Central Rho-Kinase Inhibition and Ang II-induced Sympatho-excitation

描述（由申请人提供）：本申请描述了一个研究计划和研究过程中导致博士学位。作为M.D.的一部分/博士内布拉斯加大学医学中心的项目。研究计划的首要假设是，血管紧张素II（Ang II）激活中枢Rho激酶有助于疾病状态（如心力衰竭，高血压）的交感神经兴奋，Rho激酶抑制可能是这些疾病的治疗靶点。为解决这一问题，提出了三个具体目标。具体目标1：确定中枢Rho激酶抑制对慢性全身性血管紧张素II输注引起的自主神经张力调节的影响。这些实验将在长期使用仪器的清醒新西兰白色家兔中进行。Rho激酶抑制剂法舒地尔将在Ang II输注期间脑室内给药，并通过肾交感神经活动、压力反射敏感性、心率变异性、美托洛尔和阿托品血浆去甲肾上腺素后的心率变化以及神经节阻滞后动脉压的变化，评估法舒地尔和溶剂处理家兔的自主神经张力。具体目标二：确定中枢Rho激酶抑制对交感神经核中交感兴奋性活性氧（ROS）和交感抑制性一氧化氮（NO）之间平衡的影响，以响应全身性Ang II输注。我们将使用几种方法来量化氧化应激和各种蛋白质相关的活性氧和NO的生物利用度在自主中枢的脑干和下丘脑。这些研究将在特定目标1中使用的家兔组织上进行。具体目标3：确定RhoA在体外的过表达是否增强了Ang II介导的ROS增加和NO生物利用度降低。在神经元细胞系中，我们将使用与特异性目标2相同的分子终点，评估Rho激酶激活剂RhoA过表达对ROS和NO之间平衡的影响，以响应Ang II刺激。总体而言，这些研究将提供新的信息，神经元通路，可能是一个治疗目标，在交感神经兴奋性疾病状态。我的培训教学部分也被描述，包括期刊俱乐部，研讨会介绍，并在先进的生理学，统计学，工程和科学写作课程。我的综合考试将安排在2013年底或2014年初。我将得到支持参加科学会议，介绍我的研究成果。在我读博士的时候。在接受培训后，我将继续接触临床医学，在免费诊所协助工作，跟随联合国医务委员会的医生，并参加临床研讨会。

DESCRIPTION (provided by applicant): This application describes a research plan and course of study leading to the Ph.D. degree as part of the M.D./Ph.D. Program at The University of Nebraska Medical Center. The overarching hypothesis of the research plan is that activation of central Rho-kinase by Angiotensin II (Ang II) contributes to sympathoexcitation in disease states (like heart failure, hypertension) and that Rho-kinase inhibition may be a therapeutic target in these diseases. To address this issue, three specific aims are proposed. Specific Aim 1: Determine the effect of central Rho-kinase inhibition on the regulation of autonomic tone in response to chronic systemic Ang II infusion. These experiments will be carried out in chronically instrumented, conscious New Zealand White rabbits. The Rho-kinase inhibitor, fasudil, will be administered intracerebroventricularly during Ang II infusion, and autonomic tone will be assessed in fasudil- and vehicle-treated rabbits by renal sympathetic nerve activity, baroreflex sensitivity, heart rate variability, changes in heart rate after metoprolol and atropine plasma norepinephrine, and changes in arterial pressure in response to ganglionic blockade. Specific Aim 2: Determine the effects of central Rho-kinase inhibition on the balance between sympatho-excitatory reactive oxygen species (ROS) and sympatho-inhibitory nitric oxide (NO) in sympathetic nuclei in response to systemic Ang II infusion. We will use several methods to quantify oxidative stress and a variety of proteins relevant to ROS and NO bioavailability in autonomic centers of the brainstem and hypothalamus. These studies will be performed on tissue from the rabbits used in Specific Aim 1. Specific Aim 3: Determine if overexpression of RhoA in vitro potentiates Ang II-mediated increases in ROS and decreases in NO bioavailability. In a neuronal cell line, we will evaluate the effects of overexpression of RhoA, the activator of Rho-kinase, on the balance between ROS and NO in response to Ang II stimulation, using the same molecular endpoints as Specific Aim 2. Overall, these studies will provide novel information on a neuronal pathway that could be a therapeutic target in sympathoexcitatory disease states. The didactic component of my training is also described, including journal club, seminar presentations, and courses in advanced physiology, statistics, engineering, and scientific writing. My comprehensive exam will be scheduled for late 2013 or early 2014. I will be supported to attend scientific conferences to present the results of my research. During my Ph.D. training, I will continue to be exposed to clinical medicine by assisting at a free clinic, shadowing UNMC physicians, and attending clinical seminars.