Atherosclerotic arteries are well known to exhibit impaired endothelium-dependent relaxation (EDR), but the exact mechanism of this impairment remains unclear. Recently, endothelial constitutive nitric oxide synthase (ECNOS), which generates nitric oxide and mediates EDR, was cloned, and ECNOS mRNA expression was reported to be modified by various cytokines, lipoproteins, and shear stress. To investigate the expression of ECNOS mRNA and protein in atherosclerotic arteries with impaired EDR, thoracic aortas isolated from Watanabe heritable hyperlipidemic (WHHL) rabbits were examined by using in situ hybridization and immunohistochemistry. Compared with thoracic aortas from Japanese White rabbits, WHHL aortas exhibited significantly impaired EDRs, although both in situ hybridization and immunohistochemistry exhibited enhanced expression of ECNOS mRNA and protein in WHHL aortas. There was no significant relationship between expression of ECNOS mRNA and protein of endothelial cells and age of the examined WHHL aortas. These data suggest that the mechanism of impaired EDR in atherosclerotic arteries is not due to the decrease in ECNOS mRNA and protein.