MOLECULAR PHYSIOLOGY OF NEUROMUSCULAR DISEASES

许多遗传性肌肉疾病是由 肌膜高钾性周期性麻痹的电兴奋性 (HPP)虚弱发作与海拔高度相关 胞外钾。在发作过程中，肌肉去极化， 电不能兴奋的。一种相关疾病--先天性副肌强直 (PMC)，特点是局部冷致僵直(肌强直)和 轻微的虚弱。肌强直是由重复的后放电引起的 起源于受影响的肌肉，独立于神经元输入。 生理学和遗传学证据的结合已经证实 HPP、PMC和马形周期性瘫痪都是由 成人骨骼肌异构体α亚基突变的研究 钠离子通道。我们在前面已经证明了主函数 HPP的缺陷是钠电流失活的破坏。损失的损失 细胞外信号增强人HPP肌管的失活 [K].这为袭击的间歇性性质提供了解释， 但在生物物理学上是出乎意料的。这项建议的一个主要目的是 确定细胞外K是否直接改变突变的门控 渠道，探索K如何发挥其影响，并阐明 持续钠流的动力学基础。钠的门控行为 从未在对温度敏感的表型中测量到通道， PMC。由PMC突变产生的功能缺陷将通过以下方式定义 突变的cDNA在哺乳动物细胞中的异源表达。多个PMC 突变发生在III-IV细胞质环中，额外的位点- 将执行定向突变来定义该结构域如何 参与失活过程。异常的钠通道 HPP和PMC中的行为将被合并到计算机中 模拟和动物模型以探索其病理生理学基础 这些表型的显性表达。在马匹周期性麻痹中 所有受影响的动物在阿尔法亚单位都有相同的点突变， 苯妥英钠还可以减少发作的频率。么正钠流 将从马的肌管中记录以确定功能缺陷和 测定苯妥英钠对异常通道门控的影响。 建议的研究旨在提供一个完整的了解 人类两种神经肌肉疾病的分子生理学基础。在……里面 此外，这些结果将进一步加深我们对钠离子通道的理解 在分子水平上的作用，将提供对 为这些疾病设计合理的治疗方法，并将作为典范 一种用于了解改变的电的其他障碍的系统 兴奋性(癫痫、心律失常)。

Many inherited disorders of muscle are caused by an abnormality in the electrical excitability of the sarcolemmal hyperkalemic periodic paralysis (HPP) episodes of weakness occur in association with an elevation in extracellular potassium. During an attack, muscles are depolarized and electrically inexcitable. A related disorder, paramyotonia congenita (PMC), is characterized by localized cold-induced stiffness (myotonia) and mild weakness. Myotonia arises from repetitive after-discharges that originate in affected muscle independent from neuronal input. A combination of physiologic and genetic evidence has established that HPP, PMC and an equine form of periodic paralysis are all caused by mutations in the alpha subunit of the adult skeletal muscle isoform of the sodium channel. We have shown previously that the primary functional defect in HPP is a disruption of Na current inactivation. The loss of inactivation in human HPP myotubes was enhanced by raised extracellular [K]. This provided an explanation for the episodic nature of the attacks, but was unexpected biophysically. A major aim of this proposal is to determine whether extracellular K directly alters gating in mutant channels, to explore how K exerts its influence, and to elucidate the kinetic basis for the persistent Na current. Gating behavior of Na channels has never been measured in the temperature-sensitive phenotype, PMC. The functional defects produced by PMC mutations will be defined by heterologous expression of mutant cDNAs in mammalian cells. Many PMC mutations occur in the III-IV cytoplasmic loop, and additional site- directed mutagenesis will be performed to define how this domain participates in the process of inactivation. The aberrant Na channel behaviors in HPP and PMC will be incorporated into both a computer simulation and an animal model to explore the pathophysiologic basis for the dominant expression of these phenotypes. In equine periodic paralysis all affected animals have the same point mutation in the alpha subunit, and the frequency of attacks is reduced by phenytoin. Unitary Na currents will be recorded from equine myotubes to define the functional defect and to measure the effects of phenytoin on aberrant channel gating. The proposed studies are designed to provide a complete understanding of the molecular physiologic basis of two human neuromuscular diseases. In addition, these results will further our understanding of Na channel function at the molecular level, will provide insights from which to design rational therapy for these diseases, and will serve as a model system for understanding other disorders of altered electrical excitability (epilepsy, cardiac dysrhythmias).