The NO scavenger, N-methyl-D-glucamine dithiocarbamate-Fe2+ (MGD-Fe), was used to characterize spontaneous and agonist-stimulated NO activity arising from rat aortic endothelium. Scavenging of NO in solution by MGD-Fe without tissue was confirmed by electron paramagnetic resonance spectroscopy. The addition of MGD-Fe to aortic ring segments suspended in vitro under resting tension did not cause any direct contraction, but when added to rings contracted with phenylephrine elicited a rapid additional increment in tension which was not altered by indomethacin. This increase in tension was absent in rings pretreated with L-nitroarginine or in rings without endothelium; however, the developed tension in both instances was similar in magnitude to that seen after MGD-Fe suggesting complete scavenging of basal NO production. Acetylcholine produced relaxation (maximum relaxation = 100 +/- 1%) which was completely eliminated in the presence of L-nitroarginine or upon removal of the endothelium. Relaxation to acetylcholine was reduced to 56 +/- 8% in the presence of MGD-Fe (3.0/0.6 mM ratio). Higher concentrations of MGD-Fe caused further decreases in the maximum relaxation to acetylcholine; however, a residual component (approx. 30%) of relaxation persisted. This effect was mimicked by using another NO trapping agent, carboxy-PTIO. These data suggest that MGD-Fe scavenges agonist-stimulated NO, but also reveals a NO synthase-dependent component which is unavailable to interact with MGD-Fe.