Kormos Anita, Nagy Norbert, Acsai Károly, Váczi Krisztina, Ágoston Szabina, Pollesello Piero, Levijoki Jouko, Szentandrássy Norbert, Papp Julius Gy, Varró András, Tóth András
Department of Pharmacology & Pharmacotherapy, University of Szeged, Dóm tér 12., 6722 Szeged, Hungary.
MTA-SZTE Research Group of Cardiovascular Pharmacology, Hungarian Academy of Sciences, Szeged, Hungary.
Eur J Pharmacol. 2014 Oct 5;740:539-51. doi: 10.1016/j.ejphar.2014.06.033. Epub 2014 Jun 27.
In this study we evaluated the effects of selective Na+/Ca2+ exchanger (NCX) inhibition by ORM-10103 on the [Ca2+]i transient (CaT), action potential (AP), and cell viability in isolated canine ventricular cardiomyocytes exposed to a simulated ischemia/reperfusion protocol performed either alone (modeling moderate low-flow ischemia) or with simultaneous strophantidine challenge (modeling more severe low-flow ischemia). CaTs were monitored using a Ca2+-sensitive fluorescent dye, APs were recorded by intracellular microelectrodes, and anaerobic shifts in cellular metabolism were verified via monitoring native NADH fluorescence. Simulated ischemia increased the NADH fluorescence, reduced the amplitudes of the AP and CaT and induced membrane depolarization. APs moderately shortened, CaTs prolonged. Diastolic [Ca2+]i ([Ca2+]iD) level increased significantly during ischemia and further elevated following strophantidine application. Reperfusion normalized the NADH level, the amplitude of the AP and duration of the [Ca2+]i transient, but only partially restored action potential triangulation and the amplitude of the CaT. [Ca2+]iD decreased in untreated, but further increased in strophantidine-treated cells. 10 µM ORM-10103 significantly reduced the ischemic [Ca2+]i raise in both untreated and strophantidine-treated cells. During reperfusion ORM-10103 decreased [Ca2+]i and eliminated its diastolic elevation in untreated and strophantidine-treated cardiomyocytes. Following the application of ORM-10103 the detrimental effect of ischemia/reperfusion on cell viability and the reperfusion-induced increase in AP and CaT variabilities were substantially reduced, but ischemia-induced shifts in AP morphology were barely influenced. In conclusion, selective NCX inhibition by ORM-10103 is highly effective against ischemia/reperfusion induced pathologic alterations in [Ca2+]i homeostasis, however, it fails to normalize untoward arrhythmogenic changes in AP morphology.
在本研究中，我们评估了ORM-10103选择性抑制钠/钙交换体（NCX）对分离的犬心室心肌细胞中钙离子瞬变（CaT）、动作电位（AP）和细胞活力的影响。这些心肌细胞暴露于单独进行的模拟缺血/再灌注方案（模拟中度低流量缺血）或同时进行毒毛花苷挑战的方案（模拟更严重的低流量缺血）。使用钙敏荧光染料监测CaT，通过细胞内微电极记录AP，并通过监测天然NADH荧光来验证细胞代谢中的无氧变化。模拟缺血增加了NADH荧光，降低了AP和CaT的幅度，并诱导了膜去极化。AP适度缩短，CaT延长。缺血期间舒张期钙离子浓度（[Ca2+]iD）水平显著升高，应用毒毛花苷后进一步升高。再灌注使NADH水平、AP幅度和钙离子瞬变持续时间恢复正常，但仅部分恢复动作电位三角化和CaT幅度。[Ca2+]iD在未处理的细胞中降低，但在毒毛花苷处理的细胞中进一步升高。10 μM ORM-10103显著降低了未处理和毒毛花苷处理细胞中缺血时的钙离子浓度升高。在再灌注期间，ORM-10103降低了钙离子浓度，并消除了未处理和毒毛花苷处理的心肌细胞中舒张期钙离子浓度的升高。应用ORM-10103后，缺血/再灌注对细胞活力的有害影响以及再灌注诱导的AP和CaT变异性增加得到了显著降低，但缺血诱导的AP形态变化几乎未受影响。总之，ORM-10103选择性抑制NCX对缺血/再灌注诱导的钙离子稳态病理改变具有高度有效性，然而，它未能使AP形态的不良致心律失常变化恢复正常。
