MECHANISMS OF EXERCISE INDUCED MUSCLE FIBER INJURY

这项研究的目的是了解 运动时肌膜的短暂撕裂可能导致长期的 肌肉细胞内的损伤。杜氏肌营养不良症的肌肉， 最初的病变是短暂的肌膜 撕裂，表现出与离心运动相似的超微结构损伤。 杜氏营养不良的肌肉发展更大的活动，钙特异性 泄漏通道，这反过来又负责提高休息 细胞内游离钙（[Ca 2 +]i）和更高的钙- 依赖性蛋白水解我们假设，在运动过程中， 钙离子通过短暂的泪液流入导致局部激活 钙通过蛋白水解渗漏通道。最终，积累 激活的渗漏通道将增加钙内流，静息[Ca 2 +]i， 和钙依赖性降解途径的激活。瞬态 肌膜在收缩活动中撕裂，特别是在营养不良的 肌肉，可能代表激活过程的初始步骤 最终导致肌肉细胞死亡 本研究将通过以下问题来检验这一假设： (1)[Ca 2 +]i的时空变化是什么？ 严格定义的，可重复的伤害事件(2)受伤会导致 局部激活泄漏通道，如果是这样，什么变化（Ca 2 +]i 需要激活通道吗是伤口引起的泄漏通道 激活介导的蛋白水解和细胞骨架的影响？ (3)泄漏通道的活性是如何被蛋白水解或 在切除的斑块中操纵细胞骨架？(4)是激活 收缩活动后的钙依赖性降解过程 依赖于钙通过更活跃的渗漏通道流入？（提问 1)培养的小鼠骨骼肌管上的精确创伤将被追踪 通过测量[Ca 2 +]i的空间和时间变化。（问题2） [Ca 2 +]i变化的知识将被用来确定 使用贴片时钙内流与通道激活之间的关系 钳位法（问题3）通过以下方式调节通道活动： 蛋白水解和调节的细胞骨架将检查使用 切除的由内而外的膜片钳配置。（问题4）Myotubes将 经受长时间的收缩活动或静止， 泄漏通道的二氢吡啶抑制剂与抑制相关 渗漏通道的钙依赖性酶活性降低 流程.

The goal of this research is to understand the mechanisms by which transient tears at the sarcolemma during exercise might lead to long-term damage within a muscle cell. Muscles in Duchenne muscular dystrophy, in which the initial lesion is greater levels of transient sarcolemmal tearing, exhibit similar ultrastructural damage as eccentric exercise. Duchenne dystrophic muscles develop greater activity of calcium-specific leak channels which in turn are responsible for elevated resting intracellular free calcium ([Ca2+]i) and higher rates of calcium- dependent proteolysis. We hypothesize that, during exercise, local calcium influx through transient tears leads to local activation of calcium leak channels via proteolysis. Eventually, accumulation of activated leak channels will increase calcium influx, resting [Ca2+]i, and activation of calcium-dependent degradative pathways. Transient sarcolemmal tears during contractile activity, especially in dystrophic muscle, may represent the initial step in the activation of processes which eventually mediate muscle cell death. This study will examine this hypothesis through the following questions: (1) What are the spatial and temporal changes in [Ca2+]i near a rigorously defined, reproducible wounding event? (2) Does wounding lead to local activation of leak channels, and, if so, what changes in (Ca2+]i are required to activate channels? Is wound-induced leak channel activation mediated by proteolysis and by effects on the cytoskeleton? (3) How is activity of the leak channel modulated by proteolysis or manipulation of the cytoskeleton in excised patches? (4) Is activation of calcium-dependent degradative processes after contractile activity dependent on calcium influx through more active leak channels? (Question 1) Precise wounds on cultured mouse skeletal myotubes will be followed by measurement of spatial and temporal changes in [Ca2+]i. (Question 2) Knowledge of [Ca2+]i changes will then be employed to determine the relationship between calcium influx and channel activation using patch clamp methods. (Question 3) The regulation of channel activity by proteolysis and modulation of the cytoskeleton will be examined using the excised inside-out patch clamp configuration. (Question 4) Myotubes will be subject to long periods of contractile activity or quiescence and dihydropyridine inhibitors of the leak channel to correlate inhibition of leak channels with decreased rates of calcium-dependent enzymatic processes.