High Doses of Norfloxacin Nicotinate Induce Apoptosis, Developmental Neurotoxicity, and Aberrant DNA Methylation in Zebrafish (Danio rerio) Larvae

This study aimed to evaluate the response mechanisms of zebrafish larvae to Norfloxacin nicotinate (NOR-N) exposure. Embryos were exposed to NOR-N from 4 h post-fertilization (hpf) until 96 hpf. The exposure concentrations included 0.002, 0.2, 1, and 5 mg/L (simulating both normal and exceptionally high environmentally relevant levels of NOR), as well as a high dose of 25 mg/L. Subsequent analyses focused on apoptosis, neurodevelopment, and DNA methylation in the resulting zebrafish larvae. The results showed that high-dose NOR-N (≥5 mg/L) induced obvious apoptotic cell death in zebrafish larvae, accompanied by increased activities of Cas3 and Cas9, up-regulated <i>P53</i>, <i>Bax</i>, <i>Puma</i>, <i>Apaf1</i>, <i>Cas3</i> and <i>Cas9</i> genes expression, and reduced <i>Mdm2</i> levels and <i>Bcl2/Bax</i> ratio. Moreover, exposure to 5 and/or 25 mg/L NOR-N resulted in a significant up-regulation of neurodevelopment-related genes (<i>Sox2</i>, <i>Sox3</i> and <i>Sox19a</i>), concomitantly with a marked decline in the transcription of DNA methylation genes, including <i>Dnmt1</i>, <i>Dnmt3a1</i>, <i>Dnmt3b1</i>, <i>Dnmt3b2</i> and <i>Dnmt3b4</i>. Overall, our findings demonstrated that NOR-N exposure could induce apoptosis, developmental neurotoxicity and aberrant DNA methylation in zebrafish larvae. These findings provide insights to guide the safe application of NOR-N in aquaculture and support the assessment of its potential ecological risks to aquatic ecosystems.