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This study investigates the impact of the introduction of high-speed rail (HSR) on regional income and innovation, focusing on peripheral regions in Japan. Average income per capita (IN) and number of patent applications per employee (PA) are assumed to represent regional income and innovation, respectively. “Peripheries” are defined based on each municipality’s employment density and its accessibility, the latter is measured by effective density, an index representing interregional communication opportunities. Municipality-level panel data from Japan for a 40-year period are used. A difference-in-differences analysis of municipalities with variable treatment timings is employed to investigate the impact of HSR on the IN and PA of peripheries. The findings indicate that the introduction of HSR has been able to contribute to the mitigation of regional inequalities by positively affecting the IN and PA of peripheries. Furthermore, the results estimated with the accessibility-based peripheries show that the impacts of HSR on IN are smaller and on PA are larger in these peripheries than in core regions. These suggest the crucial role that accessibility may play in peripheries for catching up with the economic growth of core regions, particularly in a context of geographical concentration of economic activities, as in the case of Japan. It is also argued that the enhanced PA in peripheries with poor accessibility might not have been well exploited for economic gains because of their underdeveloped industrial structures and distance to large hub cities, leading to challenges in catching up.