Study for the Abnormality of Intracellular Ca^<2+> Regulation in Failing Hearts

在心肌细胞中，钙诱导的肌浆网（SR）钙释放在钙瞬态中起关键作用。FK-506结合蛋白（FKBP）结合SR - Ca释放通道（ryanodine receptor: RyRs），稳定SR - Ca释放通道门控。最近的双分子层研究表明，免疫抑制剂FK506可以将FKBP与RyRs分离，从而增加SR Ca泄漏。虽然FKBP位移是心力衰竭的一个重要方面，但其与生理钙调节的相关性仍不明确。我们利用激光扫描共聚焦显微镜和fluo-3检测了FK506对大鼠心室肌细胞稳态（SS）抽搐Ca瞬态（CaT）、休息后（PR）咖啡因诱导的CaT （SR Ca含量指标）和Ca火花（SR Ca泄漏指标）的影响。(1) FK506 （50 μM）可增加静息期SS （0.5Hz）抽动CaT振幅和Ca火花频率，但对PR抽动CaT和咖啡因CaT无明显影响。(2)在分别用thapsigargin和低细胞外Ca浓度处理的肌细胞中，FK506没有改变SS抽搐和PR咖啡因CaT；(3)在同时用thapsigargin和低细胞外Ca浓度处理的肌细胞中，SS抽搐CaT和PR咖啡因CaT大量降低，FK506显著加速了这种降低。综上所述，由RyRs中FKBP的移位引起的SR Ca泄漏可能不足以干扰完整肌细胞正常的兴奋-收缩偶联。然而，当SR Ca摄取减少，肌层Ca挤压时，位移会导致SR Ca的明显损失。

In cardiac myocytes the Ca-induced Ca release from the sarcoplasmicr reticulum (SR) plays pivotal roles in Ca transients. FK-506 binding protein (FKBP) binds to the SR Ca release channels (ryanodine receptors : RyRs) and stabilizes the SR Ca release channel gating. Recent bilayer studies showed that an immunosuppressant agent, FK506 can dissociate FKBP from RyRs, thereby increasing SR Ca leak. Although the displacement of FKBP is an important aspect in failing heart, its relevance to physiological Ca regulation is still undefined. We examined the effects of FK506 on steady-state (SS) twitch Ca transients (CaT), post rest (PR) caffeine-induced CaT (as an index of SR Ca content) and Ca sparks (as an index of SR Ca leak) using laser scanning confocal microscopy and fluo-3 in rat ventricular myocytes. (1) In intact myocytes, FK506 (50 μM) increased SS (0.5Hz) twitch CaT amplitude and Ca spark frequency during rest periods, but did not change the PR twitch- and caffeine- CaT. (2) In myocytes treated either with thapsigargin and low extracellular Ca concentration, FK506 did not change the SS twitch and PR caffeine CaTs, and (3) In myocytes treated with both thapsigargin and low extracellular Ca concentration, SS twitch CaT and PR caffeine CaT largely reduced, and FK506 significantly accelerated the reduction. In conclusion, the SR Ca leak by displacement of FKBP from RyRs may not be enough to disturb normal excitation-contraction coupling in intact myocytes. However, when the SR Ca uptake was reduced and the sarcolemmal Ca extrusion was, the displacement could induce a significant SR Ca loss.