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Effect of sodium nitrite on the execution of defensive and inhibitory conditioned reflexes.

Administration of sodium nitrite (an NO-generating preparation) doses of 11 mg/kg to rabbits was followed by motor disinhibition, with increases in myogram amplitudes in responses to nonreinforced flashes presented on a background of continuous illumination which served as a conditioned inhibitory stimulus; these changes appeared 1-1.5 h after administration and lasted throughout the 4-h observation period. After sodium nitrite doses of 5.5 mg/kg, responses to conditioned inhibition showed only a tendency to motor disinhibition. These results can be explained in terms of the ability of sodium nitrite to inhibit the function of GABAergic receptors, as development of internal inhibition is accompanied by increased inhibitory hyperpolarization processes in which the GABAergic mediator system is involved. The lower and higher sodium nitrite doses produced no increases in myogram amplitudes in responses to combinations of flashes of light and pain reinforcement, as occurred when these stimuli were presented to controls.

Animals↗

Nitrate and nitrite transport in bacteria.

The topological arrangements of nitrate and nitrite reductases in bacteria necessitate the synthesis of transporter proteins that carry the nitrogen oxyanions across the cytoplasmic membrane. For assimilation of nitrate (and nitrite) there are two types of uptake system known: ABC transporters that are driven by ATP hydrolysis, and secondary transporters reliant on a proton motive force. Proteins homologous to the latter type of transporter are also involved in nitrate and nitrite transport in dissimilatory processes such as denitrification. These proteins belong to the NarK family, which is a branch of the Major Facilitator Superfamily. The mechanism and substrate specificity of transport via these proteins is unknown, but is discussed in the light of sequence analysis of members of the NarK family. A hypothesis for nitrate and nitrite transport is proposed based on the finding that there are two distinct types of NarK.

Anion Transport Proteins↗

Is there a role of taurine bromamine in inflammation? Interactive effects with nitrite and hydrogen peroxide.

OBJECTIVE AND DESIGN: The myeloperoxidase system of neutrophils generates chlorinating and brominating oxidants in vivo. The major haloamines of the system are taurine chloramine (TauCl) and taurine bromamine (TauBr). It has been demonstrated in vitro that TauCl exerts both antiinflammatory and anti-bacterial properties. Much less is known about TauBr. The present study was conducted to compare bactericidal and immunoregulatory capacity of TauBr with that of the major chlorinating oxidants: HOCl and TauCl. Moreover, the effect of nitrites and H(2)O(2) on TauBr activity was investigated. MATERIALS: TauBr was prepared by reaction of HOBr with taurine. The reaction was monitored by UV absorption spectra. METHODS: Bactericidal activity of TauBr, TauCl and HOCl was tested by incubation of E. coli with the compounds and determined by the pour-plate method. To test the anti-inflammatory activity the compounds were incubated with LPS and IFN-gamma stimulated murine peritoneal macrophages. The production of following mediators was measured: nitrites by Griess reaction; TNF-alpha, IL-6, IL-10, IL-12p40 using capture ELISA. In some experiments the compounds were incubated with either nitrites or H(2)O(2). RESULTS: In our experimental set-up TauBr and HOCl exerted strong bactericidal effects on E. coli (MBC = 110 microM and 8 microM, respectively), while TauCl (< 1000 microM) did not kill test bacteria. However, both, TauBr and TauCl, at noncytotoxic concentrations (< 300 microM) inhibited the cytokine and nitric oxide production by macrophages. H(2)O(2) completely abolished the biological activities of TauBr but not those of TauCl. Nitrites did not affect any activity of TauBr or TauCl while they diminished the HOCl(-) mediated bacterial killing. CONCLUSION: TauBr, despite very low concentration of Br(-) in body fluids, may support TauCl and HOCl in the regulation of inflammatory response and in killing of bacteria by neutrophils. However, TauBr activity in vivo will depend on the presence of H(2)O(2) and possible other mediators of inflammation which can compete with target molecules for TauBr.

Animals↗

Rhodococcus sp. RB1 grows in the presence of high nitrate and nitrite concentrations and assimilates nitrate in moderately saline environments.

