Metals are increasingly polluting aquatic ecosystems, accumulating in biota and causing adverse effects. Previous studies that have explored the accumulation of metals in fish tissues and examined potential chemical changes are limited to adults. Therefore, this study uses zebrafish (Danio rerio) embryos and larvae to investigate regions of copper (Cu) accumulation in larvae exposed to environmentally relevant concentrations of Cu and determine whether there are any adverse effects to the regions of Cu accumulation. Embryos at 24 h post fertilisation were exposed to ≤11 µg L-1 Cu for 48 and 96 h. Synchrotron X-ray fluorescence microscopy (XFM) revealed increases in Cu and zinc (Zn) in the ocular tissues in fish exposed to just Cu, and further analysis of this tissue, using X-ray absorption near edge structure (XANES) spectroscopy, showed a change to the local atomic environment of the accumulated Zn after 96 h of Cu exposure, relative to the controls. Moreover, synchrotron Fourier-transform infrared microscopy (FTIR) revealed the larvae exposed to Cu for 96 h exhibited chemical changes in the retinal pigment epithelium, largely to aldehyde and Schiff-base, suggesting a decrease in the abundance of rhodopsin. In contrast, it was only at the earlier time point of 48 h that exposure to ≤11 µg L-1 of Cu caused the differential expression of proteins involved in retinol biosynthesis, the visual cycle and phototransduction, as shown using proteomics. This study employed multiple techniques to explore the fate of Cu in larval zebrafish and observed the accumulation and disruption to ocular tissues, which ultimately may disrupt visual processing and key visual behaviours necessary for survival in the wild.