Influence of endogenous vs exogenous α-ketoglutarate in the renal transport of organic anions

Epifluorescence microscopy and intact S2 segments of rabbit proximal tubules were used to study peritubular organic anion transport Preloading tubules with 100 μM α-ketoglutarate (αKG) in HEPES buffer increased initial rates of 1 μM fluorescein (FL) uptake 4.3-fold. Incubating tubules in HCO₃-buffered media, however, increased basal FL transport to the same level achieved in αKG-preloaded, HEPES-buffered tubules Preloading tubules with 100 μM αKG in HCO₃ buffer also stimulated FL uptake, but only by two-fold. Inhibition of transaminase activity by 1 mM aminooxyacetate stimulated FL uptake into tubules incubated in HEPES-buffer, suggesting that elevated cytosolic αKG concentrations were the basis for the higher rate of transport noted in HCO₃-buffered media. Mimicking the in situ condition, steady-state exposure of tubules to 10 μM αKG in a HCO₃-buffer increased FL transport by 40% Inhibiting peritubular Na-dicarboxylate cotransporter with 2 mM Li under these conditions immediately reduced transport by 20%. Inhibiting the Na-dicarboxylate cotransporter in the absence of αKG (but the presence of HCO₃) using either 2 mM Li or 1 mM methylsuccinate, also inhibited FL transport by 20% Thus, dicarboxylate transport increases organic anion transport by a direct coupling effect and by increasing cytosolic concentrations of αKG We conclude that 60% of the exchangeable dicarboxylate that drives organic anion uptake across the peritubular membrane of proximal tubules is supplied from endogenous sources. Activity of the Na-dicarboxylate cotransporter supplies approximately 50% to 60% of the exchangeable dicarboxylate by providing exogenous αKG and "recycling" endogenous dicarboxylate.