Chronic exposure to environmental concentrations of 6PPDQ induces neurotoxicity via perturbing multiple neurotransmission pathways in zebrafish

Previous research has demonstrated acute toxicity of 6PPD-quinone (6PPDQ), the ozonation product of tire-manufacturing additive 6PPD; however, chronic effects on aquatic organisms under real-world conditions remain unclear. We exposed 2-month-old zebrafish to 6PPDQ at concentrations of 2, 20 and 200 µg/L for 30 days. Whole-body desorption electrospray ionization-mass spectrometry (DESI-MS) revealed accumulation mainly in the muscle, abdomen, and brain of zebrafish exposed to 200 µg/L 6PPDQ, which was verified by MS quantification of muscle and brain tissues. Based on velocity and distance moved, 2 µg/L 6PPDQ induced zebrafish hyperactivity, whereas 20 and 200 µg/L significantly inhibited activity. H&E staining revealed that chronic exposure to 6PPDQ caused brain damage in zebrafish, characterized by vacuolation and ventriculomegaly. Transcriptomic analyses indicated that 6PPDQ exposure disturbed neuron-related signaling pathways, especially glutamatergic/GABAergic, dopaminergic, cholinergic, and serotoninergic pathways. Downregulation of genes related to neurotransmission pathways was noted following exposure to 200 µg/L 6PPDQ. Reduced neurotransmitters, serotonin, 5-hydroxyindolacetic acid, dopamine, γ-aminobutyric acid, glutamate, acetylcholinesterase, 3,4-dihydroxyphenylacetic acid and homovanillic acid levels were also observed after 20 µg/L 6PPDQ exposure. Here, prolonged exposure to environmentally relevant concentrations of 6PPDQ induced neurotoxicity in zebrafish, and the neurotransmission system may be central to this process. These findings may help identify acceptable levels of aquatic 6PPDQ.