The cyclic cationic octapeptide octreotide is known to be taken up by hepatocytes, with more than 70% of an intravenous dose being excreted into bile in unchanged form. We have already reported that a transporter other than the canalicular multispecific organic anion transporter (cMOAT) is responsible for the biliary excretion of octreotide in vivo. The aim of this study is to obtain an insight into the transporter of octreotide in bile canalicular membrane. The effect of various compounds on the ATP-dependent uptake of octreotide by rat bile canalicular membrane vesicles (CMV) was investigated. The ATP-dependent uptake of [14C]octreotide by CMV was inhibited by verapamil, vincristine and PSC833 in a concentration-dependent manner, the maximum inhibitory effects of these compounds being almost equal to that of excess unlabeled octreotide, while taurocholic acid (TCA) caused no inhibition at concentrations much higher than the Km of TCA uptake by CMV. Mutual inhibition between octreotide and dinitrophenylglutathione (DNP-SG), a representative substrate for cMOAT was only minor and could only be observed at concentrations much higher than the Km for each ligand uptake. To examine the contribution of P-glycoprotein to the biliary excretion of octreotide in vivo, biliary excretion of octreotide was compared between P-glycoprotein-induced rats by phenothiazine (PTZ) treatment and normal rats. A significant increase in the biliary excretion rate was observed in PTZ-treated rats. Only a slight decrease in biliary excretion was observed in mdr1a knock-out mice compared with normal mice, which may be explained by the associated induction of mdr1b. These results demonstrate that the transporter for octreotide is different from cMOAT and the bile acid transporter. The involvement of P-glycoprotein in the biliary excretion of octreotide is suggested.