Classically, drug penetration through the blood‐brain barrier depends on the lipid solubility of the substance, except for some highly lipophilic drugs, like colchicine and vinblastine, both substrates of P‐glycoprotein, a drug efflux pump present at the luminal surface of the brain capillary endothelial cells. Colchicine and vinblastine uptake into the brain was studied in the rat using the in situ brain perfusion technique and two inhibitors of P‐glycoprotein, verapamil and SDZ PSC‐833. When rats were pretreated with PSC‐833 (10 mg/kg, intravenous bolus), colchicine and vinblastine uptake was enhanced 8.42‐ and 9.08‐fold, respectively, in all the gray areas of the rat brain studied. The mean colchicine distribution volume was increased from 0.67 ± 0.41 to 5.64 ± 0.70 µl/g and vinblastine distribution volume from 2.74 ± 1.15 to 24.88 ± 4.03 µl/g. When rats were pretreated with verapamil (1 mg/kg, intravenous bolus), colchicine distribution volume was increased 3.70‐fold. The increase in colchicine and vinblastine did not differ between the eight brain gray areas. PSC‐833 and verapamil pretreatment had no influence on the distribution volume of either drug in the choroid plexus. Nevertheless, distribution volumes remained small, considering the highly lipophilic nature of the substances. We suggest that P‐glycoprotein is either only partially inhibited (difficulty of fully saturating P‐glycoprotein, especially under in vivo conditions) or not the only barrier to these two drugs.