Fentanyl is a basic amine shown to have extensive first-pass pulmonary uptake. To evaluate the role of the pulmonary endothelium in this uptake process, the simultaneous pharmacokinetics of [3H]fentanyl and two marker drugs, blue dextran, and [14C]antipyrine, were evaluated in a flow-through system of pulmonary endothelial cells. Fentanyl equilibrium kinetics were determined in a static culture system. The flow-through system consisted of monolayers of bovine pulmonary artery endothelial cells cultured on solid microcarrier beads placed in a chromatography column and perfused at 1.0 ml/min (37 degreesC). Fentanyl and the markers were injected into the perfusate at the top of the column and samples were collected from the eluate at 9-s intervals for 10 min. The pharmacokinetic analyses were based on determinations of mean transit time and flow. Fentanyl was partitioned into the pulmonary endothelial cells 60 times more than the tissue water space marker antipyrine. In the static system, monolayers of bovine pulmonary artery endothelial cells were cultured in 3.8-cm2 wells to which were added 0 to 946 micromol (0-500 microgram/ml) of unlabeled fentanyl citrate and 0.14 micromol of [3H]fentanyl. After a 10-min incubation, solubilized cells were assayed for [3H]fentanyl. Pulmonary endothelial cells contained a higher relative fentanyl concentration at lower fentanyl supernatant concentrations than would be expected if uptake occurred by diffusion alone. These observations can be explained with a model of fentanyl uptake that includes both passive diffusion and saturable active uptake. This suggests that the extensive first-pass pulmonary uptake of fentanyl observed in vivo is due largely to vascular endothelial drug uptake by both a passive and a saturable active uptake process.