The links between the mechanisms of Na+, K+ and H+ movements in M. mycoides var. Capri cells undergoing glycolysis were studied. Certain properties of the membrane-bound ATPase of Mycoplasma cells were investigated with special reference to ionic requirements and sensitivity to specific inhibitors. Although Na+ stimulated ATPase activity, K+ did not affect it, N,N''-dicyclocarboidimide and 7-chloro-4-nitrobenzo-2-oxa-1,3-diazole (NBD) were potent inhibitors of the basal ATPase activity, which was unaffected by vanadate and ouabain. The movements of Na+ and H+ were also studied. When Na+-loaded cells previously equilibrated with 22Na+ were diluted in a sodium-free medium, addition of glucose induced a rapid efflux of 22Na+. This energy-dependent efflux was independent of the presence of KCl in the medium. Studies of the changes in internal pH by 9-aminoacridine fluorescence or [14C]methylamine distribution indicated that the movement of Na+ was coupled to that of protons moving in the opposite direction, a finding that supports the presence of an Na+/H+ antiport. When Na+-loaded cells are diluted in an Na+-rich medium the Na+/H+ antiport is still active, but cannot decrease the intracellular Na+ concentration. Under such conditions, net 22Na+ extrusion is specifically dependent on the presence of K+ in the medium. The present results and those derived from the study of K+ accumulation can be rationalized by assuming that Mycoplasma mycoides var. Capri cells contain 2 transport systems for Na+ extrusion: an Na+/H+ antiport and an ATP-consuming Na+/K+-exchange system.