Isoform-specific roles of the Na,K-ATPase in skeletal muscle

描述(由申请人提供)：骨骼肌无力和疲劳是心力衰竭、骨质疏松症、恶病质、肌肉营养不良、慢性阻塞性肺疾病和其他疾病的一种常见的、使人衰弱的情况。Na，K-ATPase活性在肌肉收缩过程中被极大地刺激，是维持力量和运动表现所必需的。然而，很少有研究研究Na，K-ATPase亚型在肌肉无力和疲劳中的作用。成人骨骼肌主要表达Na，K-ATPase的？2亚型，而大多数其他组织主要表达？1亚型。？2亚型在骨骼肌中的作用以及刺激其活性的机制尚不清楚。这项研究的中心假设是，Na，K-ATPase？2亚型提供了一种储备能力，这种储备能力在静止时大部分不活跃，但在收缩时被迅速刺激，以满足对Na/K运输增加的需求；它的调节部分是通过肌肉特异性Na，K-ATPase的FXYD1亚单位的磷酸化实现的。这一假设将通过一种新的靶向基因小鼠进行验证，这种小鼠在成年骨骼肌(SK？？2-/-)中特别缺乏Na，K-ATPase？2，并表现出明显的运动耐受性和骨骼肌无力。其具体目的是：1)确定Na，K-ATPase？2酶在维持力量和运动能力中的急性作用；2)确定Na，K-ATPase？2酶在横管膜兴奋和抗疲劳中的作用；以及3)确定FXYD1磷酸化在收缩过程中通过？2-肾上腺素能受体激活而急性刺激Na，K-ATPase？2活性的作用；b，确定改变的Na，K-ATPase内容或功能对心力衰竭时肌肉疲劳和运动性耐受的贡献。这些目标将使用一种综合策略来实现，该策略将小鼠的遗传操作与从动物到细胞和亚细胞水平的功能测量相结合。总而言之，该项目将促进我们对Na，K-ATPase？2亚型在骨骼肌中的生理作用和调节机制的理解，并为肌肉无力和疲劳的治疗干预确定新的分子靶点。

DESCRIPTION (provided by applicant): Skeletal muscle weakness and fatigue is a common, debilitating condition in heart failure, sarcopenia, cachexia, muscular dystrophies, COPD, and other disorders. Na,K-ATPase activity is dramatically stimulated during muscle contraction and is absolutely required to maintain force and exercise performance. However, few studies have examined the role the Na,K-ATPase isoforms in muscle weakness and fatigue. Adult skeletal muscles express mainly the ?2 isoform of the Na,K-ATPase, in contrast to most other tissues which express mainly the ?1 isoform. The role of the ?2 isoform in skeletal muscle and the mechanisms by which its activity is stimulated are not known. The central hypotheses of this research is that the Na,K-ATPase ?2 isoform provides a reserve capacity which is largely inactive at rest but is rapidly stimulated during contraction, to meet the increased demand for Na/K transport; and that its regulation is achieved in part by phosphorylation of the muscle-specific FXYD1 subunit of the Na,K-ATPase. This hypothesis will be tested using a novel gene targeted mouse which specifically lacks the Na,K-ATPase ?2 in adult skeletal muscles (sk??2-/-) and shows marked exercise intolerance and skeletal muscle weakness. The Specific Aims are to: 1) Determine the acute role of the Na,K-ATPase ?2 enzyme in maintaining force and exercise performance; 2) Determine the role of the Na,K-ATPase ?2 enzyme in membrane excitation in the transverse tubules and resistance to fatigue; and 3) a, Define the role of FXYD1 phosphorylation in the acute stimulation of Na,K-ATPase ?2 activity by ?2- adrenergic receptor activation during contraction; b, Determine the contribution of altered Na,K-ATPase content or function to muscle fatigue and exercise intolerance in heart failure. These Aims will be approached using an integrated strategy which combines genetic manipulations in the mouse with functional measurements from the animal to the cellular and subcellular level. Collectively, this project will advance our understanding of the physiological roles of the Na,K-ATPase ?2 isoform and the mechanisms by which it is regulated in skeletal muscle, and identify new molecular targets for therapeutic interventions in muscle weakness and fatigue.