Abstract
Cumulative evidence suggests that several organic anions are excreted from the brain to the blood across the blood-brain barrier. In the present study, we carried out a kinetic investigation of the transport activity in MBEC4, an immortalized cell line established from BALB/c mouse cerebral microvessel endothelial cells. The presence of an efflux system in intact cells was examined by using monochlorobimane (MCB), which is conjugated with glutathione intracellularly to produce glutathione bimane (GS-B). The efflux of GS-B was inhibited by ATP depletion and also by 1-chloro-2,4-dinitrobenzne, a precursor of 2,4-dinitrophenyl-S-glutathione, in a concentration-dependent manner. Using this MBEC4 monolayer, we investigated the direction of this transport activity. Although the efflux of GS-B was observed on both luminal and abluminal sides of MBEC4 monolayer, the profile differed for the two sides with respect to the concentration dependence of MCB; the analysis suggested the presence of high-affinity transport system on the luminal side. To investigate the mechanism for the transport, we examined the ATP-dependent uptake of GS-B into the membrane vesicles prepared from MBEC4. ATP-dependent uptake systems with high (Km = 35 nM) and low (Km = 14 μM) affinities were identified. These results suggested that this high-affinity transport system of glutathione conjugates is expressed on the luminal side of the blood-brain barrier and is involved in the detoxification of xenobiotics.
Footnotes
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Send reprint requests to: Yuichi Sugiyama, Doctor of Philosophy, Graduate School of Pharmaceutical Sciences, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan.
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↵1 This work was supported in part by a grant-in-aid from the Ministry of Education, Science, Sports and Culture of Japan, and the Core Research for Evolutional Sciences and Technology of Japan Sciences and Technology Corporation.
- Abbreviations:
- BBB
- blood-brain barrier
- CDNB
- 1-chloro-2,4-dinitrobenzene
- CNS
- central nervous system
- DNP-SG
- 2,4-dinitrophenyl-S-glutathione
- GS-B
- glutathione bimane
- GS-X
- glutathione S-conjugates
- MCB
- monochlorobimane
- MRP
- multidrug resistance- associated protein
- 3-OMG
- 3-O-methyl-D-glucose
- Received March 24, 1998.
- Accepted July 31, 1998.
- The American Society for Pharmacology and Experimental Therapeutics
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