Relationship between permeability status of the blood–brain barrier and in vitro permeability coefficient of a drug

Objective: The aim was to test the hypothesis that the assessment of basal and drug-induced changes in permeability of the blood–brain barrier (BBB) during in vitro drug transport assays is essential for an accurate estimation of the permeability coefficient of a drug. Methods: An in vitro BBB model was used, comprising of brain capillary endothelial cells (BCEC) and astrocytes co-cultured on semi-permeable filter inserts. Experiments were performed under control and challenged experimental circumstances, induced to simulate drug effects. The apparent BBB permeability coefficient for two markers for paracellular drug transport, sodium fluorescein ( P app,FLU, M w 376 Da) and FITC-labeled dextran ( P app,FD4, M w 4 kDa), was determined. Transendothelial electrical resistance (TEER) was used to quantify basal and (simulated) drug-induced changes in permeability of the in vitro BBB. The relationship between P app and TEER was determined. Drug effects were simulated by exposure to physiologically active endogenous and exogenous substances (i.e., histamine, deferroxamine mesylate, adrenaline, noradrenaline, bradykinin, vinblastine, sodium nitroprusside and lipopolysaccharide). Results: P app,FLU and P app,FD4 in control experiments varied from 1.6 up to 17.6 (10 −6cm/s) and 0.3 up to 7.3 (10 −6cm/s), respectively; while for individual filters P app,FLU was 4 times higher than P app,FD4 ( R 2=0.97). As long as TEER remained above 131·Ω cm 2 for FLU or 122·Ω cm 2 for FD4 during the transport assay, P app remained independent from the basal permeability of the in vitro BBB. Below these TEER values, P app increased exponentially. This nonlinear relationship between basal BBB permeability and P app was described by a one-phase exponential decay model. From this model the BBB permeability status independent permeability coefficients for FLU and FD4 ( P FLU and P FD4) were estimated to be 2.2±0.1 and 0.48±0.03 (10 −6cm/s), respectively. In the experimentally challenged experiments, a reliable indication for P FLU and P FD4 could be estimated only after the (simulated) drug-induced change in BBB permeability was taken into account. Conclusions: The assessment of basal BBB permeability status during drug transport assays was essential for an accurate estimation of the in vitro permeability coefficient of a drug. To accurately extrapolate the in vitro permeability coefficient of a drug to the in vivo situation, it is essential that drug-induced changes in the in vitro BB