Acrolein is a widely distributed pollutant produced from various sources such as industrial waste, organic combustion, and power plant emissions. It is also intentionally released into irrigation canals to control invasive aquatic plants. Zebrafish (Danio rerio) has a good reputation for being an attractive model organism for developmental and toxicological research. In this study, zebrafish embryos were exposed to acrolein to investigate the cardiotoxic effects. The 96-h LC50 (median lethal concentration) value of acrolein was determined as 654.385 μg/L. Then, the embryos were treated with the sublethal experimental concentrations of acrolein (1, 4, 16, 64, and 256 μg/L) for 96 h. Embryos were examined at 48, 72, and 96 h post-fertilization (hpf). Acrolein affected the cardiac morphology and function of the embryos. Sinus venosus-bulbus arteriosus (SV-BA) distance of 64 μg/L and 256 μg/L acrolein groups was elongated compared with the control samples. Immunostaining with MF20 antibody clearly exhibited that the atrium positioned posterior to the ventricle which indicated cardiac looping inhibition. Histological preparations also showed the mispositioning and the lumens of the chambers narrowed. Acrolein-induced increased heart rate was noted in the 4, 16, 64, and 256 μg/L treatment groups. Taken together, these results indicated that acrolein disrupted the heart development and cardiac function in zebrafish, suggesting that its water-borne risks should be considered seriously.