The effect of calcium channel blockers and calcium on methotrexate accumulation in rat hepatocytes.

The effects of diltiazem (DIL), verapamil (VRP) and Ca2+ on the accumulation of methotrexate (MTX) were investigated in isolated rat hepatocytes. At the physiological 2 mM Ca2+, the calcium-channel blockers DIL (100 microM) and VRP (50 microM) significantly reduced the hepatocellular accumulation of MTX. By increasing the Ca2+ concentration to 7 mM control MTX levels (at 2 mM Ca2+) were restored with VRP, and resulted in MTX levels above the controls for DIL. Ca2+ at 7 mM significantly enhanced MTX accumulation in the hepatocyte suspensions after 60 min. The concentration time curves for MTX indicated that for the first 10 min influx was the dominating process. Dixon plot analysis of this uptake phase revealed Ki values of 140 microM for DIL and 75 microM for VRP. The data suggested that DIL was a non-competitive, and VRP a competitive inhibitor of MTX influx. Hence, the inhibitory effect on MTX accumulation mediated by DIL and VRP could be due to different mechanisms.