Molecular Physiology of Myotonia and Periodic Paralysis

肌强直和周期性麻痹的分子生理学

• 批准号: 7820641
• 负责人: STEPHEN C. CANNON
• 金额: $ 49.91万
• 依托单位: UT SOUTHWESTERN MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-09-28 至 2011-09-27
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7820641
• 关键词: Accounting; Action Potentials; Activities of Daily Living; Address; Adult; Affect; Age; American; Animal Model; Arginine; Arts; Behavior; Breeding; Calcium Channel; Capital; Carbonic Anhydrase Inhibitors; Charge; Chloride Channels; Computer Simulation; Critiques; Defect; Dependence; Development; Disease; Dose; Economics; Employment; Environment; Equilibrium; Equipment; Exposure to; Extravasation; Failure; Family; Fiber; Frequencies; Functional disorder; Funding; Gated Ion Channel; Gender; Gene Dosage; Generations; Genes; Genetically Engineered Mouse; Glucose; Goals; Grant; Human; Hypokalemia; Hypokalemic periodic paralysis; In Vitro; Individual; Inherited; Insulin; Intravenous infusion procedures; Investigation; Ions; Knock-in Mouse; Laboratories; Lead; Life Expectancy; Link; Measures; Membrane Potentials; Missense Mutation; Modeling; Molecular; Monovalent Cations; Morbidity - disease rate; Mus; Muscle; Muscle Contraction; Muscle Fibers; Mutant Strains Mice; Mutation; Myopathy; Myotonia; Optical Methods; Paralysed; Pathogenesis; Pathologic; Pathway interactions; Patients; Penetrance; Performance; Phenotype; Physiological; Physiology; Pilot Projects; Point Mutation; Positioning Attribute; Postdoctoral Fellow; Potassium Channel; Predisposition; Preparation; Productivity; Property; Protons; Recommendation; Recovery; Reporting; Role; Schools; Severities; Skeletal Muscle; Sodium Channel; Stimulus; Symptoms; System; Testing; Therapeutic Intervention; Tubular formation; United States National Institutes of Health; Work; base; clinical phenotype; depressed; gain of function mutation; graduate student; hyperkalemia; in vivo; interest; ionic balance; loss of function; mathematical model; meetings; mouse model; mutant; mutant mouse model; novel strategies; novel therapeutics; public health relevance; response; sensor; voltage; voltage clamp

DESCRIPTION (provided by applicant): This application is submitted in response to Notice Number NOT-OD-09-058: "NIH Announces the Availability of Recovery Act Funds for Competitive Revision Applications". The proposed studies expand the scope of the original application (R37-AR42703), by extending our studies on the pathomechanism of periodic paralysis to include Ca channel (CaV1.1) mutations responsible for hypokalemic periodic paralysis (HypoPP). The original application was focused exclusively on sodium channel (NaV1.4) defects in myotonia and periodic paralysis. The expanded studies in this competitive revision were not part of the original proposal. More specifically, the new proposed studies are not a line of investigation that failed to meet approval for recommendation of support by prior Scientific Review Group critique. HypoPP is the most common form of inherited periodic paralysis and is a significant cause of morbidity and lost productivity. We are excited to have successfully generated a knock-in CaV1.1 mutant mouse model of HypoPP, which in preliminary studies has a clear phenotype. This is the only animal model of HypoPP created to date and offers a unique opportunity to understand disease mechanism and to test therapeutic interventions. This project is not supported by any active grant to our laboratory, and support from NOT-OD-09-058 will be vital to continue to project in preparation for eventual submission of a new grant (R0-1). The proposed studies will comply with the economic objectives of the Recovery Act by creating 2.5 new positions of employment (postdoc, graduate student, technician) and adding new capital equipment to accelerate the pace of discovery on this project. PUBLIC HEALTH RELEVANCE: 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 fiber. The long-term goals of the original project (AR42703) were to characterize the functional defects of mutant channels in these disorders and to determine how abnormal channel activity produces symptoms in affected individuals. In these disorders, muscle dysfunction is caused by intermittent derangements in the electrical excitability of the fiber, which may be pathologically enhanced or depressed. Myotonia is a disorder of enhanced excitability wherein a single stimulus elicits a high-frequency burst of action potentials that produces involuntary persistent muscle contraction lasting seconds. Conversely, periodic paralysis results from a depolarization-induced loss of muscle excitability. Mutations of sodium channels (NaV1.4), chloride channels (ClC-1), K channels (Kir2.1), or Ca channels (CaV1.1) are established causes of myotonia and periodic paralysis in humans. The original project was focused exclusively on mutations in the adult skeletal muscle sodium channel (NaV1.4), to address the very interesting mechanistic question of how a point mutation in a single gene may cause myotonia, periodic paralysis, or a combination of both in the same individual. In this revised application, the scope of the project will be expanded to include an investigation of the mechanism by which mutations in the L-type Ca channel (CaV1.1) produce susceptibility to Hypokalemic Periodic Paralysis (HypoPP). The scientific approach is based on a combination of physiological studies in a knock-in mutant mouse model of CaV1.1-HypoPP, expression studies of disease-associated mutant channels, and mathematical modeling of muscle fiber excitability. Impact / Significance of the Revised Studies. While life-expectancy is normal for patients with periodic paralysis, it is a disabling condition that severely impacts performance at school or work, and interferes with activities of daily living. Tremendous advances have been gained in understanding the molecular defects associated with the periodic paralyses, as prototypical ion channelopathies [3, 11, 39]. The mechanistic link between altered channel function and loss of sarcolemmal excitability during an attack of weakness, however, is only partially understood for NaV1.4 mutations [2] and remains a complete mystery for CaV1.1 mutations in HypoPP. Heterologous expression studies have revealed biophysical defects of mutant CaV1.1 channels, but these changes do not readily provide an explanation for depolarization and paralysis. Moreover, the impact of these functional channel defects on sarcolemmal excitability has been difficult to ascertain experimentally, due to the scarcity of human HypoPP muscle suitable for study and the lack of any spontaneous animal model. Our genetically-engineered mouse model with a knock-in R528H mutation provides an outstanding opportunity to study the behavior of mutant CaV1.1 channels expressed in a muscle environment, to experimentally define the mechanism by which Vrest is aberrantly depolarized to cause weakness, and to explore the mode of action by which carbonic anhydrase inhibitors reduce the severity and frequency of attacks. In addition to elucidating disease pathogenesis, these studies may reveal new roles for CaV1.1 channels in maintaining Vrest and may provide a system for the rational development and testing of new therapeutic strategies to alleviate attacks of periodic paralysis. The proposed studies will comply with the economic objectives of the American Recovery and Reinvestment Act and the request for revised applications (NOT-OD-09-058) by creating 2.5 new positions of employment (postdoc, graduate student, technician) and adding new capital equipment to accelerate the pace of discovery on this project.

