An assessment of the importance of exposure routes to the uptake and internal localisation of fluorescent nanoparticles in zebrafish (Danio rerio), using light sheet microscopy

A major challenge in nanoecotoxicology is finding suitable methods to determine the uptake and localisation of nanoparticles on a whole-organism level. Some uptake methods have been associated with artefacts induced by sample preparation, including staining for electron microscopy. This study used light sheet microscopy (LSM) to define the uptake and localisation of fluorescently labelled nanoparticles in living organisms with minimal sample preparation. Zebrafish (Danio rerio) were exposed to fluorescent gold nanoparticles (Au NPs) and fluorescent polystyrene NPs via aqueous or dietary exposure. The in vivo uptake and localisation of NPs were investigated using LSM at different time points (1, 3 and 7 days). A time-dependent increase in fluorescence was observed in the gut after dietary exposure to both Au NPs and polystyrene NPs. No fluorescence was observed within gut epithelia regardless of the NP exposure route indicating no or limited uptake via intestinal villi. Fish exposed to polystyrene NPs through the aqueous phase emitted fluorescence signals from the gills and intestine. Fluorescence was also detected in the head region of the fish after aqueous exposure to polystyrene NPs. This was not observed for Au NPs. Aqueous exposure to Au NPs resulted in increased relative swimming distance, while no effect was observed for other exposures. This study supports that the route of exposure is essential for the uptake and subsequent localisation of nanoparticles in zebrafish. Furthermore, it demonstrates that the localisation of NPs in whole living organisms can be visualised in real-time, using LSM.