Evidence for the interaction between nitric oxide and vasoactive intestinal polypeptide in the mouse gastric fundus

PubMedID: 11714881The involvement of nitric oxide (NO) and vasoactive intestinal polypeptide (VIP) in nonadrenergic noncholinergic (NANC) nerve-induced relaxation and the interaction between NO and VIP were investigated in the mouse gastric fundus. N?-nitro-L-arginine (L-NOARG; 100 µM) completely inhibited the NANC relaxations induced by electrical stimulation (ES) (0.5, 1, 2, 4, and 8 Hz; 25 V; 1 ms; 15-s trains). Hemoglobin (20 µM), hydroxocobalamin (100 µM), and 1H-[1,2,4,]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ; 10 µM) diminished ES-induced relaxations, but ?-chymotrypsin (10 U/ml) and VIP antiserum (1/200 dilution) had no effect on NANC relaxations. L-NOARG (100 µM) did not have any effect, whereas ODQ (10 µM) attenuated sodium nitroprusside (SNP; 100 nM)-induced relaxations. ?-Chymotrypsin (10 U/ml) had no effect on the response to SNP. Furthermore, ?-chymotrypsin (10 U/ml) abolished and VIP antiserum (1/200 dilution) diminished VIP (50 nM)-induced relaxations. L-NOARG (100 µM) caused an inhibition of VIP-induced relaxation that was reversed by L-arginine (1 mM) but not by D-arginine (1 mM). Similarly, ODQ (10 µM) inhibited the responses to VIP. 2-Amino-5,6-dihydro-6-methyl-4H-1,3-thiazine (5 µM) had no effect on these relaxations. L-NOARG (100 µM) and ODQ (10 µM) did not affect isoproterenol (10 nM)-induced relaxations. In conclusion, these results provide evidence that NO is involved in NANC nerve-induced relaxation and the participation of VIP (and related neuropeptides) cannot be excluded in causing relaxation of mouse gastric fundus muscle strips. These findings support the idea that VIP directly stimulates the production of NO by increasing NOS activity and thereby activating soluble guanylyl cyclase in smooth muscle

A putative role for S-nitrosoglutathione as the source of nitric oxide in photorelaxation of the mouse gastric fundus

PubMedID: 12208320Mouse gastric fundus strips were relaxed by ultraviolet light (UV) irradiation, exogenous nitric oxide (NO), isoproterenol, S-nitrosoglutathione, S-nitroso-L-cysteine and S-nitroso-N-acetyl-penicillamine. Glutathione did not affect relaxations in response to UV irradiation, exogenous NO and isoproterenol while inhibiting that with S-nitrosoglutathione. L-Cysteine inhibited responses to UV irradiation and exogenous NO, but not in the presence of exogenous Cu2+/Zn2+ superoxide dismutase. However, L-cysteine alone or in combination with Cu2+/Zn2+ superoxide dismutase did not affect the relaxations in response to S-nitroso-L-cysteine. Ethacrynic acid and diamide inhibited photorelaxations but not the relaxations with exogenous NO and isoproterenol. This inhibition was prevented by glutathione, but not by L-cysteine. S-nitrosoglutathione-induced relaxations were abolished by diamide and ethacrynic acid, whereas responses to S-nitroso-L-cysteine and S-nitroso-N-acetyl-penicillamine were only inhibited by ethacrynic acid. These results suggest that S-nitrosoglutathione may, at least in part, be the putative S-nitrosothiol, which is converted to NO in response to UV irradiation in mouse gastric fundus strips. © 2002 Elsevier Science B.V. All rights reserved

Selective modifiers of glutathione prevent restoration of photorelaxations in mouse gastric fundus

