Functional expression of P-glycoprotein in rat brain microglia

In the central nervous system, the primary targets of the human immunodefic iency virus-1 (HIV-1) are microglia, resulting in a disorder called HIV-1 d ementia. P-glycoprotein (P-gp), a membrane-associated ATP-dependent efflux transporter, limits entry into the brain of numerous xenobiotics, including anti-HIV drugs (i.e., protease inhibitors). This project investigates the functional expression of P-gp in the endogenous immune cells of the brain, a parenchymal compartment not previously studied. We used a cell line (MLS- 9) derived from rat microglia to study the transport of digoxin, a known P- gp substrate. Reverse transcriptase-polymerase chain reaction analysis dete cted mRNA for only mdr1b in IMLS-9 cells, whereas both mdr1a and mdr1b mRNA were expressed in primary cultured microglia from which they were derived. Western blot analysis with the C219 antibody detected a single band at sim ilar to 170 to 180 kDa in MLS-9 cells, which is the size previously reporte d for P-gp. Immunocytochemical analysis with the monoclonal antibodies C219 , MRK16, and MAB-448 labeled P-gp protein along the plasma membrane and nuc lear envelope of MLS-9 cells. [H-3]Digoxin accumulation by monolayers of ML S-9 cells was significantly enhanced in the presence of any of several P-gp inhibitors (verapamil, cyclosporin A, quinidine, PSC 833), protease inhibi tors (i.e., saquinavir, indinavir, and ritonavir), and sodium azide, an ATP ase inhibitor. These results provide the first evidence for the functional expression of P-gp in microglia and imply that entry of pharmacological age nts, including protease inhibitors, may be prevented within the brain paren chyma, as well as at the blood-brain barrier.