CHARACTERIZATION OF EFFLUX TRANSPORT OF ORGANIC-ANIONS IN A MOUSE-BRAIN CAPILLARY ENDOTHELIAL-CELL LINE

Cumulative evidence suggests that several organic anions are actively effluxed from the brain to the blood across the blood-brain barrier (B BB). We examined the possibility of the presence of primary active tra nsporters for organic anions (multidrug resistance associated protein (MRP) and canalicular multispecific organic anion transporter (cMOAT)) on the BBB by measuring the ATP-dependent uptake of 2,4-dinitrophenyl -S-glutathione (DNP-SG) and leukotriene C-4 (LTC4) into membrane vesic les prepared from a cell line derived from mouse brain capillary endot helial cells (MBEC4). The ATP-dependent uptake of DNP-SG into the memb rane vesicles was osmotically sensitive and was also supported by GTP, but not by AMP or ADP. An ATPase inhibitor, vanadate, blocked the ATP -dependent uptake of DNP-SG. The ATP-dependent uptake process was satu rable, with K-m values of 0.56 and 0.22 mu M, and V-max values of 5.5 and 27.5 pmol/min/mg protein for DNP-SG and LTC4, respectively. Northe rn and Western blot analyses showed the expression of murine MRP but n ot cMOAT in MBEC4 cells. Western blot analysis of the rat cerebral end othelial cells indicated the expression of protein(s) that is detectab le with MRPr1, an antibody against MRP. These results, together with p revious Findings that both DNP-SG and LTC, are good ligands for MRP, s uggest that MRP is responsible for the unidirectional, energy-dependen t efflux of organic anions from the brain into the circulating blood a cross the BBB.