INTRACELLULAR ALPHA-KETOGLUTARATE CONTROLS THE EFFICACY OF RENAL ORGANIC ANION TRANSPORT

Renal organic anion secretion is driven by indirect coupling to the Na+ gradient at the basolateral membrane through Na+-dicarboxylate cotransport and dicarboxylate-organic anion exchange. The impact of changing intracellular alpha-ketoglutarate (alpha KG) concentrations and gradient on p-aminohippurate (PAH) transport was assessed in rat renal cortical slices. Fluorimetric analysis of alpha KG indicated that freshly isolated slices averaged 137 +/- 4 nmol/g wet weight (similar to 265 mu M in cellular water). This value was sustained over several hours at 4 degrees C. On incubation at 22 degrees C, intracellular alpha KG concentrations rose steadily, reaching levels of 2 (air) to 4 (100% O-2) times that of fresh tissue. When internal alpha KG was increased by preincubation and PAH uptake was determined at a fixed gradient, PAH transport increased with increasing internal alpha KG. Conversely, at a fixed internal alpha KG concentration, PAH uptake was a linear function of the driving force provided by the alpha KG gradient. Thus, intracellular alpha KG is a major determinant of the efficacy of renal organic anion transport, and events that alter internal alpha KG concentration, gradient, or both are poised to exert significant control over organic anion secretion. Kinetic analysis of alpha KG-PAH exchange indicated that the K-m, for alpha KG was 151 mu M in basolateral membrane vesicles and 131 mu M in slices. Because PAH transport increased at intracellular alpha KG concentrations that should have been saturating, this finding indicates that cytoplasmic alpha KG levels must be substantially lower than total tissue concentration, i.e., that much intracellular alpha KG must be sequestered.