Lactate uptake by skeletal muscle occurs under diverse conditions, including hypoxia and electrical stimulation. A possible metabolic fate of lactate in resting muscle is its conversion to pyruvate followed by carboxylation to malate in the cytosolic malic reaction. To test this hypothesis, we measured hindlimb lactate uptake in hypoxic mechanically ventilated rabbits. Rabbits were given intravenous infusions of hydroxymalonate, an inhibitor of the malic reaction (200 mM; n = 7), or normal saline (n = 7) at 1.1 ml/min. Hindlimb lactate uptake/release was calculated as femoral blood flow times the arteriovenous lactate difference. Saline or hydroxymalonate was infused continuously during sequential 30-min periods of normoxia (arterial PO2 approximately 150 Torr), hypoxemia (arterial PO2 approximately 30 Torr), and reoxygenation (arterial PO2 approximately 150 Torr). Hindlimb O2 transport decreased with hypoxemia, but O2 consumption remained unchanged in both groups. During hypoxemia there was net uptake of lactate by the hindlimb of the group given normal saline [4.5 +/- 0.9 (SE) mumol/min]. The hindlimb of the hydroxymalonate group continued to release lactate (-0.5 +/- 1.0 mumol/min). The inhibition of lactate uptake by hydroxymalonate supports the hypothesis that the malic reaction plays a major role in the metabolism of lactate by resting rabbit skeletal muscle.