1. The influence of chronic treatment with molsidomine, pro-drug of the nitric oxide (NO) donor, 3-morpholino-sydnonimine (SIN-1), on fatty streak development and release of NO and prostacyclin (PGI2) was studied in the aorta of normal and cholesterol-fed rabbits. 2. Groups of 10 rabbits received standard diet (150 g day-1), or diets with 0.3% cholesterol, with 0.02% molsidomine or with the combination of cholesterol and molsidomine for 16 weeks. Lesion area and thickness, maximum change in isometric force (Emax) and sensitivity (-log EC50 or pD2) to constricting and relaxing agonists were assessed in segments of arch, thoracic and abdominal aorta. Bioassay was used to assess NO release. 3. Cholesterol-induced fatty streaks tapered off towards the abdominal aorta. Area, thickness, weight and cholesterylester content of the lesions were augmented by the NO donor, whereas the hypercholesterolaemia remained unchanged. The exacerbation was attributed to co-release of superoxide anion from the sydnonimine. 4. As fatty streaks progressed, amplitude and pD2 of acetylcholine (ACh)-induced relaxations decreased, whereas cyclic GMP and cyclic AMP second messenger systems were not influenced, since Emax and sensitivity to SIN-1 and forskolin remained unchanged. However, extensive lesions apparently trapped some NO, as the pD2 of authentic NO decreased. 5. The fatty streaks curtailed the biosynthesis of PGI2 and the overflow of NO from the perfused thoracic aorta. The latter defect was not restored by L-arginine and appears to be consistent with a functional change of the endothelial muscarinic receptors. 6. The NO donor desensitized the aorta to cyclic GMP-mediated relaxations (ACh, SIN-1 and NO), without affecting cyclic AMP-mediated relaxation to forskolin or constrictor responses to phenylephrine and 5-hydroxytryptamine. 7. The drug also suppressed the ACh-induced overflow of NO, without changing PGI2 release. This selective reduction of endothelial NO release and the desensitization of cyclic GMP-mediated relaxations occurred independently of fatty streak formation. 8. The results indicate that chronic exposure to exogenous NO downregulates endothelial NO release and cyclic GMP-mediated relaxations, and provide evidence for the existence of negative feed-back regulations of the L-arginine NO pathway under in vivo conditions.