Lang Di, Sato Daisuke, Jiang Yanyan, Ginsburg Kenneth S, Ripplinger Crystal M, Bers Donald M
From the Department of Pharmacology, University of California, Davis.
Circ Res. 2017 Dec 8;121(12):1379-1391. doi: 10.1161/CIRCRESAHA.117.312050. Epub 2017 Oct 2.
Intercellular uncoupling and Ca (Ca) mishandling can initiate triggered ventricular arrhythmias. Spontaneous Ca release activates inward current which depolarizes membrane potential (V) and can trigger action potentials in isolated myocytes. However, cell-cell coupling in intact hearts limits local depolarization and may protect hearts from this arrhythmogenic mechanism. Traditional optical mapping lacks the spatial resolution to assess coupling of individual myocytes.
We investigate local intercellular coupling in Ca-induced depolarization in intact hearts, using confocal microscopy to measure local V and intracellular [Ca] simultaneously.
We used isolated Langendorff-perfused hearts from control (CTL) and heart failure (HF) mice (HF induced by transaortic constriction). In CTL hearts, 1.4% of myocytes were poorly synchronized with neighboring cells and exhibited asynchronous (AS) Ca transients. These AS myocytes were much more frequent in HF (10.8% of myocytes, <0.05 versus CTL). Local Ca waves depolarized V in HF but not CTL hearts, suggesting weaker gap junction coupling in HF-AS versus CTL-AS myocytes. Cell-cell coupling was assessed by calcein fluorescence recovery after photobleach during intracellular [Ca] recording. All regions in CTL hearts exhibited faster calcein diffusion than in HF, with HF-AS myocyte being slowest. In HF-AS, enhancing gap junction conductance (with rotigaptide) increased coupling and suppressed V depolarization during Ca waves. Conversely, in CTL hearts, gap junction inhibition (carbenoxolone) decreased coupling and allowed Ca wave-induced depolarizations. Synchronization of Ca wave initiation and triggered action potentials were observed in HF hearts and computational models.
Well-coupled CTL myocytes are effectively voltage-clamped during Ca waves, protecting the heart from triggered arrhythmias. Spontaneous Ca waves are much more common in HF myocytes and these AS myocytes are also poorly coupled, enabling local Ca-induced inward current of sufficient source strength to overcome a weakened current sink to depolarize V and trigger action potentials.
细胞间解偶联和钙（Ca）处理不当可引发触发型室性心律失常。自发钙释放激活内向电流，使膜电位（V）去极化，并可在分离的心肌细胞中触发动作电位。然而，完整心脏中的细胞间耦合限制了局部去极化，并可能保护心脏免受这种致心律失常机制的影响。传统光学映射缺乏评估单个心肌细胞耦合的空间分辨率。
我们使用共聚焦显微镜同时测量局部V和细胞内[Ca]，研究完整心脏中钙诱导去极化时的局部细胞间耦合。
我们使用了来自对照（CTL）和心力衰竭（HF）小鼠（经主动脉缩窄诱导的HF）的离体Langendorff灌注心脏。在CTL心脏中，1.4%的心肌细胞与相邻细胞同步性差，表现出异步（AS）钙瞬变。这些AS心肌细胞在HF中更为常见（占心肌细胞的10.8%，与CTL相比P<0.05）。局部钙波使HF心脏中的V去极化，但CTL心脏中未出现，这表明HF-AS心肌细胞与CTL-AS心肌细胞相比，缝隙连接耦合较弱。在细胞内[Ca]记录期间，通过光漂白后钙黄绿素荧光恢复来评估细胞间耦合。CTL心脏的所有区域钙黄绿素扩散都比HF快，其中HF-AS心肌细胞最慢。在HF-AS中，增强缝隙连接电导（使用rotigaptide）可增加耦合，并抑制钙波期间的V去极化。相反，在CTL心脏中，缝隙连接抑制（使用羧苄青霉素）会降低耦合，并使钙波诱导的去极化发生。在HF心脏和计算模型中观察到钙波起始和触发动作电位的同步。
耦合良好的CTL心肌细胞在钙波期间有效地进行电压钳制，保护心脏免受触发型心律失常的影响。自发钙波在HF心肌细胞中更为常见，并且这些AS心肌细胞耦合也较差，使得局部钙诱导的内向电流具有足够的源强度，以克服减弱的电流汇使V去极化并触发动作电位。
