Objective: In sepsis, severe hypotension and hyporeactivity to vasoconstrictor agents involves over production of nitric oxide. Previously, we showed that the in vitro incubation of vessels with the nitric oxide donor S-nitroso-acetyl-DL-penicillamina (SNAP) induced a long-lasting hiporeactivity to alpha-adrenergic agonists, in association with induction of dose-dependent relaxation to the thiol, aminoacid L-cysteine (L-CYS). This study addressed possible mechanisms involved in L-CYS-mediated vasorelaxation. Method and Results: Rat aorta rings were prepared for isometric tension recording, cumulative concentration-response curves to L-CYS (CCR; 1, 10 and 100 µM) were constructed, in presence or absence of endothelium after precontraction with phenylephrine (1 nM - 100 µM). Rings were incubated with SNAP (200 µM; 30 min) and washed thereafter for 1 hours before L-CYS. L-CYS induced maximal relaxation  73,8 ± 7,0 % and 84,2 ± 4,4 % in rings with and without endothelium, respectively. The guanylate ciclase inhibitor (ODQ; 10 µM) reduced the relaxation due to L-CYS (34,7 ± 7,5 % to rings with endothelium and 4,0 ± 4,1 % to rings without endothelium). In addition, pre-incubation of vessels with pharmacological agents [KT5823, KT 5720 (1 µM; inhibitor of protein kinase G and A, respectively); tetraethylammonium (TEA; 10 mM; a non-selective inhibitor of K⁺ channels); 4-aminopyridine (1 mM; inhibitor of voltage-activated K⁺ channels); glibenclamide (10 µM; inhibitor of ATP-activated K⁺ channels ); clotrimazole (10 µM; inhibitor of intermediate Ca⁺²-activated K⁺ channels) and charybdotoxin (100 nM; inhibitor of intermediate and large conductance Ca⁺²-activated K⁺ channels)] did not affect the vasorelaxing effect of L-CYS. Importantly, 80 mM KCl totally inhibited relaxation induced by L-CYS, causing contraction of the rings in 4,3 ± 3,6 %. Conclusion: We conclude that relaxation to L-CYS after previous exposure to nitric oxide depends only partially on soluble guanylate cyclase activation and opening of an yet unidentified K⁺ channel sub-type, but not on protein kinase G and A pathways. Thus, the mechanisms for nitric oxide-dependent hyporeactivity may differ from those of vasorelaxation. Financial support: CAPES, CNPq and PRONEX.