Although reabsorption across the apical (AP) membrane of the renal proximal tubule cell plays a vital role in the conservation of plasma 5-methyltetrahydrofolate, basolateral (BL) membrane-directed secretory pathways may also be important in regulating the urinary excretion of folate. Folate transport proteins, folate receptor and the reduced folate carrier have been implicated in the renal conservation of folate across the AP membrane, but their role in BL membrane-directed folate transport has not been studied. 5-Methyltetrahydrofolate transport across the AP and BL membranes of human proximal tubule cells was studied in cells grown on membrane inserts to allow optimum differentiation of AP and BL domains. Colchicine, an inhibitor of vesicular-mediated endocytosis, inhibited AP binding and AP-directed transport without affecting BL transport. Probenecid, an inhibitor of anion exchange, did not affect binding, but inhibited both AP and BL-directed transport with a greater effect on BL transport. Folic acid abolished AP binding of 5-methyltetrahydrofolate, but diminished AP-mediated transport by only 50%. These data suggest that both the folate receptor and the reduced folate carrier participate in AP uptake of folates by human kidney cells, but that BL-mediated uptake occurs primarily by the reduced folate carrier. Folate transport from the secretory direction occurred as readily as that from the reabsorptive direction, indicating that altered secretion could mediate excess urinary folate excretion.

Deficiency of folate in heavy-drinking alcoholic populations can occur partly because of an increased urinary folate excretion. Ethanol directly reduces the reabsorption of folate in the renal proximal tubule (PT) by acting on either of 2 folate transport proteins, the reduced folate carrier (RFC) and the folate receptor (FR). This study was designed to determine the effects of ethanol on the transport of folate by PT cells and to examine the effects of ethanol on RFC and the FR protein expression. Normal human PT (HPT) cells were cultured on membrane inserts to study intracellular transport of 5-methyltetrahydrofolate from the apical or basolateral direction in the presence of ethanol [11-109 mmol/L (50-500 mg/dL)]. The long-term effect of ethanol on the renal folate transport protein content was determined by western blot in treated HPT cells and in vivo in rats pair-fed control diets or ethanol-containing liquid diets. A 1-h treatment of HPT cells with ethanol (> or = 65 mmol/L) reduced the apically directed transport of folate by 20-25% without affecting the basolateral transport. A 5-d exposure of HPT cells to ethanol dose-dependently increased the content of both the FR and RFC proteins, with a greater effect on the RFC. Similarly, a 14-d exposure of rats to ethanol increased the in vivo expression of both the RFC and FR. These studies demonstrate that ethanol decreases the reabsorptive transport of folate by renal PT cells, which would increase urinary folate excretion. In contrast, subchronic exposure of PT cells, both in vivo and in vitro, to folate-depleting concentrations of ethanol leads to an upregulation of the 2 folate transport proteins. The increase in folate transporters partly counteracts the inhibitory effects of ethanol on folate transport activity, which explains the lower magnitude of ethanol's effect on transport with subchronic exposure compared with that with acute exposure.

The reabsorption of 5-methyltetrahydrofolic acid (5-CH3-H4PteGlu) by the renal proximal tubule has an important role in the maintenance of plasma folate concentrations. However, the mechanism by which this vitamin traverses the renal epithelium remains to be determined. Studies in cultured cells have suggested that the folate receptor in association with a probenecid-sensitive anion carrier may be involved in the transmembrane transport of the vitamin. Because 5-CH3-H4PteGlu is reabsorbed and metabolized in the isolated perfused rat kidney (IPRK) in a smaller manner to in vivo models, the IPRK was used to evaluate pathways involved in folate reabsorption. Reabsorption of 5-CH3-H4PteGlu could not be saturated in the isolated perfused rat kidney, even at concentrations up to 2 mumol/L. Folic acid (PteGlu) was used as a competitive inhibitor of FR-dependent reabsorption of 5-CH3-H4PteGlu. When 5-CH3-H4PteGlu was maintained at 1 nmol/L (a concentration at which receptor-mediated transport should be maximal), PteGlu (up to 100 nmol/L) had no effect on reabsorption. The addition of probenecid (1 mmol/L) did not affect the reabsorption of 5-CH3-H4PteGlu but inhibited the fractional excretion of the anion para-aminohippurate. Probenecid also inhibited the urinary excretion of 5-CH3-H4PteGlu metabolites, indicating that reabsorbed 5-CH3-H4PteGlu was metabolized to products that were subsequently secreted into the urine by anion exchange pathways. The physiological importance of a folate receptor-mediated reabsorption of 5-CH3-H4PteGlu appears to be minor in the isolated perfused rat kidney, whereas nonspecific pathways appear to play a major role in the renal folate reabsorption.