Insights into spirotetramat-induced thyroid disruption during zebrafish (Danio rerio) larval development: An integrated approach with in vivo, in vitro, and in silico analyses

Spirotetramat (SPT), a tetronic acid-derived insecticide, is implicated in reproductive and lipid metabolism disorders, as well as developmental toxicity in fish. While these effects are documented, the precise mechanisms underlying its developmental toxicity are not fully elucidated. In this study, zebrafish embryos (2 h post-fertilization, hpf) were exposed to four concentrations of SPT (0, 60, 120, and 240 μg/L) until 21 dpf (days post-fertilization). We delved into the mechanisms by examining its potential disruption of the thyroid endocrine system, employing in vivo, in vitro, and in silico assays. The findings showed notable developmental disturbances, including reduced hatching rates, shortened body lengths, and decelerated heart rates. Additionally, there was an increase in malformations and a decline in locomotor activity. Detailed analyses revealed that SPT exposure led to elevated thyroid hormone levels, perturbed the hypothalamic-pituitary-thyroid (HPT) axis transcript levels, amplified deiodinase type I (Dio1) and deiodinase type II (Dio2) activities, and both transcriptionally and proteomically upregulated thyroid receptor beta (TRβ) in larvae. Techniques like molecular docking and surface plasmon resonance (SPR) confirmed SPT's affinity for TRβ, consistent with in vitro findings suggesting its antagonistic effect on the T3-TR complex. These insights emphasize the need for caution in using tetronic acid-derived insecticides.