描述(由申请人提供)：本申请是对NOT-OD-09-058号通知的回应：“美国国立卫生研究院宣布恢复法资金可用于竞争性修订申请”。建议的研究扩大了最初的应用范围(R37-AR42703)，扩展了我们对周期性麻痹发病机制的研究，包括导致低钾性周期性麻痹(HypoPP)的钙通道(CaV1.1)突变。最初的应用仅集中在钠通道(NaV1.4)缺陷的肌强直和周期性瘫痪上。这一竞争性修订中的扩展研究不是最初提案的一部分。更具体地说，新提出的研究并不是一项未能获得科学审查小组先前批评意见支持的建议的调查。低PP是遗传性周期性麻痹最常见的形式，也是发病率和生产力丧失的重要原因。我们很高兴成功地建立了一个敲入CaV1.1突变的HypoPP小鼠模型，在初步研究中，该模型具有明确的表型。这是迄今为止创建的唯一的HypoPP动物模型，为了解疾病机制和测试治疗干预措施提供了独特的机会。该项目没有得到我们实验室任何正在进行的赠款的支持，来自NOT-OD-09-058的支持对于继续项目准备最终提交新的赠款(R0-1)至关重要。拟议的研究将符合复苏法案的经济目标，创造2.5个新的就业岗位(博士后、研究生、技术员)，并增加新的资本设备，以加快该项目的发现步伐。 公共卫生相关性：肌强直和周期性瘫痪是骨骼肌的遗传性疾病，电压门控离子通道的突变会改变纤维的电兴奋性。最初项目(AR42703)的长期目标是表征这些疾病中突变通道的功能缺陷，并确定异常通道活动如何在受影响的个体中产生症状。在这些疾病中，肌肉功能障碍是由纤维的电兴奋性的间歇性紊乱引起的，这可能是病理上的增强或抑制。肌强直是一种兴奋性增强的疾病，单个刺激引起动作电位的高频爆发，产生持续数秒的非自愿持续性肌肉收缩。相反，周期性瘫痪是去极化导致肌肉兴奋性丧失的结果。钠通道(NaV1.4)、氯通道(ClC-1)、钾通道(Kir2.1)或钙通道(CaV1.1)的突变是人类肌强直和周期性瘫痪的主要原因。最初的项目专门关注成人骨骼肌钠通道(NaV1.4)的突变，以解决非常有趣的机制问题，即单个基因点突变如何导致肌强直、周期性瘫痪或同一个体两者的组合。在这个修订的应用中，项目的范围将扩大到包括对L类型钙通道(CaV1.1)突变导致低血钾性周期性麻痹(HypoPP)易感性的机制的调查。这一科学方法基于CaV1.1-HypoPP敲入突变小鼠模型的生理学研究、疾病相关突变通道的表达研究以及肌肉纤维兴奋性的数学模型。修订研究的影响/意义。虽然周期性瘫痪患者的预期寿命是正常的，但这是一种严重影响学习或工作表现并干扰日常生活活动的残疾状况。在理解与周期性麻痹相关的分子缺陷方面取得了巨大的进展，作为典型的离子通道病[3，11，39]。然而，对于NaV1.4突变[2]，在虚弱发作期间，通道功能改变和肌膜兴奋性丧失之间的机制联系只被部分理解，并且对于HypoPP中的CaV1.1突变仍然是一个完全的谜。异源表达研究揭示了突变型CaV1.1通道的生物物理缺陷，但这些变化并不能很容易地解释去极化和瘫痪。此外，这些功能通道缺陷对肌膜兴奋性的影响很难通过实验来确定，因为适合研究的人下PP肌很少，而且缺乏任何自发的动物模型。我们的带有敲入R528H突变的转基因小鼠模型提供了一个绝佳的机会，可以研究肌肉环境中表达的突变CaV1.1通道的行为，从实验上确定Vrest异常去极化导致虚弱的机制，并探索碳酸酐酶抑制剂降低发作严重程度和频率的作用模式。除了阐明疾病的发病机制外，这些研究还可能揭示CaV1.1通道在维持Vrest中的新作用，并可能为合理开发和测试缓解周期性麻痹发作的新治疗策略提供一个系统。拟议的研究将通过创造2.5个新的就业岗位(博士后、研究生、技术员)和增加新的资本设备来加快该项目的发现步伐，从而符合美国复苏和再投资法案的经济目标和修订申请的请求(非-OD-09-058)。