OBJECTIVE The aim was to investigate the effects of adenosine on nitric oxide (NO) synthesis in vascular smooth muscle cells. METHODS NO and cAMP synthesis was measured in confluent rat vascular smooth muscle cells in culture at passage 5-10, using Griess reagent and an enzyme immunoassay kit, respectively. The expression of inducible NO synthase mRNA was assayed by Northern blotting. RESULTS Incubation of cultures with interleukin-1 beta (10 ng/ml) for 24 h caused a significant increase in nitrite production. The interleukin-1 beta-induced nitrite production by vascular smooth muscle cells was significantly increased by adenosine or its stable analogue, 2-chloroadenosine, in a dose-dependent manner. The adenosine A2a receptor antagonist, KF17837, but not the A1 receptor antagonist, DPCPX, significantly inhibited 2-chloroadenosine-mediated nitrite production. The 2-chloroadenosine-mediated nitrite production by interleukin-1 beta-stimulated cells was accompanied by increased inducible NO synthase mRNA accumulation. In the presence of dibutyryl-cAMP (1 mM), interleukin-1 beta-induced nitrite accumulation was further increased, but the effect of 2-chloroadenosine was not additive or synergistic. Addition of 2-chloroadenosine dose-dependently increased intracellular cAMP levels of vascular smooth muscle cells. CONCLUSIONS These results indicate that adenosine acts on A2 receptors and augments NO synthesis in interleukin-1 beta-stimulated vascular smooth muscle cells, at least partially through a cAMP-dependent pathway.