As social demands for biodiversity restoration and conservation rise, rapid declines in riverine biodiversity have been documented globally. Threats to biodiversity, particularly freshwater fish, have been documented in Japan, yet scientific evidence to support these claims is lacking. We collected 25 years of data from the National Census of the River Environment, which was continued for 109 class A rivers in Japan since 1990 and the survey methodology was established in 1996, to examine temporal changes in riverine fish assemblages. Fish surveys in each river were conducted once every five years, though the survey year varied by river. We used generalized additive mixed models to represent various trends in living planet index and fish populations and over a 25-year period. Rivers were treated as random effects. The living planet index fluctuated, but not in the monotonous decline that has been documented globally. Many fish species exhibited increasing trends, suggesting that conservation and restoration measures implemented in Japanese rivers have had some effect. However, bitterling and stickleback fishes have declined over the past 25 years, with some species approaching zero population density, and riverine fishes inhabiting floodplain ponds, agricultural channels, and brackish declined more than those utilizing running water as their primary habitat. Temporal change in the LPI, which comprehensively assessed population fluctuations of all species, may mask declines in rare species due to increases in populations of dominant species. Thus, stagnant and slows-flowing area such as floodplains and estuary and connectivity should be focused on to bend the curve of fish diversity loss for conservation and restoration in Japan.