oxyhyponitrite and 1-3-dihydroxy-4-4-5-5-tetramethyl-2-(4-carboxyphenyl)tetrahydroimidazole

The nitroxyl anion (HNO) is emerging as a novel regulator of cardiovascular function with therapeutic potential in the treatment of diseases such as heart failure. It remains unknown whether tolerance develops to HNO donors, a limitation of currently used nitrovasodilators. The susceptibility of the HNO donor, Angeli's salt (AS), to the development of vascular tolerance was compared with the NO donors, glyceryl trinitrate (GTN) and diethylamine/NONOate (DEA/NO) in rat isolated aortae. Vasorelaxation to AS was attenuated (P<0.01) by the HNO scavenger l-cysteine, whereas the sensitivity to GTN and DEA/NO was decreased (P<0.01) by the NO. scavenger carboxy-[2-(4-carboxyphenyl)-4,4,5,5-tetramethyl-imidozoline-1-oxy-3-oxide]. The soluble guanylate cyclase inhibitor 1H-[1,2,4]oxadiazolo[4,3-a]quinoxaline-1-one impaired responses to GTN>or=AS>>DEA/NO. Pretreatment with 10, 30, and 100 micromol/L of GTN for 60 minutes induced a 4- (P<0.05), 13- (P<0.01), and 48-fold (P<0.01) decrease in sensitivity to GTN, demonstrating tolerance development. In contrast, pretreatment with AS or DEA/NO (10, 30, and 100 micromol/L) did not alter their subsequent vasorelaxation. All of the nitrovasodilators (30 micromol/L) displayed a similar time course of vasorelaxation and cGMP accumulation over a 60-minute period. Unlike vasorelaxation, the magnitude of peak cGMP accumulation differed substantially: DEA/NO>>AS>GTN. GTN did not induce cross-tolerance to either AS or DEA/NO. In contrast, pre-exposure to DEA/NO, but not AS, caused a concentration-dependent attenuation (P<0.01) of GTN-mediated relaxation, which was negated by the protein kinase G inhibitor guanosine 3',5'-cyclic monophosphorothioate, 8-(4-chlorophenylthio)-,Rp-isomer, triethylammonium salt. In conclusion, vascular tolerance does not develop to HNO, nor does cross-tolerance between HNO and GTN occur. Thus, HNO donors may have therapeutic advantages over traditional nitrovasodilators.

Topics: Animals; Aorta, Thoracic; Benzoates; Cyclic GMP-Dependent Protein Kinases; Cysteine; Drug Tolerance; Enzyme Inhibitors; Free Radical Scavengers; Hydrazines; Imidazoles; In Vitro Techniques; Male; Nitric Oxide; Nitric Oxide Donors; Nitrites; Nitrogen Oxides; Nitroglycerin; Oxadiazoles; Quinoxalines; Rats; Rats, Inbred WKY; Time Factors; Vasodilation

Nitric oxide (NO) plays an important role in the control of vascular tone. Traditionally, its vasorelaxant activity has been attributed to the free radical form of NO (NO*), yet the reduced form of NO (NO-) is also produced endogenously and is a potent vasodilator of large conduit arteries. The effects of NO- in the resistance vasculature remain unknown. This study examines the activity of NO- in rat small isolated mesenteric resistance-like arteries and characterizes its mechanism(s) of action. With the use of standard myographic techniques, the vasorelaxant properties of NO* (NO gas solution), NO- (Angeli's salt), and the NO donor sodium nitroprusside were compared. Relaxation responses to Angeli's salt (pEC50=7.51+/-0.13, Rmax=95.5+/-1.5%) were unchanged in the presence of carboxy-PTIO (NO* scavenger) but those to NO* and sodium nitroprusside were inhibited. l-Cysteine (NO- scavenger) decreased the sensitivity to Angeli's salt (P<0.01) and sodium nitroprusside (P<0.01) but not to NO*. The soluble guanylate cyclase inhibitor ODQ (3 and 10 micromol/L) concentration-dependently inhibited relaxation responses to Angeli's salt (41.0+/-6.0% versus control 93.4+/-1.9% at 10 micromol/L). The voltage-dependent K+ channel inhibitor 4-aminopyridine (1 mmol/L) caused a 9-fold (P<0.01) decrease in sensitivity to Angeli's salt, whereas glibenclamide, iberiotoxin, charybdotoxin, and apamin were without effect. In combination, ODQ and 4-aminopyridine abolished the response to Angeli's salt. In conclusion, NO- functions as a potent vasodilator of resistance arteries, mediating its response independently of NO* and through the activation of soluble guanylate cyclase and voltage-dependent K+ channels. NO- donors may represent a novel class of nitrovasodilator relevant for the treatment of cardiovascular disorders such as angina.

Topics: Animals; Arteries; Benzoates; Culture Techniques; Cysteine; Enzyme Activation; Enzyme Inhibitors; Free Radical Scavengers; Guanylate Cyclase; Imidazoles; Male; Nitric Oxide Donors; Nitrites; Nitrogen Oxides; Nitroprusside; Oxadiazoles; Potassium; Potassium Channel Blockers; Potassium Channels, Voltage-Gated; Quinoxalines; Rats; Rats, Inbred WKY; Vascular Resistance; Vasodilation

1. The effects of agents that inactivate free radical nitric oxide (carboxy-PTIO, hydroxocobalamin and pyrogallol) were tested on relaxations produced by the nitroxyl anion (NO(-)) donor Angeli's salt in rat aortic rings and anococcygeus muscles. The amount of NO(*) generated from Angeli's salt in the presence of these agents was measured using a NO(*)-selective electrode sensor. 2. Carboxy-PTIO (100, 300 microM), hydroxocobalamin (30, 100 microM) and pyrogallol (10, 30 microM) significantly reduced relaxations produced by Angeli's salt (0.3 microM) in aortic rings but not in anococcygeus muscles. 3. NO(*) generated from Angeli's salt (0.1 - 10 microM), as detected by the sensor electrode, was less than 0.5% of the amount of Angeli's salt added. Carboxy-PTIO (100 microM) and hydroxocobalamin (30 microM), but not pyrogallol significantly increased the amount of NO(*) detected. 4. In the presence of an oxidizing agent copper [II] (as CuSO(4) 100 microM), the amount of NO(*) detected from 0.3 microM of Angeli's salt increased from an undetectable level of 142.7+/-15.7 nM (equivalent to 47.6% of Angeli's salt added). Under these conditions, carboxy-PTIO, hydroxocobalamin and pyrogallol significantly reduced the amount of NO(*) detected from Angeli's salt as well as the signal generated by an equivalent amount of authentic NO (0.33 microM). 5. The difference in effects of these agents on relaxations to Angeli's salt in the aorta and the anococcygeus muscle may be explained by the ready conversion of NO(-) to NO(*) in the aorta through an unidentified mechanism, which makes NO(-) susceptible to inactivation by these agents. Furthermore, in addition to inactivating NO(*), carboxy-PTIO and hydroxocobalamin may themselves oxidize NO(-) to NO(*), albeit slightly.

Topics: Animals; Antioxidants; Aorta, Thoracic; Benzoates; Free Radicals; Hematinics; Hydroxocobalamin; Imidazoles; In Vitro Techniques; Male; Muscle Relaxation; Muscle, Smooth; Nitric Oxide; Nitrites; Nitrogen Oxides; Pyrogallol; Rats; Rats, Sprague-Dawley