Novel role of central AT2R in blood pressure regulation

描述(由申请者提供)：项目摘要/摘要高血压是一个普遍的健康问题，也是心血管疾病的主要风险因素，在美国，心血管疾病是导致死亡的主要原因。尤其值得关注的是耐药高血压，它伴随着交感神经系统活动的增强，表明血压升高是由神经源性起源引起的。近三分之一的高血压患者属于这一类，目前还没有有效的药物治疗，确定治疗或预防神经源性高血压的策略对公众健康具有重要意义。我的总体职业目标是成为一名独立的学术科学家，研究导致神经源性高血压的神经回路损伤，以及缓解这些损伤的治疗方法的开发。我们的期望是，我的研究将对了解神经源性高血压的原因和开发用于治疗的疗法做出实质性贡献。本申请中提出的活动旨在促进TIS目标的实现，并将探索一种治疗神经源性高血压的新靶点--脑内表达的血管紧张素2型受体(AT2R)。这些实验测试了一个总体假设，即激活AT2R在投射到下丘脑室旁核(PVN；一个控制交感神经流出和血压的重要大脑区域)的神经元上的激活对血压有负面调节作用，有可能使AT2R的激活成为降压药物的合适靶点。我的研究生训练使用实验室的啮齿动物来研究血管紧张素-II是如何影响体重和葡萄糖代谢的神经控制的。血管紧张素-II是一种与高血压和心血管疾病的发展密切相关的多肽。这一系列研究向我介绍了对血管紧张素-II敏感的中枢通路，这激发了我对神经源性高血压的兴趣。因此，我选择了佛罗里达大学科林·萨姆纳斯博士的实验室来进行博士后培训。萨姆纳斯博士是神经源性高血压领域的领先专家，他的实验室是加州大学高血压中心的一部分，该中心由致力于研究高血压的核心设施和近50名教职员工组成。这种培训环境有助于我在博士后成功地提出NRSA建议，使我有能力评估啮齿动物的心血管功能，或许更重要的是，我发现实验诱导的高血压引起了控制血压的神经元的电生理特性的变化，最终增加了它们的兴奋度。考虑到这些结果，我决定有效的疗法应该减少或逆转这种增加的兴奋；然而，我也决定，旨在了解神经元电生理特性的概念和技术方法的额外培训对于发展这一研究路线是必不可少的。因此，K99阶段的主要目标是回答有关特定AT2R的结构和功能的一些基本问题，这些AT2R被定位为减少交感神经流出和血压，同时为体外膜片钳电生理学提供额外的培训。预计确定AT2R对神经源性高血压的治疗效用和亚细胞神经电生理学方面的专业知识可以与启动我的独立研究生涯的专业发展活动相辅相成。在第一个目标中，实验将结合遗传学和神经解剖学技术来测试特定的假设，即与下丘脑室旁核内自主神经前神经元接触的AT2R表达神经元表达抑制性神经递质(GABA)，从而定位它们以降低血压和自主神经功能。在目标2中，实验将使用膜片钳电生理技术来验证特定的假设，即在投射到PVN的GABA神经元上激活AT2R将促进抑制性(即GABA能)神经传递，这将导致PVN自主前神经元的活动减少。这些实验不仅将准确地确定AT2R的激活如何影响神经元网络中的活动，而且它们还将成为我学习膜片钳电生理学的训练工具，这项技术对于理解离散神经元群体中的亚细胞变化如何影响整个动物生理学至关重要。重要的是，我的背景，再加上电生理学方面的专业知识，将使我的研究计划独一无二，因为它将允许研究血管紧张素-II如何通过AT2R影响控制心血管功能的特定神经元的兴奋性。AIM 3将包含在R00阶段，并将结合我过去的培训和我新获得的技能来确定AT2R在基础和神经源性高血压期间的血压调节中的作用。利用CRE/LOX系统和药理学方法，我将选择性地激活或抑制投射到PVN的神经元上的AT2R，并测试这些AT2R负向调节血压和交感神经系统流出的具体假设。总体而言，建议的研究具有重要意义，因为它们可能揭示治疗神经源性高血压的新靶点，同时为我建立一个独立的研究计划做好准备，以解决高血压的问题。

DESCRIPTION (provided by applicant): Project Summary/Abstract Hypertension is a widespread health problem and a major risk factor for cardiovascular disease, the leading cause of death in the USA. Of particular concern is drug-resistant hypertension, which is accompanied by enhanced sympathetic nervous system activity, indicating that the increased blood pressure arises from neurogenic origins. Nearly one-third of hypertensive patients fall into this category for which there are no effective medications and determining strategies to treat or prevent neurogenic hypertension has great significance for public health. My overall career goal is to become an independent academic scientist that studies impairments in neural circuits that elicit neurogenic hypertension, as well as the development of therapeutics that alleviate these impairments. The expectation is that my research will contribute substantively to the understanding of the causes of neurogenic hypertension and to the development of therapeutics used to treat it. The activities proposed in this application are designed to facilitate reaching tis goal and will investigate a novel therapeutic target for neurogenic hypertension - angiotensin type-2 receptors (AT2R) expressed within the brain. The experiments test the overall hypothesis that activation of AT2R on neurons that project to the paraventricular nucleus of the hypothalamus (PVN; a brain region important for controlling sympathetic outflow and blood pressure) negatively-regulate blood pressure, potentially making activation of AT2R a suitable target for antihypertensive medications. My graduate training used laboratory rodents to examine how angiotensin-II, a peptide heavily implicated in the development of hypertension and cardiovascular disease, influenced the neural control of body weight and glucose metabolism. This line of research introduced me to the central pathways that were sensitive to angiotensin-II, which piqued my interest in neurogenic hypertension. Consequently, I chose