The effects of nitroblue tetrazolium (NBT), a well-known scavenger of superoxide anions and an inhibitor of nicotinamide adenine dinucleotide (NADPH)-dependent oxidations, were assessed on the metabolism of glyceryl trinitrate (GTN) to nitric oxide (NO) by bovine aortic smooth muscle cells (SMC). The extent of this metabolism was determined by measuring NO formed, using the inhibition of thrombin-induced platelet aggregation and relaxation of rabbit aortic strips as bioassay systems. In addition, NO produced from GTN by SMC was measured as nitrite (NO2-), one of its breakdown products. The antiplatelet effect of GTN (44 microM) was potentiated by SMC (0.12-0.46 x 10(5) cells) treated with indomethacin (10 microM) and this was inhibited in a concentration-dependent manner when the cells were pretreated with NBT (100 microM). NBT (3-100 microM) also reduced the formation of NO2- from GTN (600 microM) by SMC (3 x 10(5) cells). Furthermore, relaxations of endothelium-denuded strips of the rabbit aorta by GTN (10(-9)-10(-6) M) were attenuated when the strips were pretreated with NBT (100 or 500 microM). The formation of NO from L-arginine (L-Arg) by SMC was not affected by NBT. The hypotensive responses to GTN (0.25-1 mg/kg, i.v.) in anaesthetized rats were inhibited by pretreatment with NBT (1.25 mg/kg, i.v.) but NBT did not alter the hypotensive responses induced by SIN-1. Thus, NBT inhibited the bioconversion of GTN to NO both in vitro and in vivo. NBT may be a useful pharmacological tool to investigate the enzymic pathway(s) involved in the conversion of GTN to NO by smooth muscle cells or other cells.