Valproic acid uptake by bovine brain microvessel endothelial cells: role of active efflux transport.
The basis for low brain permeability of valproic acid (VPA) appears to be the result of efflux transport at the blood-brain barrier (BBB); however, the identity of the putative efflux transporter has not been investigated. The objective of our studies was to determine whether the multidrug resistance-associated protein (MRP) might be involved in efflux transport of VPA. Brain microvessel endothelial cells (BMEC) were isolated from cow brains and grown to confluence. MRP messenger RNA (mRNA) in BMEC were verified by reverse transcriptase-polymerase chain reaction (RT-PCR). Functional activity was demonstrated using the steady-state retention of calcein and MRP inhibitors, indomethacin (IND) and probenecid (PRB). Probenecid (0.50 mM) and indomethacin (10 microM) produced a 26 and 13% ( P<0.05 ) elevation in steady-state cellular VPA uptake following a 30-min-incubation with tracer 3H-VPA and 30 microM cold VPA. In contrast, at higher concentrations of probenecid (2 mM) and indomethacin (500 microM), an 11 and 31% reduction in VPA uptake was observed. The biphasic pattern of VPA uptake suggested concurrent inhibition of uptake and efflux transporters by the inhibitor with differing sensitivities, i.e. the efflux transporter being more susceptible to inhibition than the influx transporter. Similar results were obtained in the MRP overexpressing cell line A549. Overall, the results suggest that MRP(s) is(are) involved in the efflux transport of VPA, but do not preclude the possible contribution(s) of other organic anion transporters. The findings also adds to the growing evidence that up-regulation of active drug efflux transporters at the BBB may contribute to the development of drug resistance to antiepileptic drug therapy.
Gibbs
John P
JP
Department of Pharmaceutics, University of Washington, Seattle, WA, USA. john_p_gibbs@groton.pfizer.com
Adeyeye
Moji Christianah
MC
Yang
Ziping
Z
Shen
Danny D
DD
D000230:Adenocarcinoma; D000818:Animals; D001693:Biological Transport, Active / Q000502:physiology; D001921:Brain / Q000166:cytology*; D002417:Cattle; D002478:Cells, Cultured; D016861:Cyclooxygenase Inhibitors / Q000494:pharmacology; D004140:Dinitrophenols / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004347:Drug Interactions; D042783:Endothelial Cells / Q000187:drug effects* / Q000378:metabolism; D005452:Fluoresceins / Q000378:metabolism; D018682:GABA Agents / Q000494:pharmacology*; D006801:Humans; D007213:Indomethacin / Q000494:pharmacology; D008175:Lung Neoplasms; D027425:ATP-Binding Cassette, Sub-Family C Proteins / Q000378:metabolism; D010190:Pancreatic Neoplasms; D011339:Probenecid / Q000494:pharmacology; D012333:RNA, Messenger / Q000096:biosynthesis; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D012402:Rotenone / Q000494:pharmacology; D013696:Temperature; D014316:Tritium / Q000493:pharmacokinetics; D014475:Uncoupling Agents / Q000494:pharmacology; D014528:Uricosuric Agents / Q000494:pharmacology; D014635:Valproic Acid / Q000493:pharmacokinetics / Q000494:pharmacology*
D016861:Cyclooxygenase Inhibitors; D004140:Dinitrophenols; D005452:Fluoresceins; D018682:GABA Agents; D007213:Indomethacin; D027425:ATP-Binding Cassette, Sub-Family C Proteins; D011339:Probenecid; D012333:RNA, Messenger; D012402:Rotenone; D014316:Tritium; D014475:Uncoupling Agents; D014528:Uricosuric Agents; D014635:Valproic Acid; C007740:fluorexon