Comprehensive Evaluation of the Embryotoxicity and Ototoxicity of a Car Diffuser on Zebrafish Larvae: Impacts on Morphology and Hair Cells

Diffusers release various chemical compounds into the environment, some of which may have ototoxic effects. Despite their widespread use, the safety profiles of these compounds remain largely unexplored. This study evaluated the potential toxicity of a car diffuser extract, Diff3, on zebrafish larvae by assessing morphology, survival rate, hair cell integrity, and rheotactic behavior. Forty wild-type zebrafish larvae at 4 h post-fertilization (hpf) were exposed to Diff3 and observed at 3 and 5 days post-fertilization (dpf) using a stereomicroscope. Hair cell damage was assessed using YO-PRO-1 iodide staining in four neuromasts and oxidative stress was measured in transgenic (Brn3C:EGFP) larvae using CellROX Deep Red Reagent. Rheotactic behavior was analyzed using a specialized behavioral testing system. No significant morphological changes or survival differences were observed between groups. However, hair cell counts were significantly reduced in Diff3-treated larvae, and ROS levels were notably higher in the treatment group. However, in additional experiments, no significant changes in rheotactic behavior were observed. These findings suggest that Diff3 induces hair cell loss through oxidative stress, which may affect sensory function. This study highlights the need to evaluate the ototoxic potential of household chemical products and emphasizes the importance of regulatory measures to ensure consumer safety. Further research is required to determine long-term effects and potential recovery mechanisms following exposure to such compounds.