cyclic-gmp and diethylamine

The effects of the Pseudomonas aeruginosa virulence factor pyocyanin (PCN) on the contractile function of porcine coronary arteries was investigated in vitro. Artery rings (5 mm) were suspended in organ baths containing Krebs' solution for the measurement of isometric tension. The effect of PCN on resting and precontracted coronary arteries was initially investigated with various agents. Arteries were precontracted with prostaglandin (PG) F2α or potassium chloride and endothelium-dependent relaxations were induced by various agents in the presence of PCN. Pyocyanin (0.1-10 μmol/L) evoked small-amplitude, dose-dependent contractions in resting porcine coronary arteries. In addition, PCN amplified the contractile response to PGF2α , but did not alter responses to carbachol. Pyocyanin (0.1-10 μmol/L) significantly inhibited endothelium-dependent relaxations evoked by neurokinin A. Pyocyanin also inhibited relaxations evoked by diethylamine nitric oxide (a nitric oxide donor), forskolin (an adenylate cyclase activator), dibuytyryl-cAMP (a cAMP analogue), 8-bromo-cGMP (a cGMP analogue) and P1075 (a KATP channel activator), but not isoprenaline (β-adrenoceceptor agonist). These results indicate that physiological concentrations of PCN interfere with multiple intracellular processes involved in vascular smooth muscle relaxation, in particular pathways downstream of nitric oxide release. Thus, PCN may alter normal vascular function in patients infected with P. aeruginosa.

Topics: 8-Bromo Cyclic Adenosine Monophosphate; Animals; Colforsin; Coronary Vessels; Cyclic AMP; Cyclic GMP; Diethylamines; Dinoprost; Female; Isoproterenol; Male; Muscle Contraction; Muscle Relaxation; Nitric Oxide; Pseudomonas aeruginosa; Pyocyanine; Swine; Vasodilation

Nitroxyl (HNO) displays distinct pharmacology to its redox congener nitric oxide (NO(•)) with therapeutic potential in the treatment of heart failure. It remains unknown if HNO donors are resistant to tolerance development following chronic in vivo administration. Wistar-Kyoto rats received a 3-day subcutaneous infusion of one of the NO(•) donors, glyceryl trinitrate (GTN) or diethylamine/NONOate (DEA/NO), or the HNO donor Angeli's salt (AS). GTN infusion (10 μg/kg/min) resulted in significantly blunted depressor responses to intravenous bolus doses of GTN, demonstrating tolerance development. By contrast, infusion with AS (20 μg/kg/min) or DEA/NO (2 μg/kg/min) did not alter their subsequent depressor responses. Similarly, ex vivo vasorelaxation responses in isolated aortae revealed that GTN infusion elicited a significant 6-fold decrease in the sensitivity to GTN and reduction in the maximum response to acetylcholine (ACh). Chronic infusion of AS or DEA/NO had no effect on subsequent vasorelaxation responses to themselves or to ACh. No functional cross-tolerance between nitrovasodilators was evident, either in vivo or ex vivo, although an impaired ability of a nitrovasodilator to increase tissue cGMP content was not necessarily indicative of a reduced functional response. In conclusion, HNO donors may represent novel therapies for cardiovascular disease with therapeutic potential over clinically used organic nitrates.

Topics: Acetylcholine; Animals; Aorta; Cyclic GMP; Diethylamines; In Vitro Techniques; Male; Nitric Oxide Donors; Nitrites; Nitrogen Oxides; Nitroglycerin; Rats

Critical roles for nitric oxide (NO) in regulating cell and tissue physiology are broadly appreciated, but aspects remain to be explored. In the mollusk Pleurobranchaea, NO synthase activity is high in CNS ganglia containing motor networks for feeding and locomotion, where a cAMP-gated cation current (I(Na,cAMP)) is also prominent in many neurons. We examined effects of NO on I(Na,cAMP) using voltage-clamp methods developed to analyze cAMP signaling in the live neuron, focusing on the identified metacerebral giant neuron of the feeding network. NO donors enhanced the I(Na,cAMP) response to injected cAMP by an averaged 85%. In dose-response measures, NO increased the current stimulated by cAMP injection without altering either apparent cAMP binding affinity or cooperativity of current activation. NO did not detectably alter levels of native cAMP or synthesis or degradation rates as observable in both current saturation and decay rate of I(Na,cAMP) responses to cAMP injection. NO actions were not exerted by cGMP signaling, as they were not mimicked by cGMP analogue nor blocked by inhibitors of guanylate cyclase and protein kinase G. NO potentiation of I(Na,cAMP) was broadly distributed among many other neurons of the feeding motor network in the buccal ganglion. However, NO did not affect a second type of I(Na,cAMP) found in locomotor neurons of the pedal ganglia. These results suggest that NO acts through a novel mechanism to regulate the gain of cAMP-dependent neuromodulatory pathways that activate I(Na,cAMP) and may thereby affect the set points of feeding network excitability and reactivity to exogenous input.

Topics: Animals; Carbazoles; Cyclic GMP; Diethylamines; Dose-Response Relationship, Radiation; Drug Interactions; Electric Stimulation; Enzyme Inhibitors; Feeding Behavior; Ganglia, Invertebrate; In Vitro Techniques; Indoles; Ion Channel Gating; Membrane Potentials; Neurons; Nitric Oxide; Oxadiazoles; Patch-Clamp Techniques; Pleurobranchaea; Signal Transduction; Thionucleotides; Time Factors

Hypercholesterolemia (HC) induces alterations in systemic vascular reactivity, which can manifest as an attenuated endothelium-dependent relaxation, partly consequent to an impairment in nitric oxide (NO) activity. To determine whether experimental HC has a similar effect on renal vascular function, renal artery segments obtained from pigs fed a HC (n=5) or normal (n=5) diet were studied in vitro. Endothelium-dependent relaxation was examined using increasing concentrations of acetylcholine (Ach), calcium ionophore A23187, and Ach following pre-incubation with N(G)-monomethyl-L-arginine or L-arginine (L-ARG). The NO-donor diethylamine (DEA) was used to examine smooth muscle relaxation response and cyclic GMP generation in endothelium-denuded vessels. The expression of endothelial NO synthase (eNOS) in the renal arteries was examined using Western blotting. Endothelium-dependent relaxation to Ach was significantly attenuated in the HC group compared to normal (53.3+/-9.1 vs. 98.8+/-3.7%, P<0.005), but normalized after pre-incubation with L-ARG (82.3+/-13.8%, P=0.21). Receptor-independent endothelium-dependent relaxation to A23187 was also significantly blunted in HC (75.2+/-10.5 vs. 115.5+/-4.2%, P<0. 017). Smooth muscle relaxation and cyclic GMP generation in response to DEA were greater in denuded HC vessels, while relaxation of intact vessels to nitroprusside was unaltered. In the HC vessels eNOS was almost undetectable. In conclusion, experimental HC attenuates in vitro endothelium-dependent relaxation of the porcine renal artery, possibly due to low bioavailability of NO. These vascular alterations in HC could play a role in the pathogenesis of renal disease or hypertension, supporting a role for HC as a risk factor for renovascular disease.

Topics: Animals; Arginine; Cholesterol; Cyclic GMP; Diethylamines; Endothelium, Vascular; Hemodynamics; Hypercholesterolemia; In Vitro Techniques; Nitric Oxide; Nitric Oxide Donors; Reference Values; Renal Circulation; Swine; Vasomotor System

Nitric oxide (NO) has concentration-dependent biphasic myocardial contractile effects. We tested the hypothesis, in isolated rat hearts, that NO cardiostimulation is primarily non-cGMP dependent. Infusion of 3-morpholinosydnonimine (SIN-1, 10(-5) M), which may participate in S-nitrosylation (S-NO) via peroxynitrite formation, increased the rate of left ventricular pressure rise (+dP/dt; 19 +/- 4%, P < 0.001, n = 11) without increasing effluent cGMP or cAMP. Superoxide dismutase (SOD; 150 U/ml) blocked SIN-1 cardiostimulation and led to cGMP elaboration. Sodium nitroprusside (10(-10)-10(-7) M), an iron nitrosyl compound, did not augment +dP/dt but increased cGMP approximately eightfold (P < 0.001), whereas diethylamine/NO (DEA/NO; 10(-7) M), a spontaneous NO. donor, increased +dP/dt (5 +/- 2%, P < 0.05, n = 6) without augmenting cGMP. SIN-1 and DEA/NO +dP/dt increase persisted despite guanylyl cyclase inhibition with 1H-(1,2,4)oxadiazolo-(4,3,-a)quinoxalin-1-one (10(-5) M, P < 0.05 for both donors), suggesting a cGMP-independent mechanism. Glutathione (5 x 10(-4) M, n = 15) prevented SIN-1 cardiostimulation, suggesting S-NO formation. SIN-1 also produced SOD-inhibitable cardiostimulation in vivo in mice. Thus peroxynitrite and NO donors can stimulate myocardial contractility independently of guanylyl cyclase activation, suggesting a role for S-NO reactions in NO/peroxynitrite-positive inotropic effects in intact hearts.

Topics: Animals; Cyclic GMP; Diethylamines; Drug Combinations; Enzyme Inhibitors; Glutathione; In Vitro Techniques; Male; Molsidomine; Myocardial Contraction; Nitrates; Nitric Oxide; Nitric Oxide Donors; Nitroprusside; Nucleotides, Cyclic; Oxadiazoles; Oxidation-Reduction; Quinoxalines; Rats; Rats, Wistar; Superoxide Dismutase

To characterise the inotropic response of isolated myocytes to a range of structurally unrelated NO donors and to assess the role of NO release kinetics, NO species and cyclic nucleotides in mediating the observed changes.. Guinea-pig (GP) and human myocytes were prepared by enzymatic digestion. Paced contractile amplitude was recorded at 37 degrees C. NO release was measured by reduction of oxyhaemoglobin and using an NO electrode. Cyclic nucleotides were measured using a tritium labelled competitive binding assay.. The NO donors S-nitrosoglutathione (GSNO) and diethylamine/NO (DEA/NO) produced positive inotropic effects in GP myocytes at (10(-5) M) (25 and 111% increases of contraction amplitude). The response to GSNO was significantly enhanced in the presence of a low concentration of isoprenaline (3x10(-10) M). Positive inotropy was observed with a range of both thiol and non-thiol donors, amongst which a fast rate of NO release was associated with positive inotropy. The response to GSNO was abolished by the free NO scavenger oxyhaemoglobin, but not by ODQ (soluble guanylyl cyclase [sGC] inhibitor), Rp-cAMPS (protein kinase A inhibitor) or thapsigargin (sarcoplasmic reticulum Ca(2+) uptake blocker). Direct measurement of cyclic nucleotides showed a rise in cGMP but not cAMP. Human ventricular myocytes showed a significant increase of contraction with GSNO (48+/-15.8%, n=7, P<0. 05) in the presence of isoprenaline and a marked response to DEA/NO alone.. Isolated GP and human myocytes show a positive inotropic effect with certain NO donors. This is independent of sGC and cAMP. The rate of NO release from donors appears important in mediating the effect.

Topics: Animals; Cardiotonic Agents; Cells, Cultured; Cyclic AMP; Cyclic GMP; Diethylamines; Dose-Response Relationship, Drug; Glutathione; Guinea Pigs; Humans; Isoproterenol; Male; Myocardial Contraction; Myocardium; Nitric Oxide; Nitric Oxide Donors; Nitroso Compounds; Penicillamine; S-Nitrosoglutathione; Stimulation, Chemical

The present study was undertaken to assess the effects of sodium nitroprusside (SNP) and diethylamine NO (C2H5)2N[N(O)NO]-Na+ (DEA/NO), NO donors, on an acetylcholine (ACh)-induced Cl- current in identified Onchidium neurons using voltage-clamp and pressure ejection techniques. Bath-applied SNP (10 microM) and DEA/NO (5-10 microM) reduced the ACh-induced Cl- current in the neurons without affecting the resting membrane conductance and holding current. The suppressing effect of NO donors were concentration-dependent and completely reversible. Pretreatment with 1H-[1,2,4]oxadiazolo-[4,3-a] quinoxalin-1-one (1 micro M), a specific inhibitor of NO-stimulated guanylate cyclase, and hemoglobin (50 micro M), a nitric oxide scavenger, decreased the SNP-induced inhibition of the ACh-induced current. Intracellular injection of guanosine 3',5'-cyclic monophosphate (cGMP) or bath-application of 3-isobutyl-1-methylxanthine (50 micro M), a non-specific phosphodiesterase inhibitor, inhibited the ACh-induced current, mimicking the effect of NO donors. These results suggest that SNP and DEA/NO inhibit the ACh-induced Cl- current and that this effect is mediated by an increase in intracellular cGMP.

Topics: 1-Methyl-3-isobutylxanthine; Acetylcholine; Animals; Chlorides; Cyclic GMP; Diethylamines; Electric Conductivity; Enzyme Inhibitors; Hemoglobins; Mollusca; Neurons; Nitric Oxide; Nitroprusside; Oxadiazoles; Quinoxalines

Abstract NONOates are a new class of NO donors that have proven useful for studying the effects of spontaneous and chemically predictable NO release in biologic systems. In order to assess their potential as vasodilatatory drugs in the cerebrovascular bed we have compared the relaxant effects of the classical nitrovasodilator sodium nitroprusside (SNP) and three NONOates, diethylamine/NO complex (DEA/NO), spermine/NO complex (SPER/NO), and diethylenetriamine/NO complex (DETA/NO) in isolated rabbit basilar arteries precontracted with UTP. The 4 NO donors induced full relaxation of the UTP-induced tone, with the following order of potency: SNP > DEA/NO > SPER/NO > DETA/NO. Relaxations induced by SNP and DETA/NO were not modified in rubbed (endothelium denuded) arteries in which acetylcholine-relaxations were almost abolished. On the other hand, relaxations to SNP and SPER/NO were more potent and effective in histamine-precontracted arteries than in KCl-precontracted arteries. Methylene blue significantly inhibited SPER/NO-induced relaxations in both KCl- and histamine-precontracted arteries while SNP-induced relaxations were only slightly inhibited by methylene blue in KCl-precontracted arteries. This study shows that the NO donors SNP, DEA/NO, SPER/NO and DETA/NO have quantitatively different relaxant effects in rabbit basilar arteries according to their rate of NO release. Relaxations are not mediated by endothelial factors, and are inhibited by arterial depolarization. Finally, cGMP formation is involved in relaxation induced by NONOates and much less in SNP-induced relaxation.

Topics: Acetylcholine; Animals; Basilar Artery; Cyclic GMP; Diethylamines; Endothelium, Vascular; Histamine; Male; Methylene Blue; Nitric Oxide; Nitroprusside; Polyamines; Rabbits; Spermine; Structure-Activity Relationship; Uridine Triphosphate; Vasoconstriction; Vasodilation; Vasodilator Agents

The aims of the study were to investigate whether a nitric oxide-cyclic guanosine monophosphate relaxation pathway is present in the human uterus and whether it differentially inhibits contractility during pregnancy and labor.. Myometrial strips were obtained from pregnant women who were either in labor or not in labor and from nonpregnant women. Nitrites and cyclic guanosine monophosphate production by the tissues and contractile responses to nitric oxide modifiers were measured.. Biochemical assays revealed that nitric oxide (nitrites) and cyclic guanosine monophosphate are generated by the human uterus. Cyclic guanosine monophosphate production by the uterus was increased by L-arginine (the substrate for nitric oxide) and diethylamine/nitric oxide (a nitric oxide donor) and decreased by nitro-L-arginine methyl ester (an inhibitor of nitric oxide synthase). Spontaneous contractility in vitro was increased by nitro-L-arginine methyl ester and decreased by diethylamine/nitric oxide, which furthermore produced a dose-dependent inhibition of contractility, and the median effective dose of inhibition in tissues from nonlaboring pregnant patients (1.5 +/- 0.4 mumol/L) is substantially lower than in tissues from laboring pregnant (21.7 +/- 7.4 mumol/L or nonpregnant (20.8 +/- 4.4 mumol/L) women. These studies show that the nitric oxide-cyclic guanosine monophosphate system exists in the human uterus and that it inhibits contractility. Furthermore, the relaxation responsiveness to nitric oxide is elevated during pregnancy and decreased during labor.. A nitric oxide-cyclic guanosine monophosphate relaxation pathway is present in the human uterus and may be responsible for maintaining uterine quiescence during pregnancy. A decrease in uterine relaxation responsiveness to nitric oxide at term may play a role in the initiation of labor.

Topics: Adult; Analysis of Variance; Arginine; Cyclic GMP; Diethylamines; Female; Humans; In Vitro Techniques; Labor, Obstetric; Myometrium; NG-Nitroarginine Methyl Ester; Nitric Oxide; Pregnancy; Uterine Contraction