Xenobiotic efflux pumps in isolated fish brain capillaries.
To identify specific transporters that drive xenobiotics from the central nervous system to blood, the accumulation of fluorescent drugs was studied in isolated capillaries from killifish and dogfish shark brain using confocal microscopy and quantitative image analysis. In killifish brain capillaries, luminal accumulation of fluorescent derivatives of cyclosporin A and verapamil was concentrative, specific, and energy dependent (inhibition by KCN). Transport was reduced by PSC-833, but not by leukotriene C4, indicating the involvement of P-glycoprotein. The ability of capillaries to transport the cyclosporin A derivative was unchanged over 20 h, demonstrating the long-term viability of the preparation. Luminal accumulation of the fluorescent organic anions sulforhodamine 101 and fluorescein-methotrexate was also concentrative, specific, and energy dependent. Transport of these compounds was reduced by leukotriene C4, but not by PSC-833, indicating the involvement of a multidrug resistance-associated protein (Mrp). Similar results were obtained for isolated capillaries from dogfish shark. Immunostaining localized P-glycoprotein and Mrp2 to the luminal surface of the killifish brain capillary endothelium. These findings validate a new and long-lived comparative model for studying drug transport across the blood-brain barrier and, as in mammals, implicate P-glycoprotein and Mrp2 in transport from the central nervous system to blood in fish.
Miller
David S
DS
Laboratory of Pharmacology and Chemistry, National Institute of Environmental Health Sciences, National Institutes of Health, Research Triangle Park, North Carolina 27709, USA. miller@niehs.nih.gov
Graeff
Claudia
C
Droulle
Lucy
L
Fricker
Stefanie
S
Fricker
Gert
G
D009327:4-Chloro-7-nitrobenzofurazan / Q000493:pharmacokinetics; D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000378:metabolism; D000818:Animals; D001812:Blood-Brain Barrier / Q000187:drug effects / Q000502:physiology*; D001896:Boron Compounds / Q000493:pharmacokinetics; D001921:Brain / Q000098:blood supply*; D002121:Calcium Channel Blockers / Q000493:pharmacokinetics; D002196:Capillaries / Q000502:physiology; D016572:Cyclosporine / Q000493:pharmacokinetics; D003524:Cyclosporins / Q000494:pharmacology; D004284:Dogfish; D004791:Enzyme Inhibitors / Q000493:pharmacokinetics; D005456:Fluorescent Dyes / Q000493:pharmacokinetics; D023781:Fundulidae; D066298:In Vitro Techniques; D007202:Indicators and Reagents / Q000493:pharmacokinetics; D017997:Leukotriene C4 / Q000494:pharmacology; D026901:Membrane Transport Proteins; D018613:Microscopy, Confocal; D000089762:Multidrug Resistance-Associated Protein 2; D027425:ATP-Binding Cassette, Sub-Family C Proteins / Q000378:metabolism; D011190:Potassium Cyanide / Q000494:pharmacology; D014700:Verapamil / Q000493:pharmacokinetics; D015262:Xenobiotics / Q000493:pharmacokinetics*
D009327:4-Chloro-7-nitrobenzofurazan; D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D001896:Boron Compounds; D002121:Calcium Channel Blockers; D016572:Cyclosporine; D003524:Cyclosporins; D004791:Enzyme Inhibitors; D005456:Fluorescent Dyes; D007202:Indicators and Reagents; D017997:Leukotriene C4; D026901:Membrane Transport Proteins; D000089762:Multidrug Resistance-Associated Protein 2; D027425:ATP-Binding Cassette, Sub-Family C Proteins; D011190:Potassium Cyanide; D014700:Verapamil; D015262:Xenobiotics; C095489:4,4-difluoro-4-bora-3a,4a-diaza-s-indacene; C070272:valspodar; C421109:multidrug resistance-associated protein 1