PubMedID: 16011739S-nitrosoglutathione (GSNO) has previously been shown to have a role in ultraviolet (UV) light-elicited relaxations and proposed to account for the photosensitive store in the mouse gastric fundus. Furthermore, the depletion of this photosensitive store and its replenishment via long-term electrical field stimulation were demonstrated in the same tissue. In relation to these results, the aim of the present study was to investigate the putative role of S-nitrosothiols in the restorative effect of long-term electrical field stimulation on the reduced photosensitive store. Two series of UV light-elicited relaxations (photorelaxations) were obtained, and the magnitudes of the responses were 53 ± 6 and 26 ± 3%, respectively. The second series of photorelaxations attenuated statistically when compared with those in the first series. Ethacrynic acid (1 µM), diamide (1 µM) and glutathione (1 µM) had no effect on the photorelaxations occurred in the second series of responses. Electrical field stimulation (4 Hz, 25 V, 1 ms, 60 min), applied between two series of photorelaxations, revealed a complete recovery of the attenuated photorelaxations appeared in the second series. NG- monomethyl-L-arginine (100 µM), ethacrynic acid (1 µM) and diamide (1 µM) extensively prevented the restorative effect of electrical field stimulation on photorelaxations. In addition, glutathione (1 µM) reversed the prevention achieved by ethacrynic acid and diamide. The conclusion is that the restoration accomplished by electrical field stimulation is because of the activation of nitric oxide synthase, which in turn brings about the regeneration of GSNO proposed to be the photodegradable material store. © 2005 Blackwell Publishing

Neocuproine inhibits the decomposition of endogenous S-nitrosothiol by ultraviolet irradiation in the mouse gastric fundus

PubMedID: 14757150In the present study, we investigated whether copper ions are involved in the decomposition of endogenous S-nitrosothiols by ultraviolet (UV) light irradiation in the mouse gastric fundus. The effects of copper ions and chelators of copper(I) and copper(II), neocuproine and cuprozine, respectively, were studied on relaxations in response to S-nitrosoglutathione, UV irradiation, exogenous nitric oxide (NO), added as acidified NaNO2, and isoproterenol. UV irradiation of smooth muscle strips induced fast and transient relaxations which were mimicked by exogenous NO. S-Nitrosoglutathione induced concentration-dependent relaxations, which were more sustained than those elicited by UV irradiation or NO. CuCl2 did not affect relaxations elicited by UV irradiation, exogenous NO and isoproterenol but enhanced those elicited by S-nitrosoglutathione. CuSO4 but not FeSO4 mimicked the effect of CuCl2 on relaxations elicited by S-nitrosoglutathione. Neocuproine, the copper(I)-specific chelator, inhibited both photorelaxation and S-nitrosoglutathione-induced relaxation, and this inhibition was prevented by CuCl2. In contrast, neocuproine significantly enhanced the relaxations in response to exogenous NO, without affecting the relaxations elicited by isoproterenol. Cuprizone, a specific copper(II) chelator, did not affect relaxations in response to S-nitrosoglutathione, UV irradiation, exogenous NO and isoproterenol. These results suggest that copper(I) and not copper(II) may play a role in the NO release evoked by the light-induced decomposition of endogenous S-nitrosothiols in mouse gastric fundus. Also, results with the selective copper(I) chelator, neocuproine, confirmed our recent findings that the endogenous "store" of S-nitrosoglutathione, rather than NO, acts as an intermediate in photorelaxation of the mouse gastric fundus, and that photorelaxation may be a suitable model to elucidate the nature of endogenous S-nitrosothiols. © 2003 Elsevier B.V. All rights reserved

Involvement of nitric oxide in non-adrenergic non-cholinergic relaxation and action of vasoactive intestinal polypeptide in circular muscle strips of the rat gastric fundus

