Evaluating Cardiomyocyte Contractility and Calcium Dynamics

Cell culture media was replaced with 37°C Tyrode’s salt solution (supplemented with 0.1% DMSO with or without 50 μg/mL DEHP or 100 μM Wy-14,643), and spontaneous beating rate recordings were collected using a Zeiss LSM 510 confocal imaging system (488 nm excitation/505-550 nm emission filters). Cells were then equilibrated at room temperature for 20 min [19 (link)], and pace-induced calcium transient recordings were measured. In the latter, the cell network was paced using a stimulation electrode (Harvard Apparatus, Holliston MA) to which monophasic 5 msec pacing pulses were applied (4V minimum threshold, Grass Stimulator). In a second set of studies, cardiomyocytes were exposed to 20 mM caffeine in the presence of 20 mM KCl to monitor total sarcoplasmic reticulum (SR) load. Confocal imaging was accomplished at a spatial/temporal resolution of ~ 650 μm /36 fps; xt line scan resolution was ~1300 μm/650 fps.