Active efflux transporters in the blood-brain barrier lower the brain concentrations of many drug molecules and endogenous substances and thus affect their central action. The objective of this investigation was to study the dynamics of the entire inhibition process of the efflux transporter P-glycoprotein (P-gp), using positron emission tomography (PET). The P-gp marker [(11)C]verapamil was administered to anesthetized rats as an i.v. bolus dose followed by graded infusions via a computerized pump system to obtain a steady-state concentration of [(11)C]verapamil in brain. The P-gp modulator cyclosporin A (CsA) (3, 10 and 25 mg/kg) was administered as a short bolus injection 30 min after the start of the [(11)C]verapamil infusion. The CsA pharmacokinetics was studied in whole blood in a parallel group of rats. The CsA blood concentrations were used as input to model P-gp inhibition. The inhibition of P-gp was observed as a rapid increase in brain concentrations of [(11)C]verapamil, with a maximum after 5, 7.5 and 17.5 min for the respective doses. The respective increases in maximal [(11)C]verapamil concentrations were 1.5, 2.5 and 4 times the baseline concentration. A model in which CsA inhibited P-gp by decreasing the transport of [(11)C]verapamil out from the brain resulted in the best fit. Our data suggest that it is not the CsA concentration in blood, but rather the CsA concentration in an effect compartment, probably the endothelial cells of the blood-brain barrier that is responsible for the inhibition of P-gp.