Phenotypic characterization of nitric oxide regulation of cardiac function by MRI

描述（由申请人提供）：兴奋-收缩（EC）偶联和钙（Ca 2+）循环在调节心脏收缩力和心脏疾病的发展中起重要作用。Ca 2+处理障碍在心力衰竭中发生在多个水平，并且与病理表现密切相关。然而，有有限的手段来评估体内EC偶联的改变。最近的研究表明，神经元型一氧化氮合酶（nNOS）在调节EC耦合的作用，与心肌收缩的nNOS调制的几个方面，其在心脏疾病中的作用仍然知之甚少。在心脏中，已报道nNOS与肌膜、肌浆网（SR）和线粒体相关。nNOS与其效应蛋白的共定位已被认为是心肌控制的重要机制。然而，采用全局nNOS敲除模型（NOS 1-/-小鼠）的研究无法通过空间限制解决NO作用的复杂性。因此，拟议研究的目标是：1）开发锰增强磁共振成像（MEMRI）方法，用于心肌中Ca 2+摄取的体内表征，这是Ca 2+循环的第一步; 2）将最先进的MRI技术应用于研究nNOS在心脏功能调节和心肌病发展中的不同作用。我们将表征两种nNOS破坏不同的小鼠模型，即，整体nNOS敲除小鼠和1-小肌营养不良蛋白敲除小鼠，其导致仅细胞膜中的nNOS破坏。通过结合在体内MRI表征的心脏表型，如功能和Ca 2+摄取与体外分子/细胞分析的心肌细胞收缩性和Ca 2+循环的系统比较研究的新的小鼠模型与不同模式的nNOS中断，这种方法提供了独特的机会解剖的作用，nNOS在调节心脏功能的不同亚细胞室。阐明nNOS对心肌收缩和疾病进展的作用机制将使nNOS成为心血管疾病的治疗靶点。 公共卫生相关性：兴奋-收缩偶联和钙离子循环在心脏收缩力的调节和心脏疾病的发生发展中起着重要作用。神经元型一氧化氮合酶（nNOS）调节EC偶联中的几个关键过程，在心力衰竭中异常。针对nNOS的药物干预可能是心力衰竭的有效治疗。该研究的目的是开发对钙循环改变敏感的体内成像方法，并应用该方法阐明nNOS在EC偶联和nNOS破坏小鼠模型中的心脏功能中的作用。

DESCRIPTION (provided by applicant): Excitation-contraction (EC) coupling and calcium (Ca2+) cycling play an important role in regulating cardiac contractile force and in the development of cardiac diseases. Disturbance of Ca2+ handling occurs at multiple levels in heart failure and is closely related to pathological performance. However, there are limited means to evaluate alterations in EC coupling in vivo. Recent studies have indicated the role of neuronal NOS (nNOS) in regulating EC coupling, with several aspects of nNOS modulation of myocardial contractility and its role in cardiac diseases still poorly understood. In the heart, nNOS has been reported to be associated with the sarcolemma, sarcoplasmic reticulum (SR), and mitochondria. Co-localization of nNOS with its effector proteins has been suggested to be important mechanisms in myocardial control. However, studies that employ a global nNOS knockout model, the NOS1-/- mouse, cannot address the complexities of NO action through spatial confinement. Therefore, the objectives of the proposed research are 1) to develop manganese-enhanced magnetic resonance imaging (MEMRI) methods for in vivo characterization of Ca2+ uptake in myocardium, the first-step in Ca2+ cycling; 2) to apply state-of-the-art MRI technology to the investigation of the differential roles of nNOS in the regulation of cardiac function and the development of cardiomyopathy. We will characterize two mouse models that differ in nNOS disruption, i.e., the global nNOS knockout mouse and the 1-dystrobrevin knockout mouse, which leads to the disruption of nNOS in cell membrane only. By combining in vivo MRI characterization of cardiac phenotypes such as function and Ca2+ uptake with in vitro molecular/cellular analysis of myocyte contractility and Ca2+ cycling in a systematic comparative study of novel mouse models with distinctive modes of nNOS disruption, this approach offers unique opportunity for dissecting the roles of nNOS in regulating cardiac function in distinct subcellular compartments. The mechanistic elucidation of the effects of nNOS on myocardial contraction and disease progression will allow nNOS to be a therapeutic target in cardiovascular diseases. PUBLIC HEALTH RELEVANCE: Excitation-contraction (EC) coupling and calcium cycling play an important role in regulating cardiac contractile force and in the development of cardiac diseases. Neuronal nitric oxide synthase (nNOS) regulates several key processes in EC coupling and is abnormal in heart failure. Pharmacological intervention that targets nNOS may be effective treatment for heart failure. The goal of the proposed research is to develop in vivo imaging method that is sensitive to altered calcium cycling, and to apply this method to elucidate the role of nNOS in EC coupling and cardiac function in mouse models of nNOS disruption.