An electron microscopic immunocytochemical study was undertaken to clarify ultrastructural localization and translocation of nitric oxide synthase (NOS) in endothelial cells (EC) of the human cerebral and superficial temporal arteries (STA) employing antibody against endothelial NOS (EC-NOS). NOS immunoreactivity was found in all EC examined, in association with the plasma membrane and cytoplasmic organelles such as endoplasmic reticulum, Weibel-Palade body and subplasmalemmal vesicles, and in the cytoplasm devoid of organelles and extracellular regions, irrespective of arteries. The immunoreactivity in subplasmalemmal vesicles was, however, demonstrated only in human cerebral arteries. In the human STA exposed to bradykinin which induces EC-NOS phosphorylation, the gold particles significantly increased in the cytosol and decreased in the areas associated with cytoplasmic organelles; however, the number of particles did not change significantly in the plasma membrane. The results implicate that NOS may be translocated from the area associated with cytoplasmic organelles to cytosol following EC exposure to bradykinin.