PubMedID: 11529689We examined the characteristics of the non-adrenergic non-cholinergic (NANC) nerve induced relaxation and the possible interaction between nitric oxide (NO) and vasoactive intestinal polypeptide (VIP) on the basal tone of the circular muscle of the rat gastric fundus. Electrically induced NANC relaxations were partly inhibited by N?-nitro-L-arginine (100 µM), whereas sodium nitroprusside (SNP; 10 µM) and VIP (5 nM) induced relaxations were not affected. 2-amino-5,6-dihydro-6-methyl-4H-1,3-thiazine (AMT; 5 µM) also inhibited the responses to electrical stimuli to a similar extent as N?-nitro-L-arginine but not VIP. However, AMT plus N?-nitro-L-arginine did not give an additional inhibition above that of each drug alone on NANC relaxations, and dexamethasone (10 µM) had no effect on NANC nerve induced relaxations. 1H-[1,2,4,]oxadiazolo[4,3-a]qulnoxalin-1-one (ODQ; 10 µM), a selective inhibitor of guanylate cyclase, abolished the responses to NANC nerve stimulation and SNP, while VIP responses were not influenced. N-ethylmaleimide (100 µM), an adenylate cyclase inhibitor, attenuated relaxations to NANC nerve stimulation, VIP and isoproterenol (1 nM), while having no effect on those to SNP, but in combination with N?-nitro-L-arginine, there was no additional inhibition on the responses to nerve stimulation. Alpha-chymotrypsin (10 u mi-1) severely diminished VIP induced relaxations, but did not reduce electrical responses. In conclusion, these results suggest that NO is involved in the relaxations induced by short-term electrical stimulation. However, another possible unidentified transmitter that can trigger the accumulation of cyclic GMP is not entirely ruled out and there is no interaction between NO and VIP in the circular muscle strip of the rat gastric fundus, even in the basal state of the tissue. © 2001 Academic Press.University of Delaware Research Foundation SBAG-AYD-288-19S239This work was supported by the Scientific and Technical Research Council of Turkey (TUB'TAK; SBAG-AYD-288-19S239) and C¸ ukurova University Research Foundation (TF.2000.M.26)

The effects of thiol modulators on nitrergic nerve- and S-nitrosothiols-induced relaxation in duodenum

PubMedID: 23412870Background: The aim of this study was to investigate whether thiols are involved in the nitrergic neurotransmission in mouse duodenum. Methods: The effects of thiol-modulating agents, ethacrynic acid (100 µM), a non-specific sulfhydryl alkylator, and diamide (100 µM), an alkylating agent that oxidizes protein sulfhydryl groups and depletes intracellular glutathione, on relaxations to nitrergic stimulation (electrical field stimulation, EFS;10 Hz, 25 V, 1 ms, 15 s-train), S-nitrosoglutathione (GSNO; 5 µM), S-nitrosoacetylpenicillamine (SNAP; 5 µM), and S-nitrosocysteine (CysNO; 10 µM) were investigated. Moreover, the effects of buthionine sulfoximine (100 µM), an inhibitor of ?-glutamylcysteine synthetase, and sulfobromophthalein (100 µM), an inhibitor of glutathione-S-transferase, were studied on relaxant responses to EFS and S-nitrosothiols in mouse duodenum. Results: Exogenous free thiol, glutathione (GSH, 100 µM) did not influence relaxation to EFS, GSNO, SNAP, and CysNO. Ethacrynic acid and diamide significantly decreased relaxation of duodenum to EFS, GSNO, SNAP, and CysNO. This inhibition was prevented by addition of GSH. Buthionine sulfoximine and sulfobromophthalein significantly decreased relaxation to EFS and GSNO but did not influence relaxation to SNAP and CysNO. The inhibitory effect of buthionine sulfoximine and sulfobromophthalein on the relaxant response to EFS and GSNO was prevented by addition of GSH. Conclusions: These results suggest that relaxation to nitrergic stimulation is thiol-dependent, and nitrosothiols, possibly S-nitrosoglutathione may play a role, as an intermediate compound in nitrergic neurotransmission in mouse duodenum

