We investigated the mechanisms of pyruvate transport in the erythrocytes of an ancient marine agnathan, the Pacific hagfish (Eptatretus stouti), and a sedentary euryhaline teleost, the starry flounder (Platichthys stellatus). Uptake of [C-14]pyruvate (50 muM, 10%C) by flounder erythrocytes was slow (t(1/2) (half-life) + 30 min), nonconcentrative, and mediated by the band 3 Cl-/HCO3- exchanger in combination with a process similar to the H+/monocarboxylate symporter present in freshwater teleosts and mammalian erythrocytes. In contrast, pyruvate uptake by hagfish erythrocytes (50 muM, 10%C) was rapid (t(1/2) + 1.5 min) and, in 10 min, reached an intracellular concentration more than 20-fold higher than that present in the extracellular medium. Pyruvate accounted for almost 90% of the accumulated intracellular radioactivity, the remaining label being incorporated into tricarboxylic acid cycle intermediates and glutamate. Influx of pyruvate was saturable (apparent K-m = 12 mM) and inhibited by p-chloromercuriphenylsulphonate (PCMBS) (K-i = 71 muM) and 4,4'-diisothiocyanatostilbene-2,2'-disulphonate (DIDS) (K-i = 0.49 mM). Transport was inhibited poorly by alpha -cyano-4-hydroxycinnamate (CIN) (K-i > 4 mM) and was not coupled to the movement of protons. Instead, the influx of pyruvate was Na+ dependent. A sigmoidal relationship between pyruvate transport and extracellular Na+ concentration was observed, suggesting a Na+:pyruvate coupling ratio greater than 1:1. In contrast with previously described Na+-dependent monocarboxylate transport activities in mammalian renal and intestinal epithelia, the hagfish erythrocyte system did not transport lactate.