MECHANISMS OF ENDOTHELIN-INDUCED MACROMOLECULAR LEAKAGE IN MICROVASCULAR BEDS OF RAT MESENTERY

Microvascular responses to endothelin-3 were investigated in the rat mesentery under fluorescence microscopy. Endothlin-3 in a range of 0.1-100 pM induced arteriolar constriction in a dose-dependent manner. and stimulated Ca2+ niobilization, demonstrated by fura-2-associated fluorography, in both arterioles and venules. Cyclo(-D-Trp-D-Asp-Pro-D-Val-Leu-) (BQ123), an endothelin ET(A) receptor antagonist, at a concentration of 10 muM inhibited the endothelin-3 (100 pM)-induced arteriolar constriction and Ca2+ mobilization in arterioles but not in venules. In venules. an early onset leakage of FITC (fluorescein isothiocyanate)-labeled albumin and subsequent reduction of red blood cell velocity without arteriolar constriction were observed after the superfusion of endothelin-3 with BQ123, suggesting that a non-endothelin ET(A) receptor mediates macromolecular leakage followed by a decrease in blood flow. Endothelin-3 with BQ123 neither stimulated leukocyte adhesion nor activated luminol-dependent chemiluminescence in venules, showing that endothelin-3-increased permeability may be induced by leukocyte-independent and oxyradical-independent mechanisms. These microvascular alterations of permeability and red blood cell velocity were significantly attenuated by the addition of phalloidin, an F-actin stabilizer, suggesting the involvement of endothelial cell contraction. Nicardipine (1,4-dihydro-2,6-dimethyl-4-[3-nitrophenyl]methyl-2-[methyl(phenylmethyl)amino]-3,5-pyridinedicarboxylic acid ethyl ester), a dihydropyridine-type Ca2+ channel antagonist, eliminated endothelin-3-induced arteriolar constriction; however, it did not affect albumin leakage promoted by endothelin-3 with BQ123, suggesting that a non-voltage-dependent Ca2+ channel(s) is involved in non-endothelin ET(A) receptor-mediated Ca2+ mobilization and contraction of venular endothelial cells. Overall, it is conceivable that endothelin ET(A) receptor and voltage-dependent Ca2+ channel are involved in endothelin-3-induced arteriolar constriction. In addition, the present results suggest that Ca2+ mobilization in venular endothelium, which is mediated by a non-endothelin ET(A) receptor, possibly endothelin ET(B) receptor and regulated by non-voltage-dependent Ca2+ channel(s), may cause endothelial cell contraction and subsequently increase macromolecular permeability in microvascular beds treated with endothelin-3.