the laboratory of Dr. Colin Sumners at the University of Florida to conduct my postdoctoral training. Dr. Sumners is a leading expert in the field of neurogenic hypertension and his laboratory is part of the Hypertension Center at UF, which is comprised of core facilities and nearly 50 faculty members dedicated to studying high blood pressure. This training environment contributed to my successful post-doctoral NRSA proposal that afforded competence with the assessment of cardiovascular function in rodents, and perhaps more importantly, found that experimentally-induced hypertension elicited changes within the electrophysiological properties of neurons controlling blood pressure that ultimately increased their excitation. Taking these results into account, I determined that effective therapeutics should decrease or reverse this increased excitation; however, I also determined that additional training in conceptual and technical approaches aimed at understanding the electrophysiological properties of neurons was imperative to developing this line of research. Accordingly, the primary objectives of the K99-phase are to answer some fundamental questions regarding the structure and function of specific AT2R that are positioned to decrease sympathetic outflow and blood pressure, while providing additional training for in vitro patch-clamp electrophysiology. It is anticipated that determining the therapeutic utility of AT2R for neurogenic hypertension and expertise in subcellular neural electrophysiology can be complemented by professional development activities to launch my independent research career. In the first Aim, experiments will combine genetic and neuroanatomical techniques to test the specific hypothesis that AT2R-expressing neurons that make contacts onto preautonomic neurons within the paraventricular nucleus of the hypothalamus express the inhibitory neurotransmitter (GABA), thereby positioning them to decrease blood pressure and autonomic function. In Aim 2, experiments will use patch- clamp electrophysiological techniques to test the specific hypothesis that activation of AT2R on GABA neurons that project to the PVN will facilitate inhibitory (i.e., GABAergic) neurotransmission and that this will lead to reduced activity of PVN preautonomic neurons. These experiments will not only determine precisely how activation of AT2R impact activity within a neuronal network, but they will also serve as a training vehicle for me to learn patch-clamp electrophysiology, a technique that is essential to understanding how subcellular changes in a discrete population of neurons can impact whole animal physiology. Importantly, my background, combined with expertise in electrophysiology will make my research program unique, as it will allow for the study of precisely how angiotensin-II acting through AT2R influences the excitability of specific neurons that control cardiovascular function. Aim 3 will be contained within the R00-phase and will partner my past training with my newly-acquired skills to determine the role of AT2R in blood pressure regulation basally and during neurogenic hypertension. Using the Cre/lox system and pharmacological approaches, I will selectively activate or inhibit AT2R on neurons that project to the PVN and test the specific hypothesis that these AT2R negatively regulate blood pressure and sympathetic nervous system outflow. Collectively, the proposed studies are significant because they may uncover a novel therapeutic target for treatment of neurogenic hypertension while preparing me to establish an independent research program that addresses a problem with high