Estimating risk of cardiovascular pharmaceuticals in freshwaters using zebrafish embryotoxicity test - statins threat...

Cardiovascular pharmaceuticals (CVPs) are globally present in inland waters and have also been found in the sediment and plasma of fish from the Sava River, Croatia. Based on the previous research, CVPs amiodarone (AMI), ramipril (RAM), simvastatin (SIM), and verapamil (VER) have been selected for this study. Their effect has been investigated, individually and in a mixture, on the development of the zebrafish embryo Danio rerio (Hamilton, 1822) within the first 96 h of development. Upon exposure to environmentally relevant concentrations of tested CVPs (0.1, 1, and 10 μg/L) zebrafish survival and development as apparent from observed morphological abnormalities, heartbeat rates and changes in behavior, hatching success, larval length and oxidative stress level were monitored. The CVP causing the highest mortality and pathological changes was SIM (1 and 10 μg/L), which corresponds well with the observed effects during zebrafish exposure to CVPs' mixtures (4 and 40 μg/L). All pharmaceuticals affected cardiac function and decreased heart rate. SIM (1 μg/L), VER and RAM (10 μg/L) decreased larval length, while induced oxidative stress was recorded in the SIM- and VER-exposed specimens. Behavioral alterations of zebrafish were observed only in AMI-treated group (10 μg/L). Our amino acid sequence comparison and structural and docking analysis showed a highly conserved binding site between human and zebrafish HMG-CoA reductase for SIM and its main metabolite simvastatin acid. Using these ecotoxicological bioassays on a zebrafish model with particular emphasis on sublethal endpoints, the risk of CVPs, especially statins, for fish in inland waters has been identified.