The location of photodegradable nitric oxide store in the mouse stomach fundus

PubMedID: 16978603The aim of this study was to investigate the location of photodegradable nitric oxide (NO) store using a pharmacological approach in mouse gastric fundus. The ultraviolet light irradiation (UV; 360 nm, 60 s), electrical field stimulation (EFS; 4 Hz, 25 V, 1 ms, 15s-train), exogenous nitric oxide (NO; 10 µM), nitroglycerin (100 µM) and isoproterenol (5 nM) induced relaxation in mouse gastric fundus preparations in the absence or presence of an intact mucosa. The NO scavenger, haemoglobin (20 µM), significantly inhibited the relaxation of intact and denuded mucosa stomach fundus to UV light irradiation, EFS and NO, but not to nitroglycerin and isoproterenol. The superoxide anion generator, pyrogallol (50 µM), inhibited relaxation of intact and denuded mucosa stomach fundus induced by UV light irradiation, EFS, NO, but not to nitroglycerin and isoproterenol. The inhibition observed with pyrogallol was prevented by exogenous Cu/Zn superoxide dismutase (SOD; 100 U/ml), a membrane impermeable antioxidant. The Cu/Zn SOD inhibitor, diethyldithiocarbamic acid (DETCA; 8 mM), inhibited the relaxation of intact and denuded mucosa stomach fundus to UV light irradiation, EFS, NO and nitroglycerin but not those to isoproterenol. Exogenous SOD (100 U/ml) partially prevented the inhibitory effect of DETCA on relaxation to UV light irradiation, EFS, NO but not to nitroglycerin. DETCA-induced inhibition of the nitroglycerin-induced relaxation was partially prevented by the cell-permeable polyethylene-glycol-superoxide dismutase (100 U/ml). These results indicate that photodegradable NO store is, at least in part, unlikely to be within smooth muscle cells, and furthermore, that UV light-induced relaxation is not dependent on gastric mucosal layer. © 2006 Elsevier B.V. All rights reserved.TF2002BAP104This work was supported by Cukurova University Research Foundation (TF2002BAP104)

Possible stimulation of Na+-K+-ATPase by NO produced from sodium nitrite by ultraviolet light in mouse gastric fundal strip

PubMedID: 98882691. In the present study, we investigated the roles of Na+-K+-ATPase and extracellular Na+ or Ca2+ ions in ultraviolet (UV) light-induced photorelaxation of methacholine-contracted mouse isolated gastric fundus in the presence of NaNO2 (50 µM). 2. Ouabain (1-500 µM), sodium vanadate (10 µM to 3 mM) and amiloride (1-100 µM) completely inhibited the photorelaxation in a concentration-dependent manner. 3. Metabolic inhibitors, sodium azide (10-100 µM), 2,4-dinitrophenol (100 µM to 1 mM) and sodium fluoride (100 µM to 1 mM) significantly reduced photorelaxation. 4. Substitution of sucrose, lithium or KCl with extracellular Na+ completely abolished the photorelaxant responses. 5. Replacement of all extracellular CaCl2 with BaCl2 also completely inhibited UV-induced relaxation. 6. Verapamil (1-10 µM) decreased UV-induced relaxation significantly. 7. These results suggest that nitric oxide produced from NaNO2 by UV-light in mouse gastric fundus probably stimulates Na+-K+-ATPase activity, and photorelaxation of gastric smooth muscle is dependent on extracellular Na+ and Ca2+ ions. Copyright (C) 1998 Elsevier Science Inc.This study was supported by the Çukurova University Research Foundation (TF. 96 48)

The influence of nitric oxide donors on the responses to nitrergic nerve stimulation in the mouse duodenum

PubMedID: 11399268We investigated whether exogenous nitric oxide (NO) donors have a prejunctional and/or postjunctional inhibitory effect on the nitrergic responses and whether this inhibitory effect was mediated by NO itself and in part, by cyclic GMP in mouse duodenal strips. N?-nitro-L-arginine inhibited relaxations induced by electrical field stimulation of nitrergic nerves, but not those with acidified NaNO2. Furthermore, 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ) inhibited both types of relaxations while 2-amino-5,6-dihydro-6-methyl-4H-1,3-thiazine (AMT) and N-ethylmaleimide were ineffective. NO donors, nitroglycerin and sodium nitroprusside, inhibited relaxations induced by nitrergic nerve stimulation, but not those with acidified NaNO2. Hemoglobin, exogenous Cu2+/Zn2+ superoxide dismutase, diethyldithiocarbamic acid and pyrogallol did not influence the relaxation with nitrergic nerve stimulation. However, hemoglobin, diethyldithiocarbamic acid, pyrogallol and diethyldithiocarbamic acid plus pyrogallol attenuated the inhibitory effect of NO donors on relaxation with nitrergic nerve stimulation, and exogenous superoxide dismutase potentiated this inhibitory effect. Moreover, nitrergic nerve-mediated relaxations were inhibited by 8-bromo-cyclic GMP, but not by 8-bromo-cyclic AMP. These results suggest that exogenous NO donors have a prejunctional inhibitory effect on the nerve-mediated nitrergic relaxation and that the inhibitory effects of nitroglycerin and sodium nitroprusside are NO-dependent, but not related to NO metabolites such as peroxynitrite or a nitrosothiol intermediate. However, a contribution of S-nitrosothiol formed intracellularly cannot be entirely ruled out. Also, this prejunctional inhibition is mediated, at least in part, by the cyclic GMP, but not the cyclic AMP, pathway. © 2001 Elsevier Science B.V.TF.99.6We are indebted to Dr. S. Cellek (The Wolfson Institue for Biomedical Research, University College London) for the gift of 1 H -[1,2,4]oxadiazolo[4,3- a ]quinoxalin-1-one. We thank Mr. Kenan Daglıoglu (Çukurova University Experimental Research Center) for the supply of mice. This work was supported by Çukurova University Research Foundation (TF.99.6) and parts of this work were presented at the annual meeting of the European Pharmacological Societies, Budapest, Hungary, 3–7 July 1999

Protective effect of quercetin, a polyphenolic compound, on mouse corpus cavernosum

PubMedID: 19702691Flavonoids are plant-based phenolic compounds, and quercetin is the most abundant dietary member of this family. One of the most important characteristics of quercetin is its antioxidant property. The aim of this study was to investigate antioxidant effects of quercetin on corpora cavernosa of mice. Corpora cavernosa were isolated in organ baths, precontracted with phenylephrine (0.5 µm) and relaxant responses were mediated by acetylcholine (0.1-1 µm), electrical field stimulation (EFS, 1-16 Hz, 0.5 ms, 30 V) or acidified sodium nitrite (a NaNO2, 0.5 mm). Superoxide anion generators; pyrogallol (50 µm), hydroquinone (100 µm), LY 83583 (6-Anilinoquinolin-5,8-quinone, 10 µm) and superoxide dismutase (SOD) inhibitor; diethyldithiocarbamic acid (DETCA, 8 mm) were used in order to expose corpus cavernosa to oxidant stress. Acetylcholine (0.1-1 µm) induced relaxant responses were significantly inhibited in LY 83583 (10 µm) and DETCA + LY 83583 applicated trials. EFS-induced relaxant responses were significantly inhibited in DETCA (8 mm) and DETCA + LY 83583 administrated trials. On the other hand, acidified sodium nitrite-induced responses were inhibited by all of the superoxide anion generators tested. Quercetin (10 µm) failed to improve the inhibitions on endothelium and electrically stimulated responses. Acidified sodium nitrite (0.5 mm) mediated relaxant responses were significantly restored by quercetin except the groups in which LY 83583 were used. The data suggest that quercetin acts as a protective agent in mouse corpus cavernosum, increasing the bioavailability of exogenous nitric oxide by protecting it from superoxide anion (O2-). © 2009 Société Française de Pharmacologie et de Thérapeutique