Assessment of Sarcoplasmic Reticulum Calcium Reserve and Intracellular Diastolic Calcium Removal in Isolated Ventricular Cardiomyocytes

作者：Gao, Jing; Shi, Xiaolu; He, Hong; Zhang, Juhong; Lin, Ding; Fu, Guosheng; Lai, Dongwu^*

来源：Jove-Journal of Visualized Experiments, 2017, (127): e55797.

DOI：10.3791/55797

• Cellular Biology
• Issue 127
• Sarcoplasmic reticulum
• calcium homeostasis
• calcium reserve
• intracellular calcium removal
• SERCA
• cardiomyocytes

Intracellular calcium recycling plays a critical role in regulation of systolic and diastolic function in cardiomyocytes. Cardiac sarcoplasmic reticulum (SR) serves as a Ca2+ reservoir for contraction, which reuptakes intracellular Ca2+ during relaxation. The SR Ca2+ reserve available for beats is determinate for cardiac contractibility, and the removal of intracellular Ca2+ is critical for cardiac diastolic function. Under some pathophysiological conditions, such as diabetes and heart failure, impaired calcium clearance and SR Ca2+ store in cardiomyocytes may be involved in the progress of cardiac dysfunction. Here, we describe a protocol to evaluate SRCa2+ reserve and diastolic Ca2+ removal. Briefly, a single cardiomyocyte was enzymatically isolated, and the intracellular Ca2+ fluorescence indicated by Fura-2 was recorded by a calcium imaging system. To employ caffeine for inducing total SR Ca2+ release, we preset an automatic perfusion switch program by interlinking the stimulation system and the perfusion system. Then, the mono-exponential curve fitting was used for analyzing decay time constants of calcium transients and caffeine-induced calcium pulses. Accordingly, the contribution of the SR Ca2+-ATPase (SERCA) and Na+-Ca2+ exchanger (NCX) to diastolic calcium removal was evaluated.

• 出版日期2017-9
• 单位浙江大学; 中国中医科学院