Rhodococcus sp. RB1 was able to thrive in media with up to 0.9 M NaCl or KCl and in the presence of high concentrations of nitrate (up to 0.9 M) and nitrite (up to 60 mM), but only under oxic conditions. An adaptation period was not required for salt tolerance, but a rapid extrusion of K+ and intake of Na+ was observed after addition of 0.5 M NaCl. Nitrate assimilation was limited by the carbon supply, but nitrite was not accumulated in the culture medium, even at nitrate concentrations as high as 0.8 M, thus suggesting that nitrite reduction does not limit nitrate assimilation. The presence of NaCl or KCl did not affect nitrate or nitrite uptake, which were completely inhibited by ammonium or glutamine. Rhodococcus sp. RB1 nitrate reductase had an apparent molecular mass of 142 kDa and used NADH and reduced bromophenol blue or viologens as electron donors, independently of the presence of salt. The enzyme was associated with an NADH-diaphorase activity and was induced by nitrate and repressed by ammonium or glutamine, thus showing typical biochemical and regulatory properties of bacterial assimilatory NADH-nitrate reductases. The enzyme was active in vitro in the presence of 3 M NaCl or KCI, but the maximal activity was observed at 0.5 M salt. Addition of 2 M NaCl increased the optimal temperature of the enzyme from 12 to 32 degrees C, but the optimal pH (10.3) was unaffected.

Cell Division↗

Competition and coexistence of aerobic ammonium- and nitrite-oxidizing bacteria at low oxygen concentrations.

In natural and man-made ecosystems nitrifying bacteria experience frequent exposure to oxygen-limited conditions and thus have to compete for oxygen. In several reactor systems (retentostat, chemostat and sequencing batch reactors) it was possible to establish co-cultures of aerobic ammonium- and nitrite-oxidizing bacteria at very low oxygen concentrations (2-8 microM) provided that ammonium was the limiting N compound. When ammonia was in excess of oxygen, the nitrite-oxidizing bacteria were washed out of the reactors, and ammonium was converted to mainly nitrite, nitric oxide and nitrous oxide by Nitrosomonas-related bacteria. The situation could be rapidly reversed by adjusting the oxygen to ammonium ratio in the reactor. In batch and continuous tests, no inhibitory effect of ammonium, nitric oxide or nitrous oxide on nitrite-oxidizing bacteria could be detected in our studies. The recently developed oxygen microsensors may be helpful to determine the kinetic parameters of the nitrifying bacteria, which are needed to make predictive kinetic models of their competition.

Ammonia↗

Urinary nitrite excretion and urinary variables in patients with primary nocturnal frequency of micturition: effects of indomethacin suppositories.

Urinary nitrite excretion was measured in patients with primary nocturnal frequency of micturition (PNFM) and in normal individuals. Effects of indomethacin suppository on urine volume and other urinary variables were evaluated. The study comprised seven patients with PNFM and seven healthy control (age range 30-45 years). Nitrite was assayed in spot morning urine samples; urine volume, urine osmolality and electrolytes, serum osmolality and electrolytes and functional bladder capacity (FBC) were assayed. Both groups were then given 100 mg of indomethacin suppository daily for a maximum of 10 days and urinary variables were re-evaluated during day 10. Results showed that urinary nitrite excretion of patients with PNFM was greater than that of the normal subjects (230+/-62 umol/l vs. 42+/-30 umol/l, P<0.05). The mean (SD) 24 h urine volume and osmolality, the night urine volume and osmolality, serum osmolality, FBC, creatinine clearance, fractional excretion of sodium (FENa), fractional excretion of potassium (FEK), and urinary excretion of glucose and potassium were lower in patients with PNFM as compared with normal individuals, although not statistically significantly so, except for FBC that was significantly lower in the patients. Urinary excretion of sodium, calcium, chloride, phosphorus, magnesium, day-night urinary volume ratio, spot morning osmolality, nocturnal index, and nocturnal polyuria index were higher in patients with PNFM. Indomethacin decreased the 24 h urinary volume by 21%, creatinine clearance by 12%, osmolar clearance by 14% and urinary protein excretion by 38% in the patients. These variables decreased by 26, 45, 17 and 12% respectively in the healthy subjects, whereas 24 h urinary protein excretion increased mildly by 9%. Indomethacin increased day-night urinary volume ratio by 73% in the healthy subjects. It might be concluded that urinary nitrite excretion, urinary excretion of sodium, chloride, phosphorus, calcium, and magnesium increased and FBC decreased in patients with PNFM; Indomethacin decreased urinary volume, FENa, FEK, osmolar clearance, and free water clearance in the healthy subjects and the patients. These might explain the mechanism of action of indomethacin to reduce frequency of voiding. The possible interaction of prostaglandin and NO in the pathogenesis of PNFM is discussed.

