An electroneutral Na+/H+ exchange mechanism (dimethylamiloride inhibitable, Li+ sensitive, and Ca2+ insensitive) was identified in brush-border membrane vesicles (BBMV) from Kuruma prawn hepatopancreas by monitoring Na(+)-dependent H+ fluxes with the pH-sensitive dye acridine orange and measuring 22Na+ uptake. Kinetic parameters measured under short-circuited conditions were the Na+ concentration that yielded one-half of the maximal dissipation rate (Fmax) of the preset transmembrane delta pH (KNa) = 15 +/- 2 mM and Fmax = 3,626 +/- 197 delta F.min-1.mg protein-1, with a Hill coefficient for Na+ of approximately 1. In addition, the inhibitory constant for dimethylamiloride was found to be approximately 1 microM. The electroneutral nature of the antiporter was assessed in that an inside-negative transmembrane electrical potential neither affected kinetic parameters nor stimulated pH-dependent (intracellular pH > extracellular pH) 22Na+ uptake. In contrast, electrogenic pH-dependent 22Na+ uptake was observed in lobster hepatopancreatic BBMV. Substitution of chloride with gluconate resulted in increasing KNa and decreasing delta Fmax, which suggests a possible role of chloride in the operational mechanism of the antiporter. These results indicate that a Na+/H+ exchanger, resembling the electroneutral Na+/H+ antiporter model, is present in hepatopancreatic BBMV from the Kuruma prawn Penaeus japonicus.