P-Aminohippurate Uptake Across the Basolateral Membrane of Rat Proximal Tubule Cells: Specificity and Mode of Energetization

Abstract

Uptake of [3H]PAH (p-aminohippurate) across the basolateral membrane of rat proximal tubule cells was studied in intact kidneys and membrane vesicles to determine biologically and pharmacologically important substrates of the PAH transporter and its energization by ion gradients. Effective inhibition of in situ [3H]PAH uptake by trans-2-decanoate, α-ketoglutarate, prostaglandin E2, 11-dehydrocorticosterone, and dibutyryl cyclic GMP suggests an involvement of the PAH transporter in cellular entry and exit of metabolic products, selected hormones and second messengers. The high affinity for N-acetyl-, N-benzoyl-, N-acetylcysteine-, sulfate- and glucuronide-conjugates supports the role of the PAH transporter in renal excretion of metabolites. Among the drugs, apalcillin proved an inhibitor of high affinity (app. K i,PAH 0.02 mM) and selectivity for the PAH transporter. The nephrotoxic agents, ochratoxin A, orellanin, and β-amanitin, also inhibited in situ [3H]PAH uptake indicating that their cellular uptake proceeds via the PAH transporter. In vesicles, glutarate and an out > in Na+ gradient, but not a Na+ gradient alone, drove uphill [3H]PAH accumulation. Stimulation of [3H]PAH uptake was half-maximal at 10.8 µM glutarate and 2.3 mM Na+ with 3 Na+ ions being involved. These results reflect the kinetic properties of the (Na+)3-dicarboxylate cotransporter and prove its role in uphill PAH transport. Glutarate/PAH exchange is electroneutral, suggesting either exchange of one glutarate for two PAH molecules, or the involvement of hitherto undefined ions in the exchange process. In conclusion, PAH uptake across the basolateral membrane occurs through exchange with α-ketoglutarate, which is itself intracellularly accumulated by the Na+-coupled dicarboxylate transporter.