Previous work has shown that endothelium-dependent dilation is maintained in cerebral resistance arteries with age; however, the mechanisms that regulate nitric oxide bioavailability may be altered by age and exercise training. PURPOSE: The purpose of this study was to determine the effects of age and exercise training on mechanisms that regulate endothelial nitric oxide availability in cerebral resistance arteries. METHODS: Young (4 mo) and old (24 mo) male Fischer-344 rats underwent exercise training or remained sedentary for 10 weeks. Vasodilation to acetylcholine (ACh) was determined in posterior communicating arteries (PCoA) under control conditions and during treatment with 1-(2-trifluoromethylphenyl) imidazole (TRIM) and aminoguanidine (AG) to block neuronal nitric oxide synthase (nNOS) and inducible nitric oxide synthase (iNOS), respectively. Responsiveness to ACh was also evaluated in the presence and absence of Tiron, a cell permeable form of superoxide dismutase. RESULTS: In PCoA from both young and old rats, non-specific blockade of NOS with NG-nitro-L-arginine methyl ester (L-NAME) eliminated dilation to ACh (p=0.0001). Simultaneous treatment with TRIM and AG eliminated ACh-induced dilation in PCoA from both sedentary and exercise trained old rats; however, significant dilation remained in the presence of TRIM and AG in young sedentary and exercise trained rats (p=0.003 and p=0.0001; respectively). Tiron enhanced vasodilation to ACh in PCoA from both sedentary and exercise trained old rats (p=0.008 and p=0.003; respectively), but not in PCoA from young rats, regardless of training status. CONCLUSIONS: These results, along with similar measurements of basal and ACh-stimulated NO production in PCoA from young and old rats, suggest the bioavailability of NO in PCoA decreases with age possibly due to an increase in superoxide. Furthermore, these data indicate that exercise training upregulates eNOS activity in PCoA of young but not old rats. In PCoA of old rats, the ability of exercise training to increase eNOS activity may be limited by the presence of greater oxidant stress. Supported by NIH R01HL077224 and AHA GIA0255956Y