Received for publication April 16, 2004.
Revised June 3, 2004.
Accepted for publication June 3, 2004.

RAPID REGULATION OF P-GLYCOPROTEIN AT THE BLOOD-BRAIN BARRIER BY ENDOTHELIN-1

Abstract

The ATP-driven xenobiotic transporter, p-glycoprotein, is a critical element of the blood-brain barrier. To study regulation of p-glycoprotein function, we measured specific transport (PSC833-sensitive) of a fluorescent cyclosporine A derivative (NBDL-CSA) into the lumens of isolated rat brain capillaries using confocal microscopy and quantitative image analysis. Luminal NBDL-CSA accumulation was rapidly and reversibly reduced in a concentration-dependent manner by 0.1-100 nM endothelin-1 (ET-1). In this concentration range, ET-1 did not affect junctional permeability. The ET_B receptor agonist, sarafotoxin 6c, also reduced transport. Effects of ET-1 and sarafotoxin 6c were blocked by an ET_B receptor antagonist; an ET_A receptor antagonist was without effect. Consistent with this, immunostaining and Western blotting showed expression of the ET_B receptor in brain capillary membranes. NBDL-CSA transport was also reduced by sodium nitroprusside (SNP), a NO donor, and by phorbol ester, a protein kinase C (PKC) activator. Inhibition of NO synthase (NOS) or PKC abolished the ET-1 effects. Thus, ET-1, acting through an ET_B receptor, NOS and PKC, rapidly and reversibly reduced transport mediated by p-glycoprotein at the blood-brain barrier.

Key words: Endothelin, Nitric oxide, Protein Kinase C, MDR/p-Glycoprotein, Fluorescence techniques