Hemodynamic responses elicited by systemic injections of flavin adenine dinucleotide in anesthetized rats.

Flavin adenine dinucleotide (FAD) elicits an endothelium-dependent vasodilation in isolated rat mesenteric beds via activation of P2Y-purinoceptors. The aims of this study were to characterize the hemodynamic responses elicited by systemic injections of FAD and flavin mononucleotide (FMN) in anesthetized rats and to determine the role of nitric oxide synthase (NOS), cyclooxygenase, P2Y/P2X-purinoceptors, and muscarinic receptor in these responses. FAD (0.05-1.0 micromol/kg, iv) elicited dose-dependent decreases in heart rate (HR), mean arterial blood pressure (MAP), and hindquarter vascular resistance (HQR), whereas it elicited an initial increase and then a decrease in mesenteric (MR) vascular resistance. The FAD-induced responses were not affected by the P2Y/P2X-purinoceptor antagonist suramin, the muscarinic receptor antagonist methyl-atropine, or the cyclooxygenase inhibitor indomethacin. The vasodilator actions of FAD were unaffected by the NOS inhibitor N-nitro-L-arginine methyl ester (L-NAME), whereas the bradycardia elicited by higher doses of FAD were diminished by L-NAME. FMN did not elicit hemodynamic responses in the absence or presence of L-NAME. In summary, FAD-induced bradycardia depends, in part, on the activation of NOS, whereas the vasodilator actions of FAD are not obviously due to newly synthesized nitrosyl factors. These findings and those in our companion manuscript support the concepts that the adenine moiety confers biological activity to FAD, which releases preformed pools of nitrosyl factors.

L-beta,beta-dimethylcysteine attenuates the haemodynamic responses elicited by systemic injections of peroxynitrite in anaesthetized rats.

1 There is direct chemical evidence that L-beta,beta-dimethylcysteine (L-penicillamine (L-PEN)) is a scavenger of peroxynitrite. The aim of this study was to determine whether L-PEN attenuates the haemodynamic responses elicited by peroxynitrite in pentobarbital-anaesthetized rats. 2 Peroxynitrite (1-20 micromol kg(-1), i.v.) elicited dose-dependent reductions in mean arterial blood pressure (MAP) and mesenteric and hindquarter vascular resistances. 3 L-PEN (2 mmol kg(-1), i.v.) elicited relatively minor but significant increases in MAP and vascular resistances. The initial reductions in MAP and vascular resistances elicited by peroxynitrite were not diminished after administration of L-PEN whereas they were much shorter in duration. As such, the total reductions in MAP and vascular resistances were markedly reduced by L-PEN. 4 The finding that L-PEN (2 mmol kg(-1), i.v.) did not affect the hypotensive or vasodilator responses elicited of the ATP-dependent potassium-channel agonist, cromakalim (3-18 microg kg(-1), i.v.), suggests that this dose of L-PEN is not a nonselective inhibitor of vasodilation. 5 These findings suggest that L-PEN may effectively scavenge peroxynitrite in vivo and/or interfere with the mechanisms by which peroxynitrite elicits its vasodilator responses.

Peroxynitrite elicits dysfunction of stereoselective s-nitrosocysteine recognition sites.

The aim of this study was to determine whether induction of tachyphylaxis to peroxynitrite (induced by giving 10 intravenous injections of a 10-micromol/kg dose) differentially affects the vasodilator responses elicited by systemic injections of the L- and D-isomers of S-nitrosocysteine (L-SNC and D-SNC), in pentobarbital-anesthetized rats. L- and D-SNC (12.5-200 nmol/kg, iv) elicited dose-dependent reductions in hindquarter, mesenteric, and renal vascular resistances. The L-SNC-induced vasodilator responses in the hindquarter and renal vascular beds were virtually abolished whereas the vasodilator responses in mesenteric bed were markedly diminished after administration of peroxynitrite. The D-SNC-induced vasodilator responses in the hindquarter and renal beds were slightly attenuated whereas the vasodilator responses in the mesenteric bed were not diminished after administration of peroxynitrite. The vasodilator responses elicited by the nitric oxide donor, MAHMA NONOate (5-50 nmol/kg, iv), were not attenuated by peroxynitrite. The finding that induction of tachyphylaxis to peroxynitrite diminishes the effects of L- and D-SNC but not MAHMA NONOate suggests that the stereoisomers exert their vasodilator effects by mechanisms other than their decomposition to nitric oxide. Moreover, the finding that induction of tachyphylaxis to peroxynitrite causes a more pronounced attenuation of the vasodilator effects of L- than D-SNC supports evidence that the stereoisomers differentially interact with stereoselective S-nitrosothiol recognition sites in the vasculature. Taken together, these novel results support the possibility that peroxynitrite diminishes the vasodilator potencies of L- and D-SNC by oxidation and/or nitration of amino acids in these recognition sites.

