Interaction of nonpeptidic delta agonists with P-glycoprotein by in situ mouse brain perfusion: liquid chromatography-mass spectrometry analysis and internal standard strategy.
Many opioids are substrates of the efflux transporter P-glycoprotein (P-gp) in the blood-brain barrier (BBB). In situ brain perfusion in wild-type and mdr 1a(-/-) P-gp-deficient mice was utilized to investigate potential P-gp-mediated transport of novel nonpeptidic delta agonists (AR-M delta compounds). Because radioactive compounds were not available for this series, liquid chromatography-mass spectrometric detection (LC-MS) was the assay methodology of choice. Verapamil in the perfusion buffer (0.5 microM) served as a positive control for P-gp-mediated efflux and as an experimental internal standard for P-gp modulation by AR-M delta compounds. LC-MS provided excellent assay sensitivity with no significant interferences. In P-gp-competent mice, the brain extraction of AR-M delta compounds ranged from 1.1 to 96%. The ratio of initial brain uptake clearances (Cl(up)) in P-gp-deficient and wild-type mice (P-gp effect) ranged from 0.96 to 4.91. Some compounds increased the Cl(up) of verapamil in P-gp-competent mice, consistent with P-gp inhibition. These results demonstrate that LC-MS is an appropriate assay methodology for mouse brain perfusion samples, that AR-M delta compounds may interact with P-gp in the BBB, and that the internal strategy can provide useful information concerning P-gp modulation by compounds of interest.
Dagenais
Claude
C
Division of Drug Delivery and Disposition, School of Pharmacy, University of North Carolina at Chapel Hill, 27599-7360, USA.
Ducharme
Julie
J
Pollack
Gary M
GM
D018435:ATP Binding Cassette Transporter, Subfamily B / Q000172:deficiency / Q000378:metabolism; D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D018528:ATP-Binding Cassette Transporters / Q000378:metabolism; D000818:Animals; D001921:Brain / Q000098:blood supply* / Q000378:metabolism*; D002121:Calcium Channel Blockers / Q000493:pharmacokinetics; D002853:Chromatography, Liquid / Q000379:methods / Q000706:statistics & numerical data; D008297:Male; D013058:Mass Spectrometry / Q000379:methods / Q000706:statistics & numerical data; D051379:Mice; D018345:Mice, Knockout; D010477:Perfusion / Q000379:methods; D017465:Receptors, Opioid, delta / Q000819:agonists* / Q000378:metabolism*; D014700:Verapamil / Q000493:pharmacokinetics
D018435:ATP Binding Cassette Transporter, Subfamily B; D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D018528:ATP-Binding Cassette Transporters; D002121:Calcium Channel Blockers; D017465:Receptors, Opioid, delta; C095122:multidrug resistance protein 3; D014700:Verapamil