The possible involvement of nitric oxide in mediating effect of 2-hrs immobilization stress (IMO) on testosterone production was investigated. IMO significantly decreased serum testosterone level and inhibited basal as well as hCG-stimulated testosterone production in vitro. These effects were accompanied by significant increase of nitrite, a stabile oxidation product of nitric oxide, in the testes incubation media. Bilateral intratesticular Nsup(o)-nitro-L-arginine methyl ester (NAME; 600 microgram/testis) treatment, 30 minutes before exposure to IMO, partially reduced IMO-induced increase in nitrite levels but prevented IMO-induced decrease of serum testosterone levels in vitro hCG-stimulated testosterone production, without affecting basal testosterone production. NAME treatment itself did not alter serum testosterone levels or in vitro basal and hCG-stimulated testosterone production in freely moving rats. These results indicate that nitric oxide could be involved in IMO impaired testicular steroidogenesis.