Characteristics of folic acid transport in the L1210 leukemia cell

Uptake of folic acid was studied in L1210 leukemia cells. Folic acid was rapidly metabolized in this cell system and the metabolites appeared to be exchangeable across the cell membrane. Metabolism was partially reduced when cells were pretreated with methotrexate (4-amino-4-doxy-10-methylpteroylglutamic acid) to inhibit dihydrofolate reductase. The energy dependence of folic acid transport was investigated in cells pretreated with methotrexate to minimize metabolism. These studies suggested an energy-dependent efflux mechanisms for folic acid, since (1) the ratio of intracellular to extracellular folic acid at the steady state was far below that predicted for a passive equilibrating transport system and (2) inhibition of energy metabolism resulted in a decrease in the unidirectional efflux of folic acid and a marked rise in the net uptake velocity. Although interpretation of the data is complicated by residual metabolism of folic acid, many of the findings for folic acid are similar to those observed for methotrexate-a compound which shares, at least in part, the same transport mechanism as folic acid and is not metabolized under the conditions of these experiments. Compatible with an active efflux mechanism was the observation that net uptake of [³H]folic acid is a complex process in which back flux of label becomes significant within 30 sec after the addition of folic acid to the extracellular medium and at cell folate levels far below the steady state. Following this, the rate slows to an essentially constant velocity for the remainder of the period of observation and represents a very small difference between much larger unidirectional influx and efflux rates. Net uptake is highly temperature dependent (Q₁₀ of 5) and is markedly inhibited by methotrexate and folinic acid. Addition of these substances to the extracellular medium during uptake of folic acid results in net efflux of essentially all intracellular label.