The effect of age in culture on the uptake and metabolism of methotrexate by primary cultures of hepatocytes.

Primary monolayer cultures of rat hepatocytes show an initial transient azide-sensitive capacity to concentrate methotrexate several-fold from the medium. However, after 48 h in culture they lose the azide-sensitive ability to accumulate methotrexate. The uptake of (+) 5-methyltetrahydrofolate and folate is not altered greatly with time in culture and is unaffected by azide. Once inside the cell, methotrexate is converted predominantly to di- and triglutamate derivatives. After a short exposure to methotrexate (up to 4 h) most of the intracellular methotrexate is unaltered, but by 24 h nearly 80% is present as polyglutamates. Unlike the capacity of the cells to accumulate methotrexate, the ability of the cells to form polyglutamates does not appear to diminish with the age of the culture.