Oxidation and Pharmacologic Activation of IK/SK Channels

IK/SK 通道的氧化和药理学激活

• 批准号: 7568223
• 负责人: DANIEL C DEVOR
• 金额: $ 35.5万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-02-01 至 2011-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7568223
• 关键词: Agonist; Atherosclerosis; Balloon Angioplasty; Behavior; Blood Pressure; Blood Vessels; Bradykinin; Calcium-Activated Potassium Channel; Cardiovascular Diseases; Cholinergic Agonists; Clinical; Coronary Arteriosclerosis; Development; Disease; Endothelial Cells; Endothelium; Environment; Feedback; Fluorescence Resonance Energy Transfer; Goals; Homeostasis; Hydrogen Peroxide; Hypertension; Inflammatory; Inflammatory Response; Ion Channel; Ischemia; Laboratories; Mediating; Molecular; Molecular Mechanisms of Action; Muscle relaxation phase; Mutagenesis; Oxidants; Physiological; Potassium Channel; Process; Qualifying; Reactive Oxygen Species; Regulation; Relaxation; Reperfusion Therapy; Research Personnel; Role; Series; Smooth Muscle; Structure; System; Techniques; Therapeutic; Transgenic Mice; Universities; Vascular Diseases; Vascular Smooth Muscle; Vasodilation; innovation; oxidation; patch clamp; response

The role of intermediate (IK)and small (SK) conductance, Ca -activated K+ channels have been unequivocally demonstrated in the endothelial-dependent relaxation of vascular smooth muscle. Both IK and SK channels are activated during agonist- and flow-induced vasodilation as well as in the presence of increased reactive oxygen species (ROS), which are associated with virtually all cardiovascular disease. The expression of these channels in endothelia has also been shown to be compromised following balloon angioplasty. Finally, these channels are known to be critical to EDHF-mediated vasodilation, which is compromised in a host of cardiovascular diseases. These results have led to the proposal that the pharmacological activation of endothelial IK and SK channels would be of clinical benefit in a wide array of cardiovascular diseases. Our laboratory was the first to identify a series of structurally similar pharmacological openers of IK and SK channels. To further define the role of IK and SK channels in endothelial function and how they may be pharmacologically manipulated for clinical benefit requires us to answer two critical unknowns. First, how do ROS alter IK and SK channel function and therefore endothelial function? Second, what is the molecular mechanism of action for the known openers of IK and SK channels? Thus, we propose the following aims: (i) Define the mechanisms involved in the reactive oxygen species-dependent regulation of IK and SK channels. We will utilize a combination of patch-clamp and mutagenesis techniques to define the mechanisms whereby oxidizing agents activate IK and SK channels. These studies will be carried out on both heterologously expressed channels as well as on primary cultures of endothelial cells, (ii) We will define the molecular mechanism whereby pharmacological activators of IK and SK channels increase channel activity. These studies will be carried out utilizing a combination of patch-clamp and mutagenesis techniques, (iii)We will utilize FRET to define inter- and intra-subunit domain interactions in IK and SK channels and how physiological and pharmacological regulators of channel function modify these interactions. Defining how these interactions are altered is critical to our understanding of how these channels are regulated during the inflammatory process and how they may be manipulated pharmacologically. The results of these studies will clearly define the mechanism whereby ROS activate endothelial IK and SK channels, and thus alter vascular tone, as well as define the molecular mechanism underlying pharmacological activation of these channels; thereby furthering our understanding of how these channels may be manipulated for therapeutic benefit.

中间体（IK）和小（SK）电导的作用，CA激活的K+通道已毫无疑问 在血管平滑肌的内皮依赖性松弛中证明。 IK和SK频道都是 在激动剂和流动诱导的血管舒张以及活性氧的存在期间被激活 （ROS），几乎与所有心血管疾病有关。这些通道在内皮中的表达 在球囊血管成形术后也已被证明被妥协。最后，这些渠道是至关重要的 EDHF介导的血管舒张，在许多心血管疾病中受到损害。这些结果导致了 关于内皮IK和SK通道的药理学激活的提议将在A中具有临床益处 广泛的心血管疾病。我们的实验室是第一个确定一系列结构相似的实验室 IK和SK频道的药理学开瓶器。进一步定义IK和SK频道在内皮中的作用 功能以及如何在药理学上操纵它们以获得临床益处，这需要我们回答两个关键 未知。首先，ROS如何改变IK和SK通道功能，从而改变内皮功能？第二，是什么 IK和SK通道开瓶器的作用分子机制？因此，我们提出以下内容 目的：（i）定义与IK和SK通道的活性氧依赖性调节所涉及的机制。 我们将利用斑块夹和诱变技术的组合来定义氧化的机制 代理激活IK和SK频道。这些研究将在两个异源表达的通道上进行 以及在内皮细胞的一级培养物上，（ii）我们将定义药理的分子机制 IK和SK通道的激活剂增加了通道活动。这些研究将通过结合 斑块夹和诱变技术，（iii）我们将利用FRET来定义suber-subunit域 IK和SK通道中的相互作用以及通道功能的生理和药理调节剂如何修改 这些相互作用。定义这些相互作用的改变是我们对这些渠道的理解至关重要的 在炎症过程中受到调节，以及如何通过药理操作。这些结果 研究将明确定义ROS激活内皮IK和SK通道的机制，从而改变 血管张力，并定义这些通道的药理学激活的分子机制； 从而进一步了解如何为治疗益处操纵这些渠道。