Abstract: To characterize the mechanism of H+ release from rice roots under ammonium nutrition, rice seedlings (Oryza sativa L.) were incubated in a range of NH4+ concentration (0 1 mol/L) for 4 hours. H+ release was calculated by titration of the root exudation solution with 1mol/L NaOH. The plasma membrane vesicles of roots were isolated by PEG-DEXTRAN two-phase system. The plasma membrane H+-ATPase hydrolytic activity and pump activity was tested. In addition, plasma membrane potential was investigated by micro-electrode in situ. The correlations between the H+ release, membrane potential, plasma membrane H+ ATPase activity were analyzed. The results show that: (1) the H+ release is increased with the increase of NH4+concentration in root medium, (2) in vitro, both the plasma membrane hydrolytic and pump activities are increased, (3) in situ, membrane potential depolarization is also increased, and (4) there correlations between membrane potential, plasma membrane H+ ATPase, H+ pump activity and H+ release. These results indicate that the H+ release is closely related to the plasma membrane H+ pump activity, which is induced by membrane depolarization under NH4+-N.