AT2 Receptors in the RVLM: Regulation of Sympathetic Outflow in Heart Failure

摘要 慢性心力衰竭（CHF）是美国发病率和死亡率的主要原因， 交感神经过度活跃和肾素-血管紧张素系统激活的特征。这些都是 主要治疗目标。在这个项目中，我们提出了三个目标，以探索潜力 在延髓头端腹外侧过度表达血管紧张素2型受体（AT 2 R）的益处 （RVLM）与CHF大鼠。此外，我们还将确定涉及的潜在机制， RVLM中AT 2 R过表达的交感神经抑制作用。已经确定，在 与AT 1 R的影响相反，AT 2 R易化神经元钾通道和电流， 其使膜电位超极化并抑制神经元兴奋性。另一方面我们 初步实验显示CHF大鼠RVLM中AT 2 R蛋白表达下调。 这些现象使我们假设RVLM中AT 2 R信号的减少有助于 通过提高前交感神经元的兴奋性来抑制这种综合征的交感兴奋。我们的全球 该建议中假设是通过基因转移在RVLM中过表达AT 2 R将 减少或正常化CHF中的交感神经激活，因此有益于该综合征。在 初步实验中，我们成功地建立了一个大鼠模型，其中AT 2 R选择性地超过 通过将AT 2 R病毒载体直接递送到RVLM中表达。采用这种动物模型， 我们将通过以下三个具体目标来检验我们的假设。目的1：确定 正常和CHF大鼠血流动力学、心功能和交感神经流出量 RVLM中AT 2 R的过表达。这一目标包括两个组成部分。首先，我们将确定 RVLM中过表达AT 2 R对动脉血压（AP）、心率（HR） 心功能和去甲肾上腺素排泄在清醒的正常和CHF大鼠。此外，水的摄入， 将测量尿排泄和体重。其次，我们将观察 将AT 1 R和AT 2 R的激动剂和拮抗剂微量注射到RVLM中， AP、HR和肾交感神经活动（RSNA）。我们还将探讨细胞内 AR 2 R信号传导交感神经抑制。这包括NO/cGMP和PLA 2/AA/12-LO/PP 2A途径。目的 2：确定过表达AT 2 R对单个前交感神经元活动的影响， 麻醉大鼠的RVLM。我们将直接记录RVLM的细胞外单个单位放电 AT 2 R过表达后的前交感神经元。目的3：确定 AT 2 R过表达对RVLM前交感神经元钾电流的影响。采用 采用膜片钳和脑干切片的方法，直接记录前交感神经的钾电流 AT 2 R过度表达后的神经元。这些研究将使人们更好地了解 充血性心力衰竭时前交感神经元中的血管紧张素信号传导。他们将强调以下方面的重要性 AT 1和AT 2受体信号传导在设定交感神经兴奋水平和识别 CHF治疗的可能新靶点。

ABSTRACT Chronic heart failure (CHF) is a leading cause of morbidity and mortality in the United States with the characteristics of sympathetic overactivity and activation of the renin-angiotensin system. These are the primary therapeutic targets for this syndrome. In this project, we propose three aims to explore the potential benefit of over expressing angiotensin type 2 receptor (AT2R) expression in the rostral ventrolateral medulla (RVLM) in rats with CHF. Moreover we will also determine the underlying mechanisms involved in the sympatho-inhibitory effects of AT2R overexpression in the RVLM. It has been firmly established that, in contrast to the influences of the AT1R, the AT2R facilitates the neuronal potassium channel and current, which hyperpolarizes membrane potential and suppresses neuronal excitability. On the other hand, our preliminary experiments show a down regulation of AT2R protein expression in the RVLM of rats with CHF. These phenomena lead us to postulate that a decrease in AT2R signaling in the RVLM contributes to sympatho-excitation of this syndrome by elevating the excitability of presympathetic neurons. Our global hypothesis in this proposal is that over expression of the AT2R in the RVLM by gene transfer will reduce or normalize sympathetic activation in CHF, and therefore benefit this syndrome. In preliminary experiments, we have successfully produced a rat model in which the AT2R is selectively over expressed in the RVLM by direct delivery of AT2R viral vectors into this area. Employing this animal model, we will test our hypothesis by pursuing the following 3 Specific Aims. AIM 1: To determine the hemodynamic, cardiac function, and sympathetic outflow in normal and CHF rats with overexpression of AT2R in the RVLM. This Aim includes two components. First, we will determine the chronic effects of over expressing AT2R in the RVLM on arterial blood pressure (AP), heart rate (HR), cardiac function, and norepinephrine excretion in conscious normal and CHF rats. In addition, water intake, urine excretion, and body weight will be measured. Second, we will observe the acute effects of microinjecting agonists and antagonists of AT1R and AT2R into the RVLM with AT2R over expression on AP, HR, and renal sympathetic nerve activity (RSNA). We will also explore the involvement of intracellular AR2R signaling sympatho-inhibition. This includes the NO/cGMP and PLA2/AA/12-LO/PP2A pathways. AIM 2: To determine the effects of overexpressing AT2R on single presympathetic neuronal activity in the RVLM of anesthetized rats. We will directly record extracellular single unit firing of RVLM presympathetic neurons following overexpression of AT2R. AIM 3: To determine the effects of overexpressing AT2R on potassium current of presympathetic neurons in the RVLM. Employing patch clamp and brainstem slice preparations, we will directly record potassium currents of presympathetic neurons following overexpression of AT2R. These studies will lead to an enhanced understanding of angiotensin signaling in presympathetic neurons in the setting of CHF. They will highlight the importance of a balance between AT1 and AT2 receptor signaling in setting the level of sympatho-excitation and identify possible new targets for therapy in CHF.