In skeletal muscle (SM), inward Ca-currents have no apparent role in excitation-contraction coupling (e-c coupling), however the Ca-channel blocker can affect twitch and tetanic muscle in mammalian SM. Experiments were conducted to study how diltiazem (DLZ) facilitates e-c coupling and inhibits contraction. 1) In complete (EDL) muscle and single intact fibres, 0.03 mM DLZ causes twitch potentiation and decreases force during tetanic activity, with increased fatigue. 2) In split open fibres isolated from EDL fibres, DLZ inhibits sarcoplasmic reticulum (SR) Ca-loading in a dose-dependent manner and has a potentiating effect on caffeine-induced SR Ca-release. 3) In isolated light SR (LSR) vesicles, SERCA1 hydrolytic activity is not affected by DLZ up to 0.2 mM. However, ATP-dependent Ca-uptake was inhibited in a dose-dependent manner at a concentration where e-c coupling is changed. 4) The passive Ca-efflux from LSR was reduced by half with 0.03 mM diltiazem, indicating that SR leaking does not account for the decreased Ca-uptake. 5) The denaturation profile of the SERCA Ca-binding domain has lower thermal stability in the presence of DLZ in a concentration-dependent manner, having no effect on the nucleotide-binding domain. We conclude that the effect of DLZ on SM is exerted by crossing the sarcolemma and interacting directly with the SERCA Ca-binding domain, affecting SR Ca-loading during relaxation, which has a consequence on SM contractility. Diltiazem effect on SM could be utilized as a tool to understand SM e-c coupling and muscle fatigue.

中文翻译：

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Ca2+ 通道阻滞剂地尔硫卓引起的 SERCA-1 构象变化影响哺乳动物骨骼肌功能

在骨骼肌 (SM) 中，内向 Ca 电流在兴奋-收缩耦合 (ec 耦合) 中没有明显作用，但 Ca 通道阻滞剂可以影响哺乳动物 SM 中的抽搐和强直肌。进行实验来研究地尔硫卓 (DLZ) 如何促进 ec 偶联并抑制收缩。 1) 在完整 (EDL) 肌肉和单个完整纤维中，0.03 mM DLZ 会导致抽搐增强并降低强直活动期间的力量，并增加疲劳。 2) 在从 EDL 纤维中分离出的分裂开放纤维中，DLZ 以剂量依赖性方式抑制肌浆网 (SR) Ca 负载，并对咖啡因诱导的 SR Ca 释放具有增强作用。 3) 在分离的轻 SR (LSR) 囊泡中，SERCA1 水解活性不受高达 0.2 mM 的 DLZ 的影响。然而，在 ec 偶联改变的浓度下，ATP 依赖性 Ca 摄取以剂量依赖性方式受到抑制。 4) 使用 0.03 mM 地尔硫卓时，LSR 的被动 Ca 流出减少了一半，表明 SR 泄漏并不能解释 Ca 吸收的减少。 5) SERCA Ca结合结构域的变性特征在DLZ存在下以浓度依赖性方式具有较低的热稳定性，对核苷酸结合结构域没有影响。我们得出的结论是，DLZ 对 SM 的影响是通过穿过肌膜并直接与 SERCA Ca 结合域相互作用来发挥的，影响舒张过程中 SR Ca 的负载，从而对 SM 收缩性产生影响。地尔硫卓对 SM 的影响可以作为了解 SM ec 耦合和肌肉疲劳的工具。