Laboratory of Cardiovascular Science, National Institute on Aging, National Institutes of Health, Baltimore, MD, USA.
Department of Medicine, Krannert Institute of Cardiology, Indianapolis, IN, USA.
J Mol Cell Cardiol. 2014 Jan;66:106-15. doi: 10.1016/j.yjmcc.2013.11.011. Epub 2013 Nov 22.
Basal phosphorylation of sarcoplasmic reticulum (SR) Ca(2+) proteins is high in sinoatrial nodal cells (SANC), which generate partially synchronized, spontaneous, rhythmic, diastolic local Ca(2+) releases (LCRs), but low in ventricular myocytes (VM), which exhibit rare diastolic, stochastic SR-generated Ca(2+) sparks. We tested the hypothesis that in a physiologic Ca(2+) milieu, and independent of increased Ca(2+) influx, an increase in basal phosphorylation of SR Ca(2+) cycling proteins will convert stochastic Ca(2+) sparks into periodic, high-power Ca(2+) signals of the type that drives SANC normal automaticity. We measured phosphorylation of SR-associated proteins, phospholamban (PLB) and ryanodine receptors (RyR), and spontaneous local Ca(2+) release characteristics (LCR) in permeabilized single, rabbit VM in physiologic [Ca(2+)], prior to and during inhibition of protein phosphatase (PP) and phosphodiesterase (PDE), or addition of exogenous cAMP, or in the presence of an antibody (2D12), that specifically inhibits binding of the PLB to SERCA-2. In the absence of the aforementioned perturbations, VM could only generate stochastic local Ca(2+) releases of low power and low amplitude, as assessed by confocal Ca(2+) imaging and spectral analysis. When the kinetics of Ca(2+) pumping into the SR were increased by an increase in PLB phosphorylation (via PDE and PP inhibition or addition of cAMP) or by 2D12, self-organized, "clock-like" local Ca(2+) releases, partially synchronized in space and time (Ca(2+) wavelets), emerged, and the ensemble of these rhythmic local Ca(2+) wavelets generated a periodic high-amplitude Ca(2+) signal. Thus, a Ca(2+) clock is not specific to pacemaker cells, but can also be unleashed in VM when SR Ca(2+) cycling increases and spontaneous local Ca(2+) release becomes partially synchronized. This unleashed Ca(2+) clock that emerges in a physiological Ca(2+) milieu in VM has two faces, however: it can provoke ventricular arrhythmias; or if harnessed, can be an important feature of novel bio-pacemaker designs.
窦房结细胞(SANC)的肌质网(SR)Ca2+蛋白基础磷酸化水平较高,可产生部分同步、自发性、节律性、舒张期局部 Ca2+释放(LCR),而心室肌细胞(VM)的基础磷酸化水平较低,仅偶尔出现舒张期、随机的 SR 产生的 Ca2+火花。我们假设在生理 Ca2+环境中,且不增加 Ca2+内流的情况下,SR Ca2+循环蛋白的基础磷酸化增加将使随机 Ca2+火花转变为驱动 SANC 正常自律性的周期性、高功率 Ca2+信号。我们在生理 [Ca2+]下,在兔 VM 的单个通透细胞中测量了 SR 相关蛋白(PLB 和 RyR)的磷酸化和自发局部 Ca2+释放特性(LCR),并在抑制蛋白磷酸酶(PP)和磷酸二酯酶(PDE)或添加外源性 cAMP 之前和期间,或在特异性抑制 PLB 与 SERCA-2 结合的抗体(2D12)存在的情况下。在没有上述干扰的情况下,VM 只能产生低功率和低幅度的随机局部 Ca2+释放,这可以通过共聚焦 Ca2+成像和光谱分析来评估。当通过增加 PLB 磷酸化(通过 PDE 和 PP 抑制或添加 cAMP)或 2D12 增加 Ca2+泵入 SR 的动力学时,自组织的“时钟样”局部 Ca2+释放,在空间和时间上部分同步(Ca2+小波)出现,这些节律性局部 Ca2+小波的总和产生周期性的高幅度 Ca2+信号。因此,钙钟不仅对起搏细胞具有特异性,而且当 SR Ca2+循环增加且自发局部 Ca2+释放变得部分同步时,也可以在 VM 中释放。然而,在 VM 中生理 Ca2+环境中出现的这种未被束缚的钙钟有两个方面:它可能引发室性心律失常;或者如果加以利用,它可能成为新型生物起搏器设计的重要特征。