Loss of K+ATP-channel-mediated vasodilation after induction of tachyphylaxis to peroxynitrite.

Systemic injections of peroxynitrite elicit pronounced vasodilator responses in rats by activation of ATP-dependent K+ channels (K+ATP-channels). The aim of this study was to determine whether development of tachyphylaxis to the vasodilator actions of peroxynitrite involves the loss of K+ATP-channel function. The falls in mean arterial blood pressure (MAP) and mesenteric and hindquarter vascular resistances produced by the K+ATP-channel agonist, cromakalim (3-18 microg/kg, iv), and the nitric oxide (NO) donor, sodium nitroprusside (SNP; 1-4 microg/kg, iv), were determined in pentobarbital-anesthetized rats before and after induction of tachyphylaxis to peroxynitrite induced by the administration of 10 injections of peroxynitrite (10 micromol/kg, iv). The first dose of peroxynitrite elicited pronounced falls in MAP and vascular resistances whereas the tenth injection elicited much smaller responses that were equivalent to those of decomposed peroxynitrite. Before induction of tachyphylaxis to peroxynitrite, cromakalim and SNP produced dose-dependent reductions in MAP and vascular resistances. The hemodynamic actions of cromakalim were markedly attenuated after induction of tachyphylaxis to peroxynitrite whereas the SNP-induced responses were only slightly attenuated. These results suggest that tachyphylaxis to the vasodilator actions of peroxynitrite involves the loss of K+ATP-channel function whereas tachyphylaxis to peroxynitrite minimally affects NO-mediated vasodilation. Taken together, these findings raise the possibility that peroxynitrite inhibits K+ATP-channel function by oxidation and/or nitration of amino acids in these channels.

Role of ATP-sensitive K+ -channels in hemodynamic effects of peroxynitrite in anesthetized rats.

The aim of this study was to determine whether the hypotensive and vasodilator actions of peroxynitrite in pentobarbital-anesthetized rats involve the activation of ATP-sensitive K+-channels (K+ATP-channels). The effects of the K+ATP-channel agonist, cromakalim (9-36 microg/kg, iv), peroxynitrite (0.5-10 micromol/kg iv), and L-S-nitrosocysteine (12.5-200 nmol/kg, iv) on mean arterial blood pressure (MAP) and mesenteric (MR) and hindquarter (HQR) vascular resistances were determined before and after injection of the K+ATP-channel blocker, glibenclamide (40 micromol/kg, iv). Cromakalim, peroxynitrite, and L-S-nitrosocysteine produced dose-dependent reductions in MAP, MR, and HQR. Administration of glibenclamide did not affect resting hemodynamic parameters but markedly attenuated the hemodynamic actions of cromakalim. The maximal falls in MAP and HQR produced by peroxynitrite were attenuated by glibenclamide whereas the maximal falls in MR were not affected. In addition, the duration of the hypotensive and vasodilator effects of peroxynitrite in the mesenteric and hindquarter beds were markedly diminished by glibenclamide. In contrast, glibenclamide did not affect the maximal hypotensive or vasodilator effects of L-S-nitrosocysteine or the duration of these responses. These results suggest that the hypotensive and vasodilator actions of peroxynitrite in anesthetized rats involve the activation of K+ATP-channels whereas the hemodynamic actions of L-S-nitrosocysteine do not.

Vasodilator actions of the endothelium-derived relaxing factor L-S-nitrosocysteine in anaesthetized rats are markedly diminished by peroxynitrite.

The aim of the present study was to assess the effects of the potent oxidant and nitrating agent peroxynitrite on the haemodynamic actions of the endothelium-derived S-nitrosothiol L-S-nitrosocysteine. The haemodynamic actions of L-S-nitrosocysteine (12.5-100 nmol/kg, i.v.) were determined in pentobarbital-anaesthetized rats before and after the induction of tachyphylaxis to peroxynitrite achieved by giving 10 intravenous injections of a 10 micromol/kg dose. L-S-Nitrosocysteine elicited dose-dependent reductions in mean arterial blood pressure and in hindquarter and mesenteric vascular resistance. The L-S-nitrosocysteine-induced responses were substantially attenuated after administration of peroxynitrite. We have reported previously that nitric oxide-mediated vasodilation is not diminished after the induction of tachyphylaxis to peroxynitrite. Taken together, these findings support the concept that peroxynitrite reduces the vasodilator actions of L-S-nitrosocysteine via oxidation and/or nitration of putative membrane-bound S-nitrosothiol recognition sites.