The portion of Na-K-ATPase activity on oxygen consumption was determined in suspensions of rat proximal tubules by monitoring oxygen consumption (Q<sub>O2</sub>) under different metabolic states: in the presence or absence of succinate or lactate (10 mmol/l) and increasing concentrations of ouabain (0.4; 0.8; 1.2 and 1.6 mmol/l). In the metabolic states tested, the ouabain induced decrease of Q<sub>O2</sub> was identical, which implies a fixed rate between Na-K-ATPase activity and Q<sub>O2</sub>. On the basis of these results, Ki and maximal inhibition rate were determined by Lineweaver-Burk and Eadie-Hofstee plots. K<sub>i</sub> was 0.67 mmol/l ouabain and the maximal inhibition of Q<sub>O2</sub> was 77%. This corresponds to an absolute decrease of Q<sub>O2</sub> of 1,630 µmol O<sub>2</sub> h<sup>-1</sup> g<sup>-1</sup> protein. Since this value represents the real portion of Na-K-ATPase on Q<sub>O2</sub>, the activity of Na-K-ATPase can be calculated, yielding an activity of 163 µmol ATP min<sup>-1</sup> g<sup>-1</sup> tubule protein.