine TREK-1 channels that accompany acute and chronic changes in female steroid hormone levels to study a potential functional relationship with pregnancy and delivery mechanisms.

描述（由申请人提供）：在大多数物种中，包括人类，17β-雌二醇和黄体酮的血浆水平在怀孕期间增加。妊娠后期女性类固醇激素的升高与子宫肌层平滑肌的电重塑有关。这些观察结果表明，雌性类固醇激素可以影响与子宫平滑肌兴奋性有关的离子电导的活性和/或表达。虽然有报道分娩机制与雌性激素水平下降有关，但从离子通道表达角度来看，临产收缩的机制尚未得到明确研究。在怀孕期间，子宫肌层的质量急剧增加，而宫内压力仅略有增加。为了实现这一目标，子宫肌层的平滑肌必须在怀孕期间保持放松。因此，质膜拉伸激活的 K+ 电导可能是子宫肌源性反应的重要组成部分。多个 K+ 通道参与子宫肌层静息膜电位的调节和动作电位的复极化。最近报道了独特的 K+ 通道，它由四个跨膜片段和两个孔域 (K2P) 组成。在 K2P 通道中，TREK-1、TREK-2 和 TRAAK 具有独特的功能特性，代表了第一个克隆的拉伸激活 K+ 通道。先前的研究表明，女性类固醇激素会影响许多 K+ 通道的转录表达。我们将在本提案中解决以下具体目标：目标1，我们将使用膜片钳方法识别小鼠子宫肌层中的SDK通道功能表达，并表征一氧化氮及其细胞内信号传导机制对这些通道的调节。目标 2，我们将使用传统的微电极记录、等长力测量和 Ca2+ 成像分析来表征完整子宫肌层中的拉伸依赖性超极化和松弛。目标 3，我们将使用分子和蛋白质技术来研究小鼠子宫肌层中 TREK-1 的表达与原生 SDK 通道的关系。目标 4，我们将表征非妊娠、妊娠、产后和卵巢切除小鼠模型中 TREK-1 表达的变化，以了解天然 SDK 通道在不同水平的雌性激素下的功能表达。特别是，这项研究将扩展雌激素调节 TREK-1 通道的分子机制，从而导致不同水平的子宫肌层顺应性。了解这一机制可能有助于对分娩和分娩的调节进行临床干预，减少早产和随后的疾病的数量。公共卫生相关性：过去 20 年来，早产率增长了 30% 以上。在大约 40% 的早产病例中，早产的原因尚不清楚。该提案中概述的实验旨在评估子宫 TREK-1 通道的变化，这些变化伴随着女性类固醇激素水平的急性和慢性变化，以研究与妊娠和分娩机制的潜在功能关系。