Adult↗

beta-Adrenergic agonist-induced nitrite production in isolated pig ciliary processes.

BACKGROUND: We performed an experiment to assess whether in ciliary processes adrenergic-receptor agents can modulate the production of nitrite, an oxidized metabolite of nitric oxide (NO). METHODS: The porcine ciliary processes and the iris were dissected and stored at -20 degrees C. Later, tissues were thawed, and nitrite measured by Griess reaction, before and 2 h after exposure to various drugs. RESULTS: In the ciliary processes, but not in the iris, the alpha-, beta-adrenergic receptor agonist, norepinephrine (0.01-10 microM), increased nitrite production (95 +/- 12%, P < 0.01). The norepinephrine-induced (0.1 microM) nitrite production was inhibited by the inhibitor of NO synthase, N omega-nitro-L-arginine methyl ester (0.5 mM; P < 0.05) and by the non-selective beta-adrenergic receptor antagonist, propranolol (10 microM; P < 0.01), but not by the non-selective alpha-adrenergic receptor antagonist, phentolamine (10 microM). CONCLUSION: In isolated porcine ciliary processes, beta-adrenergic receptor stimulation increases NO production.

Adrenergic beta-Agonists↗

Adrenomedullin and nitrite levels in children with Bartter syndrome.

Children with Bartter syndrome have lower than normal vascular reactivity with normotension in spite of biochemical and hormonal abnormalities which are typical of hypertension. Nitric oxide (NO) is a potent endogenous vasodilator, and plays an important role in the control of vascular tone. Adrenomedullin (AM) is a novel hypotensive peptide originally isolated from human pheochromocytoma. The possible role of NO and AM in maintaining this reduced vascular reactivity was examined by studying plasma and urinary nitrite, a stable metabolite of NO, and AM levels in ten children with Bartter syndrome, ten healthy controls, and five children with hypokalemia of causes other than Bartter syndrome (pseudo-Bartter). Urinary excretion of nitrite (mumol/mg urinary creatinine) was 8.9 +/- 1.2 in children with Bartter syndrome, 4.7 +/- 0.9 in healthy controls, and 2.9 +/- 0.8 in pseudo-Bartter (P < 0.05). Plasma nitrite levels (mumol/l) were 101.9 +/- 23.4, 59.9 +/- 14.7, and 65.0 +/- 29.7, respectively (P < 0.05), in the three groups. Urinary excretion of AM (pmol/mg urinary creatinine) was 187 +/- 40, 65 +/- 10, and 160 +/- 50, respectively (P < 0.05), in the three groups. Plasma AM levels were 47.4 +/- 1.8, 39.9 +/- 5.9, and 42.4 +/- 3.9, respectively (P > 0.05), in the three groups. The same parameters were repeated in the two groups of controls and in the Bartter patients in the 6th month of therapy. Urinary nitrite and AM levels were still higher in the Bartter patients than in the other groups. We conclude that in Bartter syndrome the increased NO production may be responsible for the reduced vascular response of the disease. Initially, increased levels of AM in Bartter syndrome and pseudo-Bartter may be a compensatory response to acute hypokalemia; however, continuation of a high level of urinary excretion of AM in children with Bartter syndrome may suggest also the possible role of AM in the reduced vascular response of the disease.

Adolescent↗

The cerebrospinal fluid oxidized NO metabolites, nitrite and nitrate, in Alzheimer's disease and vascular dementia of Binswanger type and multiple small infarct type.

The concentration of the nitric oxide (NO) metabolites nitrite (NO2-) and nitrate (NO3-) in the cerebrospinal fluid from patients with Alzheimer's disease (AD), vascular dementia of the Binswanger type (VDBT) or multiple small infarct type (MSID), and controls were determined using high performance liquid chromatography. The nitrite concentration was significantly higher in VDBT/MSID patients than in controls (p < 0.005). The nitrate concentration and the combined nitrite and nitrate concentration was significantly higher in both AD (p < 0.05) and VDBT/MSID (p < 0.001) patients than in controls, with these concentrations being significantly greater in VDBT/MSID than AD patients (p < 0.005). The combined nitrite and nitrate concentration significantly decreased as the severity of dementia progressed in AD (rs=0.70, p < 0.01), but remained elevated in all stages of VDBT/MSID. These results suggest that NO production or oxidation in the brain increases in the early stage of AD and then decreases as neuronal cell loss progresses, but increases throughout the course of disease in VDBT/MSID, which may in part contribute to neuronal degeneration in both conditions.

