We have examined the effects of verapamil and PSC 833 on cellular uptake and release of rhodamine 123 (R123) in two human cancer cell lines. Both verapamil and PSC 833 were able to increase R123 accumulation in the multidrug resistant (MDR) MV522/Q6 and KB-8-5 lines in the release study. However, the effects of these drugs on R123 accumulation during accumulation study were quite different. Incubation with PSC 833 increased R123 accumulation in both MDR lines. By contrast, incubation with verapamil only increased R123 accumulation in the KB-8-5 line. The failure of verapamil to increase R123 accumulation in the MV522/Q6 cells can be attributed to the presence of a carrier system in the parent MV522 cells that recognizes both R123 and verapamil, but not PSC 833, as substrates. These results imply that performing R123 accumulation study without first ascertaining possible role of a carrier system for cellular uptake of R123 and putative P-gp modulators might inadvertently lead one to draw improper conclusions on P-gp activity.,