Identifying cellular mechanisms of zinc-induced relaxation in isolated cardiomyocytes.
Identifying cellular mechanisms of zinc-induced relaxation in isolated cardiomyocytes.
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DOI：
10.1152/ajpheart.00025.2013
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发表时间：
2013-09
期刊：
影响因子：
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通讯作者：
Tingting Yi;Jonathan S. Vick;M. Vecchio;K. Begin;S. Bell;R. Delay;B. Palmer
中科院分区：
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作者：
Tingting Yi;Jonathan S. Vick;M. Vecchio;K. Begin;S. Bell;R. Delay;B. Palmer
我们测试了心肌细胞收缩-舒张功能的几种分子和细胞机制，这些机制可以解释暴露于细胞外Zn(2+)时观察到的收缩功能降低和舒张功能增强的原因。在维持在 37°C、以 2 或 6 Hz 刺激并暴露于 32 μM Zn(2+) 或载体的离体大鼠和小鼠心肌细胞中监测收缩-舒张功能。使用 FluoZin-3 检测到的细胞内 Zn(2+) 在 3-5 分钟内浓度升至~13 nM。 Zn(2+) 暴露后，峰值肌节缩短显着减少，舒张期肌节长度延长。 Fura-2FF 检测到的细胞内 Ca(2+) 峰值在 Zn(2+) 暴露后降低。然而，反映肌浆网（SR）Ca（2+）-ATP酶（SERCA2a）活性的胞质Ca（2+）下降率和通过快速Na（+）诱导的Ca（2+）流出评估的Na（+）/Ca（2+）交换活性率不受Zn（2+）暴露的影响。在暴露于Zn(2+)的心肌细胞中，通过快速咖啡因暴露评估的SR Ca(2+)负荷减少～50%，通过全细胞膜片钳测量的L型钙通道内向电流减少～70%。此外，用50 μM Zn(2+)灌注心脏后，兰尼碱受体(RyR) S2808和受磷蛋白(PLB) S16/T17显着去磷酸化。化学剥皮心肌条的最大张力发展和细丝Ca(2+)敏感性不受Zn(2+)暴露的影响。这些发现表明，Zn(2+) 通过降低收缩期和舒张期细胞内 Ca(2+) 浓度来抑制心肌细胞收缩功能并增强舒张功能，这是由于竞争性抑制 Ca(2+) 通过 L 型钙通道流入、减少受磷蛋白去磷酸化导致的 SR Ca(2+) 负荷以及降低 SR Ca(2+) 泄漏。 通过 RyR 去磷酸化。使用低 Ca(2+) 亲和力 Fura-2FF 可能会阻止检测舒张期 Ca(2+) 和 SERCA2a 功能的变化。在 Zn(2+) 存在的情况下检测舒张 Ca(2+) 的其他策略对于未来的工作至关重要。
We tested several molecular and cellular mechanisms of cardiomyocyte contraction-relaxation function that could account for the reduced systolic and enhanced diastolic function observed with exposure to extracellular Zn(2+). Contraction-relaxation function was monitored in isolated rat and mouse cardiomyocytes maintained at 37°C, stimulated at 2 or 6 Hz, and exposed to 32 μM Zn(2+) or vehicle. Intracellular Zn(2+) detected using FluoZin-3 rose to a concentration of ∼13 nM in 3-5 min. Peak sarcomere shortening was significantly reduced and diastolic sarcomere length was elongated after Zn(2+) exposure. Peak intracellular Ca(2+) detected by Fura-2FF was reduced after Zn(2+) exposure. However, the rate of cytosolic Ca(2+) decline reflecting sarcoplasmic reticulum (SR) Ca(2+)-ATPase (SERCA2a) activity and the rate of Na(+)/Ca(2+) exchanger activity evaluated by rapid Na(+)-induced Ca(2+) efflux were unchanged by Zn(2+) exposure. SR Ca(2+) load evaluated by rapid caffeine exposure was reduced by ∼50%, and L-type calcium channel inward current measured by whole cell patch clamp was reduced by ∼70% in cardiomyocytes exposed to Zn(2+). Furthermore, ryanodine receptor (RyR) S2808 and phospholamban (PLB) S16/T17 were markedly dephosphorylated after perfusing hearts with 50 μM Zn(2+). Maximum tension development and thin-filament Ca(2+) sensitivity in chemically skinned cardiac muscle strips were not affected by Zn(2+) exposure. These findings suggest that Zn(2+) suppresses cardiomyocyte systolic function and enhances relaxation function by lowering systolic and diastolic intracellular Ca(2+) concentrations due to a combination of competitive inhibition of Ca(2+) influx through the L-type calcium channel, reduction of SR Ca(2+) load resulting from phospholamban dephosphorylation, and lowered SR Ca(2+) leak via RyR dephosphorylation. The use of the low-Ca(2+)-affinity Fura-2FF likely prevented the detection of changes in diastolic Ca(2+) and SERCA2a function. Other strategies to detect diastolic Ca(2+) in the presence of Zn(2+) are essential for future work.