Aged↗

Kinetic and mechanistic studies of the reactions of nitrogen monoxide and nitrite with ferryl myoglobin.

Nitrogen monoxide (nitric oxide) generated endogenously has a variety of different properties. Among others it regulates blood pressure and transmission of nerve impulses, and has been shown to exert specific toxic effects, but also, paradoxically, to protect against various toxic substances. Recent studies suggest that NO* can serve as an antioxidant of the highly oxidizing ferryl myoglobin (MbFe(IV)=O), which has been proposed to be at least in part responsible for the oxidative damage caused by the reperfusion of ischemic tissues. In the present work we have determined the rate constant for the reaction between MbFe(IV)=O and NO* [(17.9+/-0.5)x10(6)M(-1)s(-1) at pH 7.5 and 20 degrees C] and we have shown that this reaction proceeds via the intermediate nitrito-metmyoglobin complex MbFe(III)ONO. Our results imply that this reaction is very likely to take place in vivo and might indeed represent a detoxifying pathway for both MbFe(IV)=O as well as NO*. Moreover, we have found that the rate of reaction of MbFe(IV)=O with nitrite is significantly lower (16+/-1 M(-1) s(-1) at pH 7.5 and 20 degrees C). Thus, this reaction probably plays a role only when NO* has been consumed completely and large concentrations of nitrite are still present. In contrast to the protecting role of NO*, the reaction with nitrite generates nitrogen dioxide which can contribute to tyrosine nitration. Indeed, we have demonstrated that nitrite can nitrate added tyrosine in the presence of iron(III) myoglobin and hydrogen peroxide.

Kinetics↗

Effects of amyl nitrite on left ventricular relaxation in patients with borderline hypertension.

The effects of systemic hypertension on left ventricular relaxation properties remain largely undefined. To assess such effects 22 normal volunteers and 15 patients with borderline hypertension were examined. The tangent to the echocardiographic left ventricular posterior wall endocardium was measured in diastole and was normalized for end-diastolic dimension to yield normalized velocity of relaxation. This velocity of relaxation was measured at rest and throughout inhalation of amyl nitrite. Mean value +/- standard deviation (SD) from rest to peak amyl nitrite effect for the normal group and for the patients with borderline hypertension was 3.3 +/- 0.6 leads to 7.2 +/- 1.1 and 3.0 +/- 0.8 leads to 4.4 +/- 1.1 s-1, respectively. All 22 persons in the normal group and 2 of the 15 patients with borderline hypertension attained normalized velocity of relaxation greater than 5.5 s-1 with administration of amyl nitrite. Multivariate analysis in the normal group identified heart rate, mean arterial pressure, and fractional shortening as the best predictors of normalized left ventricular relaxation velocity (r = 0.85; p less than 0.001). The increase in the normalized velocity of relaxation induced by amyl nitrite is blunted in patients with borderline hypertension. These changes in left ventricular relaxation identify early cardiac involvement and may prove clinically useful in hypertensive patients.

Adult↗

The efficacy of the nitrite test and microscopic urinalysis in predicting urine culture results.

The efficacy of the nitrite dipstick and microscopic urinalysis in predicting culture results were evaluated for 146 urine cultures from 56 women at risk for recurrent pyelonephritis. There were 111 negative cultures, 18 positive cultures, and 17 contaminated cultures. Nitrites were not detectable in either negative or contaminated cultures. Bacilluria was present in 10% of negative cultures and in 18% of contaminated cultures. Pyuria was present in 14% of negative and in 24% of contaminated cultures. Among the 18 positive cultures, nitrite was present in only 22%, bacilluria in only 61%, and pyuria in only 67%. These results suggest that positive cultures cannot be accurately predicted by either microscopic urinalysis or the nitrite dipstick, and that routine culturing should be performed as part of the outpatient management of women at risk for recurrent pyelonephritis.

Bacteriological Techniques↗

Nitrite reductase system involved in the terminal oxidation of the Streptomyces griseus respiratory particle.

