Regulation of Calcium Release by Calmodulin in Muscle

描述(申请人提供)：骨骼肌肌浆网(SR)钙(Ca~(2+))通过兰尼定受体(RyR)钙释放通道的变化已在许多肌肉疾病中被发现，包括恶性高热和中枢性疾病。内源性配体对肌浆网钙离子释放的调节在这些病变肌肉中也发生了改变。这表明了解内源调节剂对RyR活性的调控具有重要意义。利用激光扫描共聚焦显微镜和钙指示剂Fluo-3的技术，可以对功能完整的骨骼肌中离散的局部钙释放事件(钙火花)进行成像。对这些事件在电压或配体刺激下的特性和发生频率的研究有助于深入了解肌纤维功能的背景下SR钙释放的分子控制。 本研究的主要目的是研究功能完整的骨骼肌纤维中钙调素对肌浆网钙离子释放的局部调控作用。具体地说，我打算研究钙调蛋白对自发和电压激活的骨骼肌钙离子触发特性的影响，以深入了解钙调蛋白在骨骼肌兴奋收缩偶联中所起的作用。使用重组钙调蛋白，包括野生型和突变体，不能结合钙离子，将允许评估钙调蛋白在这些离散的局部钙释放事件的激活、增殖和终止中所起的作用。

DESCRIPTION (provided by applicant): Alterations in skeletal muscle sarcoplasmic reticulum (SR) calcium (Ca2+) release via the ryanodine receptor (RYR) Ca2+ release channel have been identified in a number of muscle diseases including malignant hyperthermia and central core disease. The regulation of SR Ca2+ release by endogenous ligands is also altered in these diseased muscles. This signifies the importance of understanding the regulation of RYR activity by endogenous modulators. Techniques utilizing laser scanning confocal microscopy and the Ca2+ indicator Fluo-3 have allowed the imaging of discrete localized Ca2+ release events (Ca2+ sparks) in functionally intact skeletal muscle. Examination of properties and frequency of occurrence of these events in response to either voltage or ligand stimulation is providing insight into the molecular control of SR Ca2+ release within the context of the functioning muscle fiber. The main objective of this proposal is to examine the modulation of the local control of SR Ca2+ release by calmodulin in functionally intact skeletal muscle fibers. Specifically, I intend to examine the effects of calmodulin on spontaneous and voltage activated skeletal muscle Ca2+ spark properties to gain insight into the role calmodulin plays in skeletal muscle excitation contraction coupling. The use of recombinant calmodulin, both wild type and a mutant that cannot bind Ca2+, will allow for the assessment of the role calmodulin plays in the activation, propagation and termination of these discrete localized Ca2+ release events.