NA+-DEPENDENT AND NA+-INDEPENDENT URIDINE UPTAKE IN AN ESTABLISHED RENAL EPITHELIAL-CELL LINE, OK, FROM THE OPOSSUM KIDNEY

The characteristics of Na+-dependent and Na+-independent uridine uptake at 22-degrees-C were determined for monolayers of OK renal epithelial cells. The majority of uridine influx in subconfluent to early confluent (day 1 postconfluency) OK monolayers was mediated via a facilitated-diffusion pathway (apparent K(m) 160 +/- 41 muM, V(max) 610 +/- 100 pmol/mg protein per min). This system was inhibited with high affinity by nitrobenzylthioinosine (NBMPR) (IC50 value 1.5 nM) and by purine and pyrimidine nucleosides. Specific [H-3]NBMPR binding sites were detected in OK monolayers (apparent K(d) 0.67 +/- 0.25 nM, B(max) 90 +/- 19 fmol/mg protein) yielding a turnover number for the carrier of 112 uridine molecules/site per s at 22-degrees-C. Na+-dependent uridine uptake was minor in subconfluent OK monolayers, but increased 8-fold with time after confluency reaching a stable plateau at 8 days postconfluency. Inhibition of Na+-dependent 1 muM uridine uptake by inosine, guanosine, adenosine and uridine was biphasic with approx. 40% of the total uptake inhibited with high affinity (IC50 value 2 to 14 muM). Concentrations of thymidine and cytidine up to 1 mM had no effect on Na+-dependent uridine uptake and no Na+-dependent thymidine influx by confluent OK monolayers was detected. Using cell monolayers grown on a permeable filter support, Na+-dependent uridine uptake occurred preferentially from the apical. surface. This high affinity component of Na+-dependent uridine uptake is suggested to represent the Na+-dependent purine preferring Nl nucleoside transporter. The Na+/uridine stoichiometry for this system was consistent with 1:1. The remaining component of Na+-dependent uridine uptake was inhibited by some nucleosides, such as guanosine and inosine, with low affinity (IC50 values of 0.6 to 5 mM). Other nucleosides showed little specific inhibition. We propose that this component of uridine uptake represents a mutated carrier that binds nucleosides but is defective in the translocation of permeant.