Thiamethoxam at environmentally relevant concentrations induces neurotoxicity in zebrafish larvae through binding with multiple receptors

Thiamethoxam (THM) is one of the most widely used insecticides globally, which was designed to selectively target nicotinic acetylcholine receptors (nAChRs) in the insect nervous system and is generally considered safe for non-targeted organisms. However, increasing evidence has demonstrated its neurotoxicity in aquatic organisms, though the underlying mechanisms, especially at environmentally relevant concentrations, remain largely unclear. In this study, the swimming distance of zebrafish was significantly shortened by 14.06%-21.64% after exposure to THM at 10-1000 ng/L. This behavioral impairment may result from the damage to nervous and visual systems, as confirmed by notable apoptosis, histological analysis of the eyes, and differential expression of numerous genes. Molecular docking and biomarkers assays found that THM can bind with nAChR and multiple hormone receptors, with binding energies varying from -3.75 to -6.74 kcal/mol. Consequently, the concentrations of a neurotransmitter (acetylcholine) and related hormones (cortisol, triiodothyronine, thyroxine, and thyroid-stimulating hormone) were significantly affected. Further investigations using a weighted gene correlation network and metabolomics suggest that THM may enter the cell via endocytosis and bind with multiple hormone receptors, potentially activating the MAPK signaling pathway. This activation may disrupt purine and pyrimidine metabolism in the cell nucleus, ultimately leading to cell apoptosis and neurotoxicity. This study reveals that THM, even at environmentally relevant concentrations, poses neurological risks to zebrafish and underscore the need for urgent attention to the ecological impacts of THM in aquatic environments.