AbstractThe main goal of this study was to clarify whether the spectral properties of retinal photoreceptors reflect the features of behaviour of closely related fish species cohabiting shallow marine and fresh waters. The spectral sensitivity of photoreceptors was compared between two smelt species, Hypomesus japonicus and Japanese smelt Hypomesus nipponensis. The spectral absorption of the visual pigments was measured using microspectrophotometry. In H. japonicus, a mostly marine species, all photoreceptors contained visual pigments based on retinal and were distributed differently in specific retinal areas. The absorbance maxima (λmax) of rods and long‐wave‐sensitive members of double cones throughout the retina amounted to 507 and 573 nm, respectively, but the λmax value of the short‐wave‐sensitive members of double cones and single cones in the temporal hemiretina showed a significant blue shift compared to the nasal hemiretina: 485 vs. 516 nm and 375 vs. 412 nm, respectively, thus enhancing the short‐wave sensitivity of the temporal hemiretina. In H. nipponensis, an euryhaline species, the estimated λmax value of both rods and cones significantly varied between the groups caught in different localities (sea, river or estuary) because of the presence of rhodopsin/porphyropsin mixtures. The long‐wavelength shift in rod and cone photoreceptors was observed because of changes in the chromophore complement in closely related but ecologically different species dwelling in freshened bodies of water. Considering the data available in the literature, several putative common opsin genes have been suggested for species under study.