Investigation of role for oxidant stress in vascular tolerance development to glyceryl trinitrate in vitro

1 The role of reactive oxygen species (ROS) during the development of vascular cellular tolerance to glyceryl trinitrate (GTN), was studied in the rat isolated aorta. 2 Nitrate tolerance induced by a 30 min incubation with GTN (30 or 100 mu M) in vitro, was not affected by pretreatment with the intracellular superoxide anion scavenger, tiron (10 mM), or the intracellular scavenger of peroxynitrite anion and hydroxyl radical, dimethylsulphoxide (DMSO, 0.2% v v(-1)). In contrast, pretreatment with the intracellular sulphydryl donor, N-acetyl-L-cysteine (NAC, 1 mM), significantly attenuated GTN-induced tolerance. 3 Pretreatment with a putative inhibitor of oxidant stress-mediated, transcription factor NF-kappa B activation, pyrrolidine dithiocarbamate (PDTC, 50 mu M), an inhibitor of gene activation by NF-kappa B, dexamethasone (1 mu M) or an inhibitor of protein synthesis, cycloheximide (10 mu M), failed to affect tolerance development to GTN. 4 Pretreatment with DMSO (0.2% v v(-1)) or PDTC (50 mu M) depressed non-tolerant vasorelaxation to GTN (1 nM-1 mu M) per se. 5 Tiron (10 mM) abolished the reduction of ferricytochrome c by a superoxide anion generating system, assessed photometrically in vitro. In contrast, DMSO (0.2% v v(-1)), NAC (1 mM) and PDTC (50 mu M) were without effect. 6 Our data suggests that neither oxidant stress nor nuclear activation, is important in the development of cellular tolerance to GTN in rat isolated aortic smooth muscle.