Resistance of endothelium-dependent relaxation to elevation of O(-)(2) levels in rabbit carotid artery.

Abstract

Endogenous superoxide anion (O(-)(2)) interferes with the bioactivity of nitric oxide (NO) in endothelium-dependent arterial relaxation (EDR). Using the lucigenin chemiluminescence assay, we measured O(-)(2) in the thoracic and abdominal aortas and the carotid artery of rabbits to determine whether ambient O(-)(2) varies among the three arteries and differentially diminishes the effect of NO. Basal levels of O(-)(2) were significantly higher in carotid arteries than in the thoracic aorta [23 +/- 6.1 vs. 3.9 +/- 1.4 chemiluminescence units (CU); P < 0.05], whereas EDR in response to ACh (10(-8)-10(-5) M) was not significantly different on ANOVA. After treatment with the superoxide dismutase (SOD) inhibitor diethyldithiocarbamate (DDC; 10 mM), O(-)(2) levels were significantly elevated, becoming greater in the carotid artery and abdominal aorta than in the thoracic aorta (185 +/- 31.2 and 202 +/- 40.3 vs. 89 +/- 18 CU; P < 0.05). DDC significantly reversed EDR in the thoracic aorta but not in the carotid artery; at 10(-6) M ACh, the decrease seen with DDC was 48 +/- 6.2 vs. 6.8 +/- 8.0% of maximal relaxation in the thoracic aorta and carotid artery, respectively. In the thoracic aorta, exogenous SOD reversed the inhibition of EDR caused by DDC. Moreover, DDC/O(-)(2)-resistant EDR in the carotid artery was ablated by the addition of nitro-L-arginine methyl ester (300 microM; P < 0.05), an NO synthase inhibitor, consistent with peroxynitrite or an O(-)(2)-resistant NO donor being involved in carotid relaxation. Indeed, exogenous peroxynitrite caused similar relaxation of the carotid artery and thoracic aorta, which was unaffected by DDC. Our studies show a greater production of nitrite and O(-)(2) per unit area by the carotid artery, suggesting a greater amount of their product peroxynitrite. These findings support the hypothesis that peroxynitrite is the relaxing agent that resists high O(-)(2) in the carotid artery.