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Zebrafish embryos have been proposed as a cost-effective vertebrate model to study heart function. Many fluorescent genetically encoded Ca 2+ indicators (GECIs) have been developed, but those with ratiometric readout seem more appropriate to image a moving organ such as the heart. Four ratiometric GECIs based on troponin C, TN-XXL, Twitch-1, Twitch-2B, and Twitch-4 were expressed transiently in the heart of zebrafish embryos. Their emission ratio reported the Ca 2+ levels in both the atrium and the ventricle. We measured several kinetic parameters of the Ca 2+ transients: systolic and diastolic ratio, the amplitude of the systolic Ca 2+ rise, the heart rate, as well as the rise and decay times and slopes. The systolic ratio change decreased in cells expressing high biosensor concentration, possibly caused by Ca 2+ buffering. The GECIs were able to report the effect of nifedipine and propranolol on the heart, which resulted in changes in heart rate, diastolic and systolic Ca 2+ levels, and Ca 2+ kinetics. As a result, Twitch-1 and Twitch-4 ( K d 0.25 and 2.8 µM, respectively) seem the most promising GECIs for generating transgenic zebrafish lines, which could be used for modeling heart disorders, for drug screening, and for cardiotoxicity assessment during drug development.

Mapping Calcium Dynamics in the Heart of Zebrafish Embryos with Ratiometric Genetically Encoded Calcium Indicators