Université Lyon 1, CNRS UMR 5534, Centre de Génétique et de Physiologie Moléculaires et Cellulaires, 43 boulevard du 11 novembre 1918, 69622 Villeurbanne cedex, France.
J Physiol. 2012 Dec 1;590(23):6027-36. doi: 10.1113/jphysiol.2012.237321. Epub 2012 Sep 24.
Contraction of skeletal muscle is triggered by the release of Ca(2+) from the sarcoplasmic reticulum (SR) in response to depolarization of the muscle membrane. Depolarization is known to elicit a conformational change of the dihydropyridine receptor (DHPR) in the tubular membrane that controls in a time- and voltage-dependent manner the opening of the ryanodine receptor (RyR), the SR Ca(2+) release channel. At rest, it is assumed that RyRs are kept in a closed state imposed by the repressive action of DHPRs; however, a direct control of the RyR gating by the DHPR has up to now never been demonstrated in resting adult muscle. In this study, we monitored slow changes in SR Ca(2+) content using the Ca(2+) indicator fluo-5N loaded in the SR of voltage-clamped mouse muscle fibres. We first show that external Ca(2+) removal induced a reversible SR Ca(2+) efflux at -80 mV and prevented SR Ca(2+) refilling following depolarization-evoked SR Ca(2+) depletion. The dihydropyridine compound nifedipine induced similar effects. The rate of SR Ca(2+) efflux was also shown to be controlled in a time- and voltage-dependent manner within a membrane potential range more negative than -50 mV. Finally, intracellular addition of ryanodine produced an irreversible SR Ca(2+) efflux and kept the SR in a highly depleted state following depolarization-evoked SR Ca(2+) depletion. The fact that resting SR Ca(2+) efflux is modulated by conformational changes of DHPRs induced by external Ca(2+), nifedipine and voltage demonstrates that DHPRs exert an active control on gating of RyRs in resting skeletal muscle.
骨骼肌的收缩是由肌浆网（SR）中的 Ca²⁺释放触发的，这是对肌肉膜去极化的反应。众所周知，去极化会引起管状膜中二氢吡啶受体（DHPR）的构象变化，这种变化以时间和电压依赖的方式控制 Ryanodine 受体（RyR）的开放，即 SR Ca²⁺释放通道。在静息状态下，假设 RyR 处于由 DHPR 产生的抑制作用所强制的关闭状态；然而，DHPR 对 RyR 门控的直接控制在静止的成年肌肉中至今从未得到证明。在这项研究中，我们使用加载在电压钳制的小鼠肌肉纤维 SR 中的 Ca²⁺指示剂 fluo-5N 来监测 SR Ca²⁺含量的缓慢变化。我们首先表明，在 -80 mV 时，去除外部 Ca²⁺会诱导可逆的 SR Ca²⁺外排，并防止去极化引起的 SR Ca²⁺耗竭后 SR Ca²⁺再填充。二氢吡啶化合物硝苯地平也会产生类似的效果。还表明，在膜电位范围更负于 -50 mV 的情况下，SR Ca²⁺外排的速率也以时间和电压依赖的方式受到控制。最后，细胞内添加 Ryanodine 会产生不可逆的 SR Ca²⁺外排，并在去极化引起的 SR Ca²⁺耗竭后使 SR 保持高度耗竭状态。事实证明，静息 SR Ca²⁺外排受外部 Ca²⁺、硝苯地平和电压诱导的 DHPR 构象变化调节，表明 DHPR 对静息骨骼肌中 RyR 的门控具有主动控制作用。
