Molecular Physiology of Neuromusclar Diseases

描述（由申请人提供）：肌强直和周期性麻痹是骨骼肌的遗传性疾病，其中电压门控离子通道的突变改变了肌膜的电兴奋性。该项目的长期目标是表征突变通道的功能缺陷，并确定异常通道行为如何产生症状。对于这些疾病，肌肉功能障碍是由间歇性的电兴奋性紊乱引起的，可能是病理性的增强或抑制。肌强直是一种兴奋性增强的障碍，其中单个刺激引起动作电位的爆发，产生持续数秒的不自主持续肌肉收缩。相反，周期性麻痹是由去极化引起的肌肉兴奋性丧失引起的。氯离子通道突变引起肌强直，而钾离子或钙离子通道突变引起周期性麻痹。骨骼肌钠通道（由SCN4A编码的NaV1.4）的错义突变可能导致同一个体的肌强直、周期性麻痹或两者兼而有之。这种临床表型变异的病理生理基础，都是由一个共同钠通道基因的突变引起的，是本提案研究的主要焦点。我们的实验方法是通过测量离子电流来确定突变通道行为的改变，然后使用计算机或动物模型来探索通道功能的特定改变是如何引起肌强直或周期性麻痹的。目的1是识别额外的、尚未表征的NaV1.4突变体的功能缺陷，从而进一步定义导致特定形式的膜兴奋性改变的门控缺陷的生物物理特征。我们和其他人之前的工作已经确定，慢失活门控是易患虚弱症的关键决定因素。与其他门控转换相比，对慢失活的了解相对较少。目的2旨在通过结合精细的电压依赖性门控方案和基于半胱氨酸扫描诱变的结构研究来提高我们对缓慢失活的理解。Aim 3通过表征NAY1.4靶向突变的小鼠模型，并使用基于计算机的肌肉兴奋性模型，探讨Na通道行为的功能缺陷如何导致肌强直或周期性麻痹。已知有30多种人类疾病是由电压门控离子通道的突变引起的。拟议的研究旨在为一组人类通道病的病理生理基础提供更完整的理解：从基因缺陷到临床症状。这些研究也将进一步加深我们在分子水平上对钠通道功能的认识，将导致对肌肉兴奋性决定因素的更好理解，将为更合理的治疗策略设计提供机制见解，并将作为理解更常见的兴奋性疾病（如癫痫或心律失常）的模型系统。

DESCRIPTION (provided by applicant): The myotonias and periodic paralyses are heritable diseases of skeletal muscle in which mutations of voltage-gated ion channels alter the electrical excitability of the sarcolemma. The long-term goals of this project are to characterize the functional defects of mutant channels and to determine how abnormal channel behavior produces symptoms. For these disorders, muscle dysfunction is caused by intermittent derangements in electrical excitability, which may be pathologically enhanced or depressed. Myotonia is a disorder of enhanced excitability wherein a single stimulus elicits a burst of action potentials that produces involuntary persistent muscle contraction lasting seconds. Conversely, periodic paralysis results from a depolarization-induced loss of muscle excitability. Chloride channel mutations cause myotonia, whereas mutations in potassium or calcium channels give rise to periodic paralysis. Missense mutations in a skeletal muscle sodium channel (NaV1.4 encoded by SCN4A) may cause myotonia, periodic paralysis, or of both in the same individual. The pathophysiological basis for this variation in clinical phenotype, all arising from mutations in a common sodium channel gene, is a major focus of the studies in this proposal. Our experimental approach is to identify alterations in the behavior of mutant channels by measuring ionic current, and then use computer or animal-based models to explore how specific alterations in channel function cause myotonia or periodic paralysis. Aim 1 is to identify the functional defects for additional, as-yet uncharacterized, NaV1.4 mutants and thereby define further the biophysical profile of gating defects that give rise to specific forms of altered membrane excitability. Prior work, by us, and others, has identified slow-inactivation gating as a critical determinant in the predisposition to attacks of weakness. In comparison to other gating transitions, relatively little is known about slow inactivation. Aim 2 seeks to improve our understanding of slow inactivation through a combination of refined voltage-dependent gating protocols and structural studies based on cysteine-scanning mutagenesis. Aim 3 explores how functional defects in Na channel behavior cause myotonia or periodic paralysis by characterizing a mouse model with a targeted mutation in NAY1.4 and using computer-based models of muscle excitability. More than 30 human disorders are known to be caused by mutations in voltage-gated ion channels. The proposed studies are designed to provide a more complete understanding of the pathophysiological basis for a group of human channelopathies: from gene defect to clinical symptoms. These studies will also further our knowledge of Na channel function at the molecular level, will lead to an improved understanding of the determinants of muscle excitability, will provide mechanistic insights for a more rational design of therapeutic strategies, and will serve as a model system for understanding more common disorders of excitability such as epilepsy or cardiac arrhythmia.