Developmental co-exposure of TBBPA and titanium dioxide nanoparticle induced behavioral deficits in larval zebrafish

Both tetrabromobisphenol A (TBBPA) and titanium dioxide nanoparticle (TiO₂ NP) have widespread commercial applications, resulting in their ubiquitous co-presence in the environment and biota. Although environmental chemicals exist as mixtures, toxicity studies are nearly always conducted with single chemicals. Few studies explore potential interactions of different chemical mixtures. In this study, we employ the sensitive developing nerve system in zebrafish to assess the neurotoxicity of TBBPA/TiO₂ NP mixtures. Specifically, zebrafish embryos were exposed to solvent control (0.1% DMSO), 2 μM TBBPA, 0.1 mg/L TiO₂ NP, and their mixture from 8 to 120 h post fertilization (hpf), and motor/social behavioral assessments were conducted on embryos/larvae at different developmental stages. Our results showed that TBBPA/TiO₂ NP single or co-exposures increased spontaneous movement, decreased touch response and swim speed, and affected social behaviors of light/dark preference, shoaling, mirror attack and social contact. In particular, many of these phenotypes were manifested with higher magnitude of changes from the mixture exposure. These behavioral deficits were also accompanied with increased cell death in olfactory region and neuromasts in the lateral line system, increased ROS in gallbladder, pancreas, liver, and intestine, as well as increased lipid peroxidation and decreased ATP levels in whole larval tissue homogenates. Further, genes coding for key cell apoptosis marker and antioxidant enzyme were significantly upregulated by these two chemicals, in particular to their mixture. Interestingly, the co-presence of TBBPA also increased the mean particle size of TiO₂ NP in the exposure solutions and the TiO₂ NP content in larval tissue. Together, our analysis suggests that TBBPA/TiO₂ NP induced behavioral changes may be due to physical accumulation of these two chemicals in the target organs, and TiO₂ NP may serve as carriers for increased accumulation of TBBPA. To conclude, we demonstrated that TBBPA/TiO₂ NP together cause increased bioaccumulation of TiO₂, and heightened responses in behavior, cell apoptosis and oxidative stress. Our findings also highlight the importance of toxicity assessment using chemical mixtures.