ABSTRACT In this study, we explored the gill mRNA expression ofNa+/K+-ATPase α1 (NKAα1) and Na+/K+/2Cl− cotransporter 1a(NKCC1a) in an amphibious mudskipper species, Periophthalmus waltoniat high salinities of mangrove tidal zones. Fish were exposed to four salinities including 35 (control), 45, 55, and 70 g kg−1. The branchial transcripts of NKAα1 were elevated in fish kept at 55 and 70 g kg−1 salinities with a strong elevation after medium-term (11 days) of exposure. No obvious changes were observed in NKCC1a expression among different salinity groups after short-term (3 days) and long-term (21 days) of exposure, except for a medium-term strong upregulation of NKCC1agene at 55 g kg−1. The NKAα1 and NKCC1a expressions returned to baseline levels after the long term exposure salinity period, suggesting complete molecular and physiological acclimation.These results demonstrate the osmoregulatory role of ion transporter enzymes in acclimation of amphibious fish to high salinities.