Altered CNS Intercellular Signaling Mechanisms in Cardiovascular Disease

描述(申请人提供)：神经体液激活，包括交感神经兴奋和循环激素水平的增加，如血管加压素，是心力衰竭(HF)病理生理学的主要参与者，直接影响这种疾病的发病率和死亡率。虽然下丘脑室旁核(PVN)和视上核(SON)在HF时对神经体液激活的作用已被证实，但对HF时这些核团内神经元活动增加的确切细胞机制仍缺乏全面的了解。神经元细胞内钙离子水平(D[Ca~(2+)]i)的活性依赖性变化不仅是影响膜兴奋性的关键信号，而且还影响神经可塑性和基因表达。兴奋性递质谷氨酸主要通过NMDAR发挥作用，是D[Ca~(2+)]i信号的主要来源(NMDA-DCa~(2+))，在神经分泌和交感前神经元活动的调节中起重要作用。重要的是，越来越多的证据支持心力衰竭患者谷氨酸功能的恶化。此外，与NMDA-DCa+直接相关的其他信号机制(如一氧化氮、ROS、GABA)也在心衰时发生改变。NMDA-DCa~(2+)的整体功能结果在很大程度上依赖于D[Ca~(2+)]i的时空模式，因此，阐明影响NMDA-DCa~(2+)特性的精确机制，以及这些机制的异常变化如何可能有助于心力衰竭神经元活动的加剧，具有重要的意义。我们已经获得了令人兴奋的初步数据，支持线粒体，传统上被视为静态的细胞动力装置，是关键的和动态的细胞器，积极影响 NMDAR的有效性，通过抑制其钙依赖的耦合到其他细胞内信号通路，进而影响整个SON/PVN神经分泌和交感前神经元的活动。此外，我们还发现，钝化的NMDAR-线粒体串扰导致NMDAR效应增强，并加剧了NMDA-DCa2+，导致依赖钙的Trp通道家族的激活增加，最终导致HF神经元活动异常增加。在这里，我们将检验一个中心假设，即线粒体结构-功能完整性的破坏会导致谷氨酸兴奋功能的加剧，谷氨酸兴奋功能通过加强与钙敏感的TRPM4通道的偶联而导致HF中神经元活动的增强。这一假说将在3个具体目标中得到验证：1-阐明线粒体在SON/PVN神经元NMDAR-[Ca~(2+)]i信号形成中的作用，2-阐明线粒体在SON/PVN神经元HF大鼠NMDAR-[Ca~(2+)]i信号改变中的结构和功能机制，以及3-确定线粒体功能障碍对NMDAR介导的心衰大鼠神经元兴奋性的影响。我们期望这项工作的结果能够拓宽我们对下丘脑调节神经体液流出的基本细胞机制的理解，以及这些机制的变化如何有助于心力衰竭中的神经体液激活。

DESCRIPTION (provided by applicant): Neurohumoral activation, including sympathoexcitation and increased circulating hormonal levels such as vasopressin, is a major player in the pathophysiology of heart failure (HF), directly influencing morbidity and mortality i this disease. While the contribution of the hypothalamic paraventricular (PVN) and supraoptic (SON) nuclei to neurohumoral activation in HF is established, a comprehensive understanding of the precise cellular mechanisms contributing to increased neuronal activity within these nuclei in HF remains elusive. Activity-dependent changes in neuronal intracellular Ca2+ levels (D[Ca2+]I)act as a critical signal influencing not only membrane excitability, but also neuroplasticity and gene expression. The excitatory transmitter glutamate, acting primarily via NMDA receptors (NMDAR), is a major source of D[Ca2+]I signaling (NMDA-DCa2+], playing an important role in the regulation of neurosecretory and presympathetic neuronal activity. Importantly, a growing body of evidence supports an exacerbated glutamate function in HF. Moreover, other signaling mechanisms that are directly linked to NMDA-DCa2+ (e.g, nitric oxide, ROS, GABA) are also altered in HF. The overall functional consequences of NMDA-DCa2+ are largely dependent on the spatiotemporal pattern of the D[Ca2+]I Thus, elucidating the precise mechanisms that influence the NMDA-DCa2+ properties, and how abnormal changes in these mechanisms may contribute to exacerbated neuronal activity in HF, is highly relevant. We have obtained exciting preliminary data supporting that mitochondria, classically viewed as static cellular power plants, are critical and dynamic organelles that actively influence NMDAR efficacy, by restraining its Ca2+-dependent coupling to other intracellular signaling pathways, influencing in turn overall SON/PVN neurosecretory and presympathetic neuronal activity. Moreover, we found that a blunted NMDAR- mitochondria crosstalk results in an enhanced NMDAR efficacy and exacerbated NMDA-DCa2+ leading to increased activation of the Ca2+- dependent family of TRP channels, and ultimately, abnormally elevated neuronal activity in HF. Here, we will test the central hypothesis that disruption of mitochondrial structurl-functional integrity results in exacerbated glutamate excitatory function, which via a strengthened coupling to Ca2+-sensitive TRPM4 channels, leads to enhanced neuronal activity in HF. This hypothesis will be tested in 3 specific aims: 1- To elucidate the role of mitochondria in shaping NMDAR-[Ca2+]i signaling in SON/PVN neurons, 2- To elucidate structural and functional mitochondrial mechanisms contributing to altered NMDAR-[Ca2+]i signaling in SON/PVN neurons HF rats, and 3- To determine the consequences of mitochondrial dysfunction on NMDAR-mediated neuronal excitability in HF rats. We expect results from this work to broaden our understanding of basic cellular mechanisms contributing to the hypothalamic regulation of neurohumoral outflows, and how changes in these mechanisms may contribute to neurohumoral activation in heart failure.