Novel role of central AT2R in blood pressure regulation

项目摘要/摘要高血压是一种普遍存在的健康问题，是心血管疾病的主要危险因素，是美国人死亡的主要原因。特别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无有效药物治疗的高血压患者，确定治疗或预防神经源性高血压的策略对公共卫生具有重要意义。我的总体职业目标是成为一名独立的学术科学家，研究引起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发展。期望我的研究将对理解神经源性高血压的原因和治疗方法的发展做出实质性的贡献。本应用程序中提出的活动旨在促进实现这一目标，并将研究神经源性高血压的新治疗靶点-脑内表达的血管紧张素2型受体（AT2R）。这些实验验证了一个整体假设，即投射到下丘脑室旁核（PVN；一个控制交感神经外流和血压的重要脑区）的神经元上的AT2R的激活对血压有负调节作用，这可能使AT2R的激活成为抗高血压药物的合适靶点。我的研究生训练使用实验室啮齿动物来研究血管紧张素- ii（一种与高血压和心血管疾病的发展密切相关的肽）如何影响体重和葡萄糖代谢的神经控制。这条研究路线使我了解了对血管紧张素- ii敏感的中枢通路，这激起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Colin Sumners博士的实验室进行我的博士后培养。Sumners博士是神经源性高血压领域的领先专家，他的实验室是佛罗里达大学高血压中心的一部分，该中心由核心设施和近50名致力于研究高血压的教职员工组成。这种训练环境促成了我成功的博士后nssa提案，提供了评估啮齿动物心血管功能的能力，也许更重要的是，发现实验诱导的高血压引起控制血压的神经元电生理特性的变化，最终增加了它们的兴奋。考虑到这些结果，我确定有效的治疗方法应该减少或逆转这种增加的兴奋；然而，我也确定，在概念和技术方法上的额外训练，旨在理解神经元的电生理特性，是发展这条研究路线的必要条件。因此，k99期的主要目的是回答一些关于特定AT2R的结构和功能的基本问题，AT2R的定位是降低交感神经流出和血压，同时为体外膜片钳电生理提供额外的训练。可以预期的是，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的专业知识可以通过专业发展活动来补充，以启动我的独立研究生涯。在第一个目标中，实验将结合遗传学和神经解剖学技术来验证一个特定的假设，即表达at2r的神经元与下丘脑室旁核内的自主神经前神经元接触，表达抑制性神经递质（GABA），从而使它们处于降低血压和自主神经功能的位置。在Aim 2中，实验将使用膜片钳电生理技术来验证特定假设，即投射到PVN的GABA神经元上的AT2R的激活将促进抑制性（即GABA能）神经传递，这将导致PVN前自主神经元的活性降低。这些实验不仅将精确地确定AT2R的激活如何影响神经元网络中的活动，而且还将作为我学习膜片钳电生理学的训练工具，这是一种对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的技术。重要的是，我的背景，结合电生理学的专业知识，将使我的研究项目独一无二，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r0 -阶段，并将我过去的培训与我新获得的技能相结合，以确定AT2R在基本血压调节和神经源性高血压中的作用。使用Cre/lox系统和药理学方法，我将选择性地激活或抑制神经元上的AT2R，并测试这些AT2R负调节血压和交感神经系统流出的特定假设。总的来说，拟议的研究意义重大，因为它们可能会发现治疗神经源性高血压的新治疗靶点，同时为我建立一个解决高血压问题的独立研究项目做准备

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