Quaternary climatic cycles have influenced marine organisms' spatial distribution and population dynamics. This study aimed to elucidate the evolutionary influences of contemporary and glacial physical barriers on the population structure, demography and colonization history of the mudskipper (Periophthalmus modestus) based on a mitochondrial gene segment (ND5) from 131 individual fish sampled in the northwestern Pacific Ocean. The current Kuroshio Current and the glacial exposure of the Taiwan Strait appeared to have restricted migration among the South China Sea, coastal East China and Japan. However, genetic homogeneity (Nm>1) also suggested contemporary larval transportation by sea circulation between the East China Sea and the South China Sea or historical dispersal along the glacial exposed shoreline among China, Japan and the Ryukyu Islands. Evolutionary signals of the strengthened East Asian Summer Monsoon in the mid-Pleistocene and regional difference in intertidal primary productions were indicated by a late-Pleistocene population expansion of P. modestus with a higher effective population size in the South China Sea than in the East China Sea. Furthermore, a potential colonization origin from the South China Sea was consistently inferred by different clues, including the populations' coalescence times, the ancestral haplotype distribution, the number of private haplotypes and species/genetic diversity.