Brush border membrane vesicles were purified from rabbit renal cortex using a calcium-precipitation procedure, and the uptake of carboxylic acids was determined by a rapid-filtration method. L-Lactate, pyruvate (monocarboxylic acids), and succinate (dicarboxylic acid) demonstrated features of Na+ cotransport: enhanced initial rate (1 s) of uptake with an inward Na+ gradient compared with the Na+ -free control condition and transient accumulation of substrate within the vesicles. Kinetic parameters derived for L-lactate and succinate show that each substrate is transported via single pathway and that the two substrates exhibit marginal cross-inhibition. A range of monocarboxylic acids including pyruvate and ketone bodies appear to interact with the monocarboxylic acid carrier. The kinetics of Nat-dependent pyruvate uptake suggest at least two transport pathways-namely, that this monocarboxylate shares both the mono- and dicarboxylic acid carriers. We conclude that isolated rabbit renal microvillus membranes possess independent transport systems for mono- and polycarboxylic acids.