When the blood-brain barrier (BBB) transport of a series of model peptides that varied in their physicochemical properties (lipophilicity, size and hydrogen-bonding potential) was determined using an in situ rat brain perfusion technique, an unexpected increase in flux with increasing peptide concentration was observed with one of the peptides. Further, inclusion of verapamil in the perfusion medium also increased permeability of the peptides. These observations were consistent with the presence of a polarized efflux system in the BBB that was saturable, could be competitively inhibited and showed substrate specificity. Such properties are similar to those of P-glycoprotein (P-gp), an apically localized efflux pump that has recently been reported to be present in the endothelial cells that constitute the BBB, and suggest that P-gp may be responsible for this activity. By measuring the BBB transport of the model peptides in the presence of verapamil (a P-gp inhibitor), the intrinsic BBB permeabilities (due to passive diffusion only) were obtained. The presence of verapamil caused a significant increase in the BBB permeabilities of six of the seven model peptides. When the intrinsic permeability coefficients were correlated with several physicochemical parameters, it was shown that hydrogen bonding potential rather than lipophilicity had the greatest influence on the passive diffusion of these model peptides across the BBB. From these studies it can be concluded that inhibition of P-gp, as well as reduction of the hydrogen bonding potential, can be used as strategies to increase peptide transport across the BBB.