Tetrachlorantraniliprole induces neurodevelopmental toxicity through oxidative stress-mediated apoptosis and dysregulation of Wnt signaling pathway

Tetrachlorantraniliprole (TCTP) is a novel bisamide insecticide and widely used to protect against lepidopteran insect species. However, the application of TCTP in rice fields often leads to water pollution, posing threats to aquatic organisms and potentially to human health. Few studies have assessed the toxic effects of TCTP on aquatic animals. In this study, we used zebrafish as a model organism to evaluate the toxicity of TCTP. Our findings indicated that TCTP adversely affected the development of zebrafish larvae, impacting parameters such as heart rate, body length, and pericardial edema. Exposure to TCTP resulted in the increased embryo mortality along with higher concentrations of the compound. The expression of neurodevelopment-related genes was inhibited in embryos exposed to TCTP. Hematoxylin and eosin (HE) staining revealed that TCTP caused damage to the brain cells of the embryos. Behavioral analysis showed a reduction in activity of the larvaes, which aligned with a decrease in acetylcholinesterase (AChE) activity. Additionally, RNA sequencing (RNA-seq) was employed to elucidate the mechanisms of toxicity. GO and KEGG analysis identified that the pathways were related to oxidative stress, apoptosis and Wnt signaling. We observed an increase of reactive oxygen species (ROS) and Ca²/Mg²-ATPase activity, while antioxidant enzyme activities (SOD, MDA, CAT, Na/K-ATPase and T-ATPase) were significantly decreased in TCTP-exposed groups. Furthermore, TCTP induced brain cells apoptosis, as evidenced by the upregulation of pro-apoptotic genes (bax, p53, TNFα, caspase3 and caspase9) and the downregulation of anti-apoptotic gene (bcl2). Moreover, TCTP increased the expression of genes involved in Wnt signaling pathway. Notably, oxidative stress and neuronal damage induced by TCTP could be mitigated by astaxanthin, an antioxidant. Additionally, IWR-1, an inhibitor of Wnt signaling pathway, effectively alleviated the upregulation of genes associated with TCTP treatment and inhibited oxidative stress-induced apoptosis. In conclusion, this study demonstrated TCTP-induced defects of neurodevelopment and the brain cells in zebrafish larvae which were primarily driven by oxidative stress-induced apoptosis and dysregulation of Wnt signaling pathway. Importantly, these toxic phenotypes can be rescued by treatment with astaxanthin or IWR-1.