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）钙释放在钙瞬变中起关键作用。FK-506结合蛋白（FKBP）与SR Ca释放通道（ryanodine受体：RyR）结合，并稳定SR Ca释放通道门控。最近的双层研究表明，免疫抑制剂FK 506可以使FKBP与RyR解离，从而增加SR Ca泄漏。虽然FKBP的移位是心力衰竭的一个重要方面，但其与生理性Ca调节的相关性仍不明确。我们使用激光扫描共聚焦显微镜和Fluo-3在大鼠心室肌细胞中检测了FK 506对稳态（SS）颤搐钙瞬变（CaT）、静息后（PR）咖啡因诱导的CaT（作为SR钙含量的指标）和钙火花（作为SR钙泄漏的指标）的影响。(1)在完整肌细胞中，FK 506（50 μM）可增加SS（2）在低钙和毒胡萝卜素处理的心肌细胞中，FK 506均不改变PR收缩和PR咖啡因收缩的CaTs。（3）在毒胡萝卜素和低钙处理的心肌细胞中，SS收缩CaT和PR咖啡因CaT显著降低，FK 506显著促进这种降低。总之，FKBP从RyRs置换引起的SR Ca泄漏可能不足以干扰完整心肌细胞的正常兴奋-收缩偶联。但当肌浆网钙摄取减少，肌膜钙排出减少时，移位可引起肌浆网钙丢失。

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.