A nitrite reductase system which was associated with the electron transfer system of the respiratory particle in Streptomyces griseus was studied. The electron transfer pathway consisted of the cytochrome oxidase and the nitrite reductase systems under aerobic and anaerobic conditions respectively, and these systems showed the exact opposite response to 2-n-heptyl-4-hydroxyquinoline-N-oxide and azide. Azide inhibited specifically the nitrite reductase system. It seems that cytochrome d works as the nitrite reductase and the reduced cytochrome b works as an intermediate electron donor for cytochrome d respectively. The respiratory particle also had a hydroxylamine reductase activity and ammonia was identified as the product of hydroxylamine reduction by the respiratory particle. A terminal electron transfer pathway in Streptomyces griseus was proposed.

Anaerobiosis↗

The interaction of nitrite with photosynthetic electrom transport under anaerobic conditions.

Chlorella cells were examined in a modulated oxygen polarograph under aerobic and anaerobic conditions. At light intensities below about 600 ergs . cm-2 . s-1 of 650 nm light, the oxygen yield and phase lag are lower under anaerobic conditions. Addition of 25 mM sodium nitrite increases both these parameters to values close to those found in the presence of oxygen. It is proposed that nitrite is reduced by Photosystem I thus diverting electrons from the cyclic electron transport pathway. The intersystem electron transport chain becomes more oxidized and this suppresses a backflow of electrons to the oxidizing side of Photosystem II, hence increasing the oxygen yield and the phase lag. The removal of oxygen from the bathing medium also alters the response of dark adapted Chlorella to a series of saturating light flashes. In terms of the Kok model of Photosystem II (Kok, B., Forbush, B. and McGloin, M. (1970) Photochem. Photobiol. 11, 457-475) there is a large increase in the parameter alpha. Addition of nitrite reverses this change and virtually restores the response seen in the presence of oxygen. The deactivation of the S2 state is greatly speeded up in the absence of oxygen but the addition of nitrite again reverses this.

Anaerobiosis↗

Nitric oxide formed by nitrite reductase of Paracoccus denitrificans is sufficiently stable to inhibit cytochrome oxidase activity and is reduced by its reductase under aerobic conditions.

Nitric oxide, generated by the action of purified nitrite reductase, inhibited the oxidase activity of both membrane vesicles from anaerobically grown Paracoccus denitrificans and bovine heart submitochondrial particles. In the former case, the inhibition was relatively short-lived and its duration was reduced either by decreasing the concentration of nitrite or raising the ratio of vesicles to nitrite reductase enzyme. These observations indicate that nitric oxide, at least at low concentrations, was sufficiently stable in the presence of oxygen to allow diffusion between proteins in aqueous solution. The shorter inhibition period with P. denitrificans membrane vesicles implies that the nitric oxide reductase of the vesicles is active in the presence of oxygen and has a sufficiently high affinity for nitric oxide to remove it from oxidase enzymes by competition. These observations are related to previous reports of potent inhibition under certain conditions of oxidase activity of P. denitrificans cells by a molecular species produced from nitrite. The implications of the deduced stability of nitric oxide in aerobic solutions are considered with respect to both the phenomenon of aerobic denitrification and the synthesis of nitric oxide in mammalian cells.

Aerobiosis↗

Stoichiometry of the reaction of oxyhemoglobin with nitrite.

During the reaction of oxyhemoglobin (HbO2) with nitrite, the concentration of residual nitrite, nitrate, oxygen, and methemoglobin (Hb+) was determined successively. The results obtained at various pH values indicate the following stoichiometry for the overall reaction: 4HbO2 + 4NO2- 4H+ leads to 4Hb+ + 4NO3- + O2 + 2H2 O (Hb denotes hemoglobin monomer). NO2- binds with methemoglobin noncooperatively with a binding constant of 340 M-1 at pH 7.4 and 25 degrees C. Thus, the major part of Hb+ produced is aquomethemoglobin, not methemoglobin nitrite, when less than 2 equivalents of nitrite is used for the oxidation.

Hemoglobin A↗

Evaluation of nitrite production by human monocyte-derived macrophages.

Reactive nitrogen intermediates are important in the anti-tumor and anti-microbial activities of rodent macrophages, but it is not known whether this is the case for human macrophages. In the present study, nitrite concentrations in vitro were used as an indicator of reactive nitrogen intermediate production by mouse, rat, and human macrophages. Human macrophages derived by culturing peripheral blood monocytