Effect of nitric oxide donorsS-nitroso-N-acetyl-dl-penicillamine, spermine NONOate and propylamine propylamine NONOate on intracellular pH in cardiomyocytes

Large (pathological) amounts of nitric oxide (NO) induce cell injury, whereas low (physiological) NO concentrations often ameliorate cell injury. We tested the hypotheses that pretreatment of endothelial cells with low concentrations of NO (preconditioning) would prevent injury induced by high NO concentrations. Apoptosis, induced in bovine aortic endothelial cells (BAECs) by exposing them to either 4 mM sodium nitroprusside (SNP) or 0.5 mM N-(2-aminoethyl)- N-(2-hydroxy-2-nitrosohydrazino)-1,2-ethylenediamine (spermine NONOate) for 8 h, was abolished by 24-h pretreatment with either 100 μM SNP, 10 μM spermine NONOate, or 100 μM 8-bromo-cGMP (8-Br-cGMP). Repair of BAECs following wounding, measured as the recovery rate of transendothelial electrical resistance, was delayed by 8-h exposure to 4 mM SNP, and this delay was significantly attenuated by 24-h pretreatment with 100 μM SNP. NO preconditioning produced increased association and expression of soluble guanyl cyclase (sGC) and heat shock protein 90 (HSP90). The protective effect of NO preconditioning, but not the injurious effect of 4 mM SNP, was abolished by either a sGC activity inhibitor 1H-[1,2,4]oxadiazolo-[4,3- a]quinoxalin-1-one (ODQ) or a HSP90 binding inhibitor (radicicol) and was mimicked by 8-Br-cGMP. We conclude that preconditioning with a low dose of NO donor accelerates repair and maintains endothelial integrity via a mechanism that includes the HSP90/sGC pathway. HSP90/sGC may thus play a role in the protective effects of NO-generating drugs from injurious stimuli.

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Platelet Ca2+ responses coupled to glycoprotein VI and Toll-like receptors persist in the presence of endothelial-derived inhibitors: roles for secondary activation of P2X1 receptors and release from intracellular Ca2+ stores

Blood ◽

10.1182/blood-2011-10-386052 ◽

2012 ◽

Vol 119 (15) ◽

pp. 3613-3621 ◽

Cited By ~ 23

Author(s):

C. Y. Eleanor Fung ◽

Sarah Jones ◽

Adwoa Ntrakwah ◽

Khalid M. Naseem ◽

Richard W. Farndale ◽

...

Keyword(s):

Nitric Oxide ◽

Phospholipase C ◽

Platelet Activation ◽

Secretory Pathway ◽

Atp Release ◽

Activation Mechanism ◽

Glycoprotein Vi ◽

Kinase C ◽

Spermine Nonoate ◽

Secondary Activation

Abstract Inhibition of Ca2+ mobilization by cyclic nucleotides is central to the mechanism whereby endothelial-derived prostacyclin and nitric oxide limit platelet activation in the intact circulation. However, we show that ∼ 50% of the Ca2+ response after stimulation of glycoprotein VI (GPVI) by collagen, or of Toll-like 2/1 receptors by Pam3Cys-Ser-(Lys)4 (Pam3CSK4), is resistant to prostacyclin. At low agonist concentrations, the prostacyclin-resistant Ca2+ response was predominantly because of P2X1 receptors activated by ATP release via a phospholipase-C–coupled secretory pathway requiring both protein kinase C and cytosolic Ca2+ elevation. At higher agonist concentrations, an additional pathway was observed because of intracellular Ca2+ release that also depended on activation of phospholipase C and, for TLR 2/1, PI3-kinase. Secondary activation of P2X1-dependent Ca2+ influx also persisted in the presence of nitric oxide, delivered from spermine NONOate, or increased ectonucleotidase levels (apyrase). Surprisingly, apyrase was more effective than prostacyclin and NO at limiting secondary P2X1 activation. Dilution of platelets reduced the average extracellular ATP level without affecting the percentage contribution of P2X1 receptors to collagen-evoked Ca2+ responses, indicating a highly efficient activation mechanism by local ATP. In conclusion, platelets possess inhibitor-resistant Ca2+ mobilization pathways, including P2X1 receptors, that may be particularly important during early thrombotic or immune-dependent platelet activation.

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Abstract P607: Nitric Oxide (NO) Regulates Paracellular Permselectivity in Isolated, Perfused Rat Thick Ascending Limbs through Claudin-19

Hypertension ◽

10.1161/hyp.68.suppl_1.p607 ◽

2016 ◽

Vol 68 (suppl_1) ◽

Author(s):

Casandra M Monzon ◽

Jeffrey Garvin

Keyword(s):

Nitric Oxide ◽

Control Period ◽

Sprague Dawley ◽

Permeability Ratio ◽

Ionic Selectivity ◽

No Donor ◽

Sprague Dawley Rats ◽

Spermine Nonoate ◽

Dilution Potential ◽

Cl Permeability

About 50% of the Na reabsorbed in thick ascending limbs (TALs) traverses the paracellular pathway. The ionic selectivity of this route is regulated by claudins in the tight junctions. TALs express claudin-19 which has been reported to regulate TAL Na permeability. We showed that nitric oxide (NO) decreases Na/Cl permeability ratio (PNa/PCl) in TALs by increasing the absolute permeabilities of both ions though PCl increased more. However, whether NO affects paracellular permeability via claudin-19 is unknown. We hypothesize that NO regulates the paracellular permselectivity in TALs through this claudin. To test this we perfused TALs from Sprague Dawley rats and measured dilution potentials (a measure of permselectivity) with and without exogenously-added or endogenously-produced NO in the presence or absence of an antibody against an extracellular domain of claudin-19 or Tamm-Horsfall protein (control). Dilution potentials were generated by reducing bath NaCl from 141 to 32 mM. For the NO donor spermine NONOate (SPM): during the control period, the dilution potential was -9.3 ± 1.8 mV. After SPM (200 μM), it was -6.7 ± 1.6 mV (n = 6; p < 0.003). In the presence of the claudin-19 antibody, SPM had no significant effect on dilution potentials (claudin-19 antibody alone: -12.7 ± 2.1 mV vs claudin-19 antibody + SPM: -12.9 ± 2.4 mV; n = 6). The claudin-19 antibody alone had no effect on dilution potentials. In the presence of the Tamm-Horsfall protein, the effect of SPM was still present (Tamm-Horsfall protein antibody alone: -9.7 ± 1.0 mV; Tamm-Horsfall protein antibody + SPM: -6.3 ± 1.1 mV, p<0.006, n = 6). For experiments with endogenously-produced NO, L-arginine the substrate for NO synthase was added. During the control period, the dilution potential was -11.0 ± 1.1 mV. After L-arginine (500 μM) treatment, they were -9.0 ± 1.2 mV (n = 9; p < 0.05). In the presence of the claudin-19 antibody, L-arginine had no significant effect on dilution potentials (claudin-19 antibody alone: -10.1 ± 0.9 mV vs claudin-19 antibody + L-arginine: -10.1 ± 1.0 mV; n = 9). In the presence of the Tamm-Horsfall protein, the effect of L-arginine was still present. We conclude that the actions of NO on the paracellular permselectivity in thick ascending limbs are at least in part mediated by claudin-19.

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Nitric Oxide Potentiates cAMP-Gated Cation Current by Intracellular Acidification in Feeding Neurons of Pleurobranchaea

Journal of Neurophysiology ◽

10.1152/jn.00021.2010 ◽

2010 ◽

Vol 104 (2) ◽

pp. 742-745 ◽

Cited By ~ 5

Author(s):

Kurt Potgieter ◽

Nathan G. Hatcher ◽

Rhanor Gillette ◽

Catherine R. McCrohan

Keyword(s):

Nitric Oxide ◽

Signaling Pathways ◽

Intracellular Ph ◽

Camp Signaling ◽

Intracellular Acidification ◽

Ph Indicator ◽

Intracellular Injection ◽

Cation Current ◽

Motor Network ◽

Important Regulator

A pH-sensitive cAMP-gated cation current ( INa,cAMP) is widely distributed in neurons of the feeding motor networks of gastropods. In the sea slug Pleurobranchaea this current is potentiated by nitric oxide (NO), which itself is produced by many feeding neurons. The action of NO is not dependent on either cGMP or cAMP signaling pathways. However, we found that NO potentiation of INa,cAMP in the serotonergic metacerebral cells could be blocked by intracellular injection of MOPS buffer (pH 7.2). In neurons injected with the pH indicator BCECF, NO induced rapid intracellular acidification to several tenths of a pH unit. Intracellular pH has not previously been identified as a specific target of NO, but in this system NO modulation of INa,cAMP via pHi may be an important regulator of the excitability of the feeding motor network.

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Nitric oxide attenuates endothelin-1-induced vasoconstriction in canine stomach

AJP Gastrointestinal and Liver Physiology ◽

10.1152/ajpgi.1996.271.1.g27 ◽

1996 ◽

Vol 271 (1) ◽

pp. G27-G35

Author(s):

J. G. Wood ◽

Q. Zhang ◽

Z. Y. Yan ◽

L. Y. Cheung

Keyword(s):

Nitric Oxide ◽

Methyl Ester ◽

Ischemia Reperfusion ◽

Nitric Oxide Donor ◽

Question Mark ◽

Endothelin 1 ◽

Vasoconstrictor Response ◽

Anesthetized Dogs ◽

Spermine Nonoate

We previously observed that endothelin-1 (ET-1)-induced gastric vasoconstriction is enhanced after ischemia-reperfusion. The purpose of our present study was to examine the role of nitric oxide in regulating ET-1-induced vasoconstriction under normal conditions and after ischemia-reperfusion. Using a mechanically perfused stomach segment from chloralose-anesthetized dogs, we examined 1) responses to NG-nitro-L-arginine methyl ester (L-NAME) alone and in combination with L-arginine, 2) whether L-NAME affects ET-1-induced vasoconstriction under normal conditions and after ischemia-reperfusion, and 3) if spermine NONOate inverted question mark1,3-propanediamine-N-[4-1-(3-aminopropyl)-2-hydroxy-2-nitrosohydrazi no] butyl; a nitric oxide donor inverted question mark attenuates the augmented response to ET-1 after ischemia-reperfusion. Our results show that 1) L-NAME significantly increased baseline vascular resistance and this response was reduced by L-arginine, 2) ET-1-induced vasoconstriction was enhanced by L-NAME, and 3) administration of spermine NONOate during reperfusion largely attenuated the vasoconstrictor response to ET-1 after ischemia-reperfusion. Our findings are consistent with the hypothesis that nitric oxide modulates responses to ET-1 under normal conditions, and loss of this vasodilator after ischemia-reperfusion results in an augmented response to ET-1.

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Nitric oxide inhibits sodium/hydrogen exchange activity in the thick ascending limb

AJP Renal Physiology ◽

10.1152/ajprenal.1999.277.3.f377 ◽

1999 ◽

Vol 277 (3) ◽

pp. F377-F382 ◽

Cited By ~ 22

Author(s):

Jeffrey L. Garvin ◽

Nancy J. Hong

Keyword(s):

Nitric Oxide ◽

Steady State ◽

Hydrogen Exchange ◽

Thick Ascending Limb ◽

Acid Base Balance ◽

No Donor ◽

Base Balance ◽

Dimethyl Amiloride ◽

Oxide Synthase ◽

Spermine Nonoate

Nitric oxide (NO) inhibits transport in various nephron segments, and the thick ascending limb (TAL) expresses nitric oxide synthase (NOS). However, the effects of NO on TAL transport have not been extensively studied. We tested the hypothesis that NO inhibits apical and basolateral Na+/H+exchange by the TAL by measuring intracellular pH (pHi) of isolated, perfused rat TALs using the fluorescent dye 2′,7′-bis-(2-carboxyethyl)-5-(and-6)-carboxyfluorescein (BCECF). The NO donor spermine NONOate (SPM, 10 μM) decreased steady-state pHi in medullary TALs from 7.18 ± 0.13 to 7.13 ± 0.14 ( P < 0.02), whereas controls did not decrease significantly. We next measured the buffering capacity of medullary TALs and the rate at which they recovered from acid loads to investigate the mechanism whereby NO reduces steady-state pHi. SPM decreased H+ flux ( J H) from 2.41 ± 0.66 to 0.97 ± 0.19 pmol ⋅ min−1 ⋅ mm−1, 55%. To assure that the decrease in J H was due to NO, another donor, nitroglycerin (NTG; 10 μM), was used. NTG decreased J H from 1.65 ± 0.11 to 1.07 ± 0.24 pmol ⋅ min−1 ⋅ mm−1, 37%. To determine the relative contributions of the apical and basolateral Na+/H+exchangers, 5-( N, N-dimethyl)amiloride (DMA; 100 μM) was added to either bath or lumen. With DMA added to the bath, SPM decreased J H from 4.78 ± 1.08 to 2.74 ± 0.54 pmol ⋅ min−1 ⋅ mm−1, an inhibition of 41%; and with DMA added to the lumen, SPM decreased J H from 2.31 ± 0.29 to 1.74 ± 0.27 pmol ⋅ min−1 ⋅ mm−1, a reduction of 26%. Addition of DMA alone to both bath and lumen resulted in an 87% inhibition of J H. We conclude that NO inhibits both apical and basolateral Na+/H+exchangers and consequently may play an important role in regulating pHi and may alter acid/base balance by directly affecting bicarbonate absorption in the TAL.

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Regulation of metalloproteinases by nitric oxide in human trophoblast cells in culture

Reproduction Fertility and Development ◽

10.1071/rd01036 ◽

2001 ◽

Vol 13 (6) ◽

pp. 411 ◽

Cited By ~ 39

Author(s):

Virginia Novaro ◽

Alejandro Colman-Lerner ◽

Felipe Vadillo Ortega ◽

Alicia Jawerbaum ◽

Dante Paz ◽

...

Keyword(s):

Nitric Oxide ◽

Embryo Implantation ◽

No Production ◽

Trophoblast Cells ◽

Human Trophoblast ◽

No Donors ◽

Multinucleated Cells ◽

Nos Inhibitors ◽

Spermine Nonoate

The process of embryo implantation requires extensive remodelling of the endometrial extracellular matrix, a function largely performed by matrix-degrading metalloproteinases (MMPs). In the present study, we used trophoblast cells isolated from human term placentas to study the regulation of MMPs by nitric oxide (NO). Using a combination of zymography, Western blot and indirect immunofluorescence, we showed that MMP-2 and MMP-9 are increased during the conversion from low-motile cytotrophoblast cells to the highly motile and differentiated syncytiotrophoblast multinucleated cells. We also observed an increase in NO production and NO synthase (NOS) expression during this cellular differentiation process. In addition, we demonstrated a positive regulatory role of NO on the activity and protein expression of MMP-2 and MMP-9, because NO donors (NOC-18 and spermine-NONOate) or the NOS substrate (L-arginine) stimulate, whereas NOS inhibitors (NG-nitro-L-arginine methyl ester and NG-monomethyl-L-arginine) reduce the expression and gelatinolytic activity of MMP-2 and MMP-9 in isolated trophoblast cells. Taken together, these results suggest that, in differentiating trophoblasts, NO regulates the induction of matrix-degrading proteases required for invasion during embryo implantation.

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Nitric oxide regulates nitric oxide synthase-2 gene expression by inhibiting NF-κB binding to DNA

Biochemical Journal ◽

10.1042/bj3220609 ◽

1997 ◽

Vol 322 (2) ◽

pp. 609-613 ◽

Cited By ~ 128

Author(s):

Song Kyu PARK ◽

Hsin Lee LIN ◽

Sean MURPHY

Keyword(s):

Nitric Oxide ◽

Nitric Oxide Synthase ◽

Interferon Γ ◽

No Production ◽

No Donor ◽

Nitric Oxide Synthase 2 ◽

Oxide Synthase ◽

Spermine Nonoate ◽

Dna Complex

Treatment of astroglial cells with interleukin 1β and interferon γ transcriptionally activates the nitric oxide synthase (NOS)-2 gene. The duration of mRNA expression is brief because of transcript instability. In addition, NO donors reduce the expression of NOS-2 mRNA dramatically by reducing the rate of transcription. In this study we observed that the NO donor, spermine NONOate did not inhibit the activation and translocation of NF-κB, a key transcription factor in the induction of NOS-2, but inhibited formation of the NF-κB–DNA complex. This effect was reversed by methaemoglobin (acting as an NO trap) and by the reducing agent dithiothreitol. Formation of the interferon-regulatory factor–DNA complex was unaffected by NO. These results suggest that NO can modulate its own production by interfering with NF-κB interaction with the promoter region of the NOS gene, a negative feedback effect that may be important for limiting NO production in vivo.

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Nitric Oxide Induces Resensitization of P2Y Nucleotide Receptors in Cultured Rat Mesangial Cells

Journal of the American Society of Nephrology ◽

10.1681/asn.v132313 ◽

2002 ◽

Vol 13 (2) ◽

pp. 313-321

Author(s):

Ruisheng Liu ◽

Antonio M. Gutiérrez ◽

Avi Ring ◽

A. Erik G. Persson

Keyword(s):

Nitric Oxide ◽

Intracellular Calcium ◽

Mesangial Cells ◽

Imaging Techniques ◽

No Production ◽

Calcium Stores ◽

Receptor Desensitization ◽

Glomerular Mesangial Cells ◽

Significant Difference ◽

Spermine Nonoate

ABSTRACT. Receptor desensitization of G protein–coupled receptors (GPCRs), which occurs during short-term (seconds to minutes) exposure of cells to agonists, is mediated by phosphorylation and receptor endocytosis. Recycling of the receptors is a requisite for resensitization of the response. The mechanisms that attenuate signaling by GPCRs are of considerable importance to regulation of intercellular signaling and maintenance of their ability to respond to agonists over time. This study evaluates the effect of nitric oxide (NO) on P2Y nucleotide receptor resensitization in cultured rat glomerular mesangial cells. The NO production in cultured mesangial cells was measured by using confocal microscopy and the fluorescence NO indicator 4,5-diaminofluorescein diacetate (DAF-2 DA). l-arginine increased and Nω-nitro-l-arginine methyl ester (l-NAME) decreased NO production significantly (P < 0.05). Calcium responses to ATP were measured with fura-2 and imaging techniques. Repeated stimulation with ATP results in receptor desensitization that is characterized by lower calcium peak amplitude. Desensitization was induced by challenging mesangial cells with four consecutive 2-min pulses of ATP (0.1 mM) separated by 4.5-min control perfusions. Intracellular calcium concentration ([Ca2+]i) increase evoked by second, third, and fourth ATP challenges were about 40%, 26%, and 18% of the first one. The NO precursor, l-arginine (10 mM), and the NO donors, spermine-NONOate (500 μM) and sodium nitroprusside (SNP) (1 mM), were added before and during a fourth ATP challenge. Spermine-NONOate and l-arginine induced a recovery of the [Ca2+]i response to the fourth ATP challenge (P < 0.01 and 0.05, respectively). The NO synthase inhibitor, l-NAME (5 mM), applied along with ATP, was shown to enhance desensitization. 1H-(1,2,4)oxadiazolo(4,3-α)quinoxalin-1-one (ODQ, 30 μM), an inhibitor of guanylate cyclase, was used along with l-arginine, SNP, or spermine-NONOate. There was no significant difference with or without ODQ. Neither ODQ nor 8-Br-cGMP, an analog of cGMP, at different concentrations showed effects on ATP-stimulated [Ca2+]i. There was no elevation of [Ca2+]i when the cells were challenged by different concentrations (1 μM, 100 μM, 1 mM, 20 mM, and 30 mM) of caffeine, caffeine plus ATP (0.1 mM), and 4-chloro-3-ethylphenol (100 μM, 500 μM, and 1 mM), a new agonist of ryanodine receptors. The results indicate that NO can increase the P2Y receptor resensitization in rat glomerular mesangial cells by acting through a cGMP-independent pathway. No evidence was found for the existence of ryanodine-sensitive intracellular calcium stores in rat mesangial cells.

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