To examine the direct effects of steroids on vascular smooth muscle, rat aortic vascular smooth muscle cells were incubated in culture either steroid-free, or with natural and synthetic corticosteroids (RU26988 were incubated, dexamethasone, corticosterone, 9.alpha.-fluorocortisol, aldosterone, dexoycorticosterone) or sex steroids (estradiol, 5.alpha.-dihydrotestosterone). At the end of 24 h, cultures were pulsed with [35S]methionine for 2 h, the cells lysed, and patterns of incorporation of isotope into protein determined by 2-dimensional gel electrophoresis and autoradiography. Neither estradiol nor 5.alpha.-dihydrotestosterone altered protein synthetic profiles compared with control (steroid-free) incubations. In contrast, cultures exposed to the 6 corticosteroids at 10-7 M showed an identical pattern of response (6 proteins increased, 6 proteins decreased). This response appears to be glucocorticoid specific, since the mineralocorticoids (9.alpha.-fluorocortisol, aldosterone and deoxycorticosterone) did not have any effects over and above those seen with the pure glucocorticoid RU26988. These data are interpreted as evidence for a putative glucocorticoid domain of at least 12 proteins in rat vascular smooth muscle cells. In contrast, there appear to be no comparable estrogen-, androgen- or mineralocorticoid-specific changes in these cells.