Wüst Rob C I, Helmes Michiel, Martin Jody L, van der Wardt Thomas J T, Musters René J P, van der Velden Jolanda, Stienen Ger J M
Department of Physiology, Institute for Cardiovascular Research, VU University Medical Centre, Amsterdam, the Netherlands.
IonOptix LLC, Milton, MA, USA.
J Physiol. 2017 Mar 15;595(6):2001-2019. doi: 10.1113/JP273589. Epub 2017 Feb 22.
Calcium ions regulate mitochondrial ATP production and contractile activity and thus play a pivotal role in matching energy supply and demand in cardiac muscle. The magnitude and kinetics of the changes in free mitochondrial calcium concentration in cardiac myocytes are largely unknown. Rapid stimulation frequency-dependent increases but relatively slow decreases in free mitochondrial calcium concentration were observed in rat cardiac myocytes. This asymmetry caused a rise in the mitochondrial calcium concentration with stimulation frequency. These results provide insight into the mechanisms of mitochondrial calcium uptake and release that are important in healthy and diseased myocardium.
Calcium ions regulate mitochondrial ATP production and contractile activity and thus play a pivotal role in matching energy supply and demand in cardiac muscle. Little is known about the magnitude and kinetics of the changes in free mitochondrial calcium concentration in cardiomyocytes. Using adenoviral infection, a ratiometric mitochondrially targeted Förster resonance energy transfer (FRET)-based calcium indicator (4mtD3cpv, MitoCam) was expressed in cultured adult rat cardiomyocytes and the free mitochondrial calcium concentration ([Ca ] ) was measured at different stimulation frequencies (0.1-4 Hz) and external calcium concentrations (1.8-3.6 mm) at 37°C. Cytosolic calcium concentrations were assessed under the same experimental conditions in separate experiments using Fura-4AM. The increases in [Ca ] during electrical stimulation at 0.1 Hz were rapid (rise time = 49 ± 2 ms), while the decreases in [Ca ] occurred more slowly (decay half time = 1.17 ± 0.07 s). Model calculations confirmed that this asymmetry caused the rise in [Ca ] during diastole observed at elevated stimulation frequencies. Inhibition of the mitochondrial sodium-calcium exchanger (mNCE) resulted in a rise in [Ca ] at baseline and, paradoxically, in an acceleration of Ca release.
rapid increases in [Ca ] allow for fast adjustment of mitochondrial ATP production to increases in myocardial demand on a beat-to-beat basis and mitochondrial calcium release depends on mNCE activity and mitochondrial calcium buffering.
钙离子调节线粒体ATP生成和收缩活动，因此在心肌能量供需匹配中起关键作用。心肌细胞中游离线粒体钙浓度变化的幅度和动力学很大程度上尚不清楚。在大鼠心肌细胞中观察到游离线粒体钙浓度随刺激频率快速增加但下降相对缓慢。这种不对称导致线粒体钙浓度随刺激频率升高。这些结果为线粒体钙摄取和释放机制提供了见解，这在健康和患病心肌中都很重要。
钙离子调节线粒体ATP生成和收缩活动，因此在心肌能量供需匹配中起关键作用。关于心肌细胞中游离线粒体钙浓度变化的幅度和动力学知之甚少。利用腺病毒感染，在培养的成年大鼠心肌细胞中表达了基于比率性线粒体靶向福斯特共振能量转移（FRET）的钙指示剂（4mtD3cpv，线粒体钙成像探针），并在37℃下不同刺激频率（0.1 - 4Hz）和细胞外钙浓度（1.8 - 3.6mM）下测量游离线粒体钙浓度（[Ca]）。在单独实验中使用Fura - 4AM在相同实验条件下评估胞质钙浓度。0.1Hz电刺激期间[Ca]的增加迅速（上升时间 = 49 ± 2毫秒），而[Ca]的下降则较慢（衰减半衰期 = 1.17 ± 0.07秒）。模型计算证实这种不对称导致在较高刺激频率下舒张期[Ca]升高。线粒体钠钙交换体（mNCE）的抑制导致基线时[Ca]升高，并且反常地加速了钙释放。
[Ca]的快速增加允许线粒体ATP生成在逐搏基础上快速调整以满足心肌需求增加，并且线粒体钙释放取决于mNCE活性和线粒体钙缓冲。
