Aquaporins (AQPs) facilitate transmembrane water and solute transport， and in addition to contributing to transepithelial water transport， they safeguard cell volume homeostasis. This study examined the expression and localization of AQP1 and AQP3 in the gills of Japanese medaka ( in response to osmotic challenges and osmoregulatory hormones， cortisol， and prolactin (PRL). mRNA was inversely regulated in response to salinity with high levels in ion-poor water (IPW)， intermediate levels in freshwater (FW)， and low levels in seawater (SW). AQP3 protein levels decreased upon SW acclimation. By comparison， AQP1 expression was unaffected by salinity. In ex vivo gill incubation experiments， mRNA was stimulated by PRL in a time- and dose-dependent manner but was unaffected by cortisol. In contrast， was unaffected by both PRL and cortisol. Confocal microscopy revealed that AQP3 was abundant in the periphery of gill filament epithelial cells and co-localized at low intensity with Na，K-ATPase in ionocytes. AQP1 was present at a very low intensity in most filament epithelial cells and red blood cells. No epithelial cells in the gill lamellae showed immunoreactivity to AQP3 or AQP1. We suggest that both AQPs contribute to cellular volume regulation in the gill epithelium and that AQP3 is particularly important under hypo-osmotic conditions， while expression of AQP1 is constitutive.