MOLECULAR PHYSIOLOGY OF HYPERKALEMIC PERIODIC PARALYSIS

高钾血症周期性麻痹的分子生理学

• 批准号: 6393144
• 负责人: LAWRENCE J HAYWARD
• 金额: $ 13.43万
• 依托单位: UNIV OF MASSACHUSETTS MED SCH WORCESTER
• 依托单位国家: 美国
• 财政年份: 1997
• 资助国家: 美国
• 起止时间: 1997-08-01 至 2002-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6393144
• 关键词: animal genetic material tag; chloride channels; genetically modified animals; hyperkalemias; ion channel blocker; laboratory mouse; mathematical model; model design /development; muscle cells; muscle function; muscle metabolism; paralysis; point mutation; potassium channel; potassium ion; sodium channel; tissue /cell culture; voltage /patch clamp; voltage gated channel; animal genetic material tag; chloride channels; genetically modified animals; hyperkalemias; ion channel blocker; laboratory mouse; mathematical model; model design /development; muscle cells; muscle function; muscle metabolism; paralysis; point mutation; potassium channel; potassium ion; sodium channel; tissue /cell culture; voltage /patch clamp; voltage gated channel

Hyperkalemic periodic paralysis (HyperPP), paramyotonia congenita (PMC), and sodium channel myotonia (SCM), are dominantly inherited myotonic disorders caused by missense mutations in the skeletal muscle sodium (Na) channel. Na channels generate the muscle action potential and influence excitability of the muscle membrane through voltage-dependent mechanisms that regulate channel availability. The proposed study will investigate the pathophysiology of these diseases by (1) analyzing electrophysiological properties of mutant Na channel expressed in a myogenic cell line and (2) evaluating the physiological consequences of over-expressed wild type or mutant Na channels upon muscle function in a transgenic mouse model. Whole-cell Na currents will be measured from MM14 muscle cells transfected with tetrodotoxin-resistant versions of wild-type or mutant Na channel cDNAs to assess muscle-specific influences upon Na channel properties. In eight transgenic mouse lines, wild type or mutant Na channel gene propagation will be verified by Southern blotting, and specificity of expression will be quantitated by an RNAse protection assay. Electromyographic screening of mice for myotonia will be performed in the absence and presence of provokation by potassium loading, exercise or cooling. Muscle force and relaxation properties of intact muscle fibers in vitro will be measured under conditions of elevated extracellular potassium. Patch clamp analysis of Na currents from sarcolemmal membrane blebs will compare native channel properties with those measured previously in heterologous cells. Histological studies will look for evidence of progressive vacuolar myopathy. These studies will test whether introduction of mutant but not wild-type Na channels in mice is sufficient to induce HyperPP through a "gain of function" mechanism. Sodium channel blockers or other drugs will be compared for effectiveness in preventing abnormal muscle excitation or ameliorating muscle cell degeneration. Kinetic models will then integrate the behavior of muscle Na, K, and Cl channels to provide quantitative predictions for understanding Na channel function and the biophysical pathology of skeletal muscle in HyperPP and related myotonic disorders. The applicant is a board-certified neurologist who has obtained experience in molecular biology and electrophysiology during three years of a postdoctoral fellowship in the laboratories of Dr. Robert H. Brown, Jr. , and Dr. Stephen C. Cannon. The proposed research will establish the critical tolls needed for the applicant to become an independent investigator focusing on an expanding family of ion channel diseases.

高钾血性周期性瘫痪（HyperPP），Parmyotonia Congenita（PMC）和钠通道Myotonia（SCM）是 由错义引起的主要继承的肌疾病 骨骼肌钠（NA）通道中的突变。 na 频道产生肌肉动作潜力和影响 肌肉膜通过电压依赖性的兴奋性 调节通道可用性的机制。 拟议的研究 将通过（1）研​​究这些疾病的病理生理学 分析突变Na通道的电生理特性 在肌原细胞系中表达，（2）评估 过表达野生型或突变体的生理后果 Na通道在转基因小鼠模型中的肌肉功能。 将从MM14肌肉细胞中测量全细胞Na电流 用野生型或耐二毒素抗性版本转染的野生型或 突变的Na通道cDNA评估肌肉特异性影响 在NA通道属性上。 在八种转基因小鼠线中，野生 类型或突变的Na通道基因传播将通过 Southern印迹和表达的特异性将被定量 通过RNase保护测定法。 肌电图筛查 在不存在和存在的情况下，将进行肌瘤的小鼠 通过钾负荷，运动或冷却来挑衅。 肌肉 体外完整肌肉纤维的力和松弛特性将 在升高的细胞外钾条件下进行测量。 斑块夹的斑块分析肌膜膜的Na电流 BLEB将将天然通道特性与所测量的特性进行比较 以前在异源细胞中。 组织学研究将寻找 进行性液泡肌病的证据。 这些研究会 测试是否引入突变体但不引入野生型Na通道 在小鼠中足以通过“获得” 功能“机制。钠通道阻滞剂或其他药物 将比较预防异常肌肉的有效性 激发或改善肌肉细胞变性。 动力学 然后，模型将整合肌肉Na，K和Cl的行为 提供定量预测以理解NA的渠道 通道功能和骨骼肌的生物物理病理学 在HyperPP和相关的肌发电症中。 申请人是 获得董事会认证的神经科医生 分子生物学和电生理学三年 罗伯特·H（Robert H.）实验室的博士后奖学金 小布朗和斯蒂芬·C·坎农博士。 拟议的研究 将确定申请人成为所需的关键通行费 一个专注于不断扩展的家庭的独立调查员 离子通道疾病。