Blood-brain barrier is involved in the efflux transport of a neuroactive steroid, dehydroepiandrosterone sulfate, via organic anion transporting polypeptide 2

We have investigated the transport characteristics of dehydroepiandrosteron e sulfate (DHEAS), a neuroactive steroid, at the blood-brain barrier (BBB) in a series of functional in vivo and in vitro studies. The apparent BBB ef flux rate constant of [H-3]DHEAS evaluated by the brain efflux index method was 2.68 x 10(-2) min(-1). DHEAS efflux transport was a saturable process with a Michaelis constant (K-m) of 32.6 mu M. Significant amounts of [H-3]D HEAS were determined in the jugular venous plasma by HPLC, providing direct evidence that most of the DHEAS is transported in intact form from brain t o the circulating blood across the BBB, This efflux transport of [H-3]DHEAS was significantly inhibited by common rat organic anion-transporting polyp eptide (oatp) substrates such as taurocholate, cholate, sulfobromophthalein , and estrone-3-sulfate. Moreover, the apparent efflux clearance of [H-3]DH EAS across the BBB (118 mu l/min-g of brain) was 10.4-fold greater than its influx clearance estimated by the in situ brain perfusion technique (11.4 mu l/min-g of brain), suggesting that DHEAS is predominantly transported fr om the brain to blood across the BBB. In cellular uptake studies using a co nditionally immortalized mouse brain capillary endothelial cell line (TM-BB B4), [H-3]DHEAS uptake by TM-BBB4 cells exhibited a concentration dependenc e with a K-m of 34.4 mu M and was significantly inhibited by the oatp2-spec ific substrate digoxin, Conversely, [H-3]digoxin uptake by TM-BBB4 cells wa s significantly inhibited by DHEAS, Moreover, the net uptake of [H-3]DHEAS at 30 min was significantly increased under ATP-depleted conditions, sugges ting that an energy-dependent efflux process may also be involved in TM-BBB 4. RT-PCR and sequence analysis suggest that an oatp2 is expressed in TM-BB B4 cells. In conclusion, DHEAS efflux transport takes place across the BBB, and studies involving in vitro DHEAS uptake and RT-PCR suggest that there is oatp2-mediated DHEAS transport at the BBB.