Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:4.6.1.2 (guanylate cyclase)
 8,497 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

It has been shown that endothelium-derived relaxing factor (EDRF) may inhibit platelet aggregation in vitro through activation of platelet-soluble guanylate cyclase. To assess whether EDRF may also affect platelet function in vivo, intravascular platelet aggregation was initiated by placing an external constrictor around endothelially injured rabbit carotid arteries. Carotid blood flow velocity was measured continuously by a Doppler flow probe placed proximal to the constrictor. After placement of the constrictor, cyclic flow reductions (CFRs), due to recurrent platelet aggregation, developed at the site of the stenosis. After CFRs were observed for 30 minutes, a solution of authentic nitric oxide (NO, n = 10) was infused into the carotid artery via a small catheter placed proximally to the stenosis. Before infusion of NO, CFR frequency averaged 18.3 +/- 2.9 cycles per hour, and CFR severity (lowest carotid blood flow as percentage of baseline values) was 6 +/- 1%. NO completely inhibited CFRs in all animals, as shown by the normal and constant pattern of carotid blood flow (CFR frequency, 0 cycles per hour, p < 0.001; carotid blood flow, 92 +/- 5%, p = NS versus baseline). These effects were transient; CFRs were restored spontaneously within 10 minutes after cessation of NO infusion. After CFRs returned, S-nitroso-cysteine (S-NO-cys), a proposed form of EDRF, was infused into the carotid artery. S-NO-cys also abolished CFRs in all animals but at a significantly lower dose than NO (0.3 +/- 0.1 versus 12 +/- 4 nmol/min). The role of endogenously released EDRF in modulating in vivo platelet function was then tested in additional experiments. In 10 animals, endogenous release of EDRF was stimulated by infusing acetylcholine into the aortic root during CFRs. Infusion of acetylcholine was also associated with a complete inhibition of CFRs, similar to that observed during exogenous infusion of NO or S-NO-cys. These antithrombotic effects of acetylcholine were completely lost when EDRF synthesis was prevented by administration of the L-arginine analogue NG-monomethyl L-arginine (L-NMMA). Furthermore, in six additional rabbits the basal release of EDRF was blocked by L-NMMA after CFRs had been previously abolished with aspirin or the combination of aspirin and ketanserin, a serotonin S2 receptor antagonist. L-NMMA caused restoration of CFRs in all animals, indicating that even the basal release of EDRF is important in modulating platelet reactivity in vivo. Taken together, the data of the present study demonstrate that endogenous EDRF might importantly contribute to the modulation of platelet function in vivo.
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PMID:Endothelium-derived relaxing factor modulates platelet aggregation in an in vivo model of recurrent platelet activation. 142 38

1. The effect of copper on the activity of the S-nitrosothiol compounds S-nitrosocysteine (cysNO) and S-nitrosoglutathione (GSNO) was investigated, using the specific copper chelator bathocuproine sulphonate (BCS), and human washed platelets as target cells. 2. Chelation of trace copper with BCS (10 microM) in washed platelet suspensions reduced the inhibition of thrombin-induced platelet aggregation by GSNO; however, BCS had no significant effect on the anti-aggregatory action of cysNO. BCS inhibited cyclic GMP generation in response to both cysNO and GSNO. 3. The effect of BCS was rapid (within 30 s), and could be abolished by increasing the platelet concentration to 500 x 10(9) l-1. 4. In BCS-treated platelet suspensions, the addition of Cu2+ ions (0.37-2.37 microM) led to a restoration of both guanylate cyclase activation and platelet aggregation inhibition by GSNO. 5. The anti-aggregatory activity of GSNO was reduced in a concentration-dependent manner by the copper (I)-specific chelators BCS and neocuproine, and to a smaller extent by desferal. No effect was observed with the copper (II) specific chelator, cuprizone, the iron-specific chelator, bathophenanthroline sulphonate, or the broader-specificity copper chelator, D-penicillamine. 6. In both BCS-treated and -untreated platelet suspensions, cys NO was more potent than GSNO as a stimulator of guanylate cyclase. In BCS-treated platelet suspensions there was no significant difference between the anti-aggregatory potency of cysNO and GSNO; however, in untreated suspensions, GSNO was significantly more potent than cysNO. Thus, when copper was available, GSNO produced a greater inhibition of aggregation than cysNO, despite being a less potent activator of guanylate cyclase. 7. The breakdown of cysNO and GSNO was measured spectrophotometrically by decrease in absorbance at 334 nm. In Tyrode buffer, cysNO (10 microM) broke down at a rate of 3.3 microM min-1. BCS (10 microM)reduced this to 0.5 microM min-1. GSNO, however, was stable, showing no fall in absorbance over a period of 7 min even in the absence of BCS.8. We conclude that copper is required for the activity of both cysNO and GSNO, although its influence on anti-aggregatory activity is only evident with GSNO. The stimulatory effect of copper is unlikely to be explained solely by catalysis of S-nitrosothiol breakdown. The enhancement by copper of the anti-aggregatory activity of GSNO, relative to cysNO, suggests that copper may be required for biological activity of GSNO which is independent of guanylate cyclase stimulation.
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PMID:Copper chelation-induced reduction of the biological activity of S-nitrosothiols. 778 Jun 43

The effects of superoxide anion generators, the nitric oxide (NO) scavenger 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoine-1-oxyl 3-oxide (carboxy-PTIO), the specific guanylate cyclase inhibitor 1H-[1,2,4]-oxadiazole-[4,3-a]-quinoxalin-1-one (ODQ), and thiol modulating agents were investigated on relaxations induced by nitrergic stimulation and exogenous NO addition in the sheep urethra. Methylene blue (MB, 10 microM), pyrogallol (0.1 mM) and xanthine (X, 0.1 mM)/xanthine oxidase (XO, 0.1 u ml(-1)) inhibited NO-mediated relaxations, without affecting those induced by nitrergic stimulation. This resistance was not diminished following inhibition of endogenous Cu/Zn superoxide dismutase (Cu/Zn SOD) with diethyldithiocarbamic acid (DETCA, 3 mM), which almost abolished tissue SOD activity. Carboxy-PTIO (0.1 - 0.5 mM) inhibited NO-mediated relaxations but had no effect on responses to nitrergic stimulation, which were not changed by treatment with ascorbate oxidase (2 u ml(-1)). Relaxations to NO were reduced, but not abolished, by ODQ (10 microM), while nitrergic responses were completely blocked. The thiol modulators, ethacrynic acid (0.1 mM), diamide (1.5 mM), or 5,5'-dithio-bis (2-nitrobenzoic acid) (DTNB, 0. 5 mM), and subsequent treatment with dithiothreitol (DTT, 2 mM) had no effect on responses to nitrergic stimulation or NO. In contrast, N-ethylmaleimide (NEM, 0.2 mM) markedly inhibited both relaxations. L-cysteine (L-cys, 0.1 mM) had no effect on responses to NO, while it inhibited those to nitrergic stimulation, in a Cu/Zn SOD-independent manner. Our results do not support the view that the urethral nitrergic transmitter is free NO, and the possibility that another compound is acting as mediator still remains open. British Journal of Pharmacology (2000) 129, 53 - 62
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PMID:Effects of superoxide anion generators and thiol modulators on nitrergic transmission and relaxation to exogenous nitric oxide in the sheep urethra. 1069 2