The cytotoxicity resulting from a 1-h exposure of Jensen tumor (JT) cells to 5-fluorouracil (FUra), quantified in terms of growth delay, was potentiated 4.1-fold if inosine was included in the culture medium for 30 min prior to and for 1 h concurrently with drug uptake. In contrast, as reported previously, protection from FUra toxicity is induced if the cells are exposed successively to inosine for 1.5 h and to FUra for 1 h. Anabolic conversion of radiolabeled FUra to free and polymeric nucleotides was stimulated by the cytotoxicity-enhancing mode of inosine exposure but inhibited by the inosine exposure that protects against FUra. Continuous exposure of JT cells to inosine after a 1-h uptake of FUra was reported earlier to potentiate drug efficacy 5-fold. Treatment of cultures by both modes of inosine exposure that enhance FUra cytotoxicity resulted in a 15.8-fold potentiation of the latter.