Xenobiotic Transport across Isolated Brain Microvessels Studied by Confocal Microscopy

Abstract

To identify specific transporters that drive xenobiotics from central nervous system to blood, the accumulation of fluorescent drugs was studied in isolated capillaries from rat and pig brain using confocal microscopy and quantitative image analysis. Luminal accumulation of daunomycin and of fluorescent derivatives of cyclosporine A (CSA) and ivermectin was concentrative, specific, and energy-dependent (inhibition by NaCN). Transport was reduced by PSC 833, ivermectin, verapamil, CSA, and vanadate, but not by leukotriene C(4) (LTC(4)), indicating the involvement of P-glycoprotein. Luminal accumulation of the fluorescent organic anions sulforhodamine 101 and fluorescein methotrexate was also concentrative, specific, and energy-dependent. LTC(4), chlorodinitrobenzene, and vanadate reduced transport of these compounds, but PSC 833 and verapamil did not, indicating the involvement of a multidrug resistance-associated protein (Mrp). Immunostaining localized P-glycoprotein and Mrp2 to the luminal surface of the capillary endothelium and quantitative polymerase chain reaction showed Mrp1 and Mrp2 expression. Finally, the HIV protease inhibitors saquinavir and ritonavir were potent inhibitors of transport mediated by both P-glycoprotein and Mrp. These results validate a new method for studying drug transport in isolated brain capillaries and implicate both P-glycoprotein and one or more members of the Mrp family in drug transport from central nervous system to blood.

Peptide Mimetic HIV Protease Inhibitors Are Ligands for the Orphan Receptor SXR

“…This protein is responsible for the biliary excretion of a number of drug metabolites. Interestingly, HIV protease inhibitors serve as substrates for MRP2 (3,4), as well as for other SXR target genes including CYP3A4 and P-glycoprotein (1).…”

“…Any protease inhibitor that escapes P-glycoprotein-mediated efflux enters the portal blood where it can ultimately be degraded by the hepatic cytochrome P450 CYP3A4. HIV protease inhibitors may also serve as substrates for MRP2 (ABCC2) (3,4), an export pump expressed on the bile canalicular membrane (5). It is thus possible that protease inhibitors may also be cleared by MRP2-dependent biliary secretion.…”

The apical conjugate efflux pump ABCC2 (MRP2)

“…However, ABCC2 was identified in brain capillaries of hippocampus specimens from patients with temporal lobe epilepsy [2] and in the rat blood-brain barrier after pilocarpine-induced epileptic seizures [52]. Moreover, some studies described Abcc2 in brain capillaries of normal rats [115,176]. Various human tissues do not express the ABCC2 protein in detectable amounts, such as the skin, exocrine pancreas, female reproductive system, lymphatic system, cardiovascular system, and connective tissue [147].…”

The MRP family of drug efflux pumps

“…In spite of the similarity in substrate range, the functions of MRP2 in the body are distinct from those of MRP1 as a result of differences in expression pattern and subcellular polarity. In contrast to MRP1, MRP2 assumes apical localization in polarized cells (Figure 2), and it is mainly expressed in liver canaliculi, with lower levels in renal proximal tubules, gut enterocytes, syncytiotrophoblast cells of the placenta and possibly brain capillaries (Kartenbeck et al, 1996;Schaub et al, 1997;Miller et al, 2000;Mottino et al, 2000;St-Pierre et al, 2000). Therefore, it is functionally similar to Pgp in its involvement in the terminal elimination of compounds and its role as a barrier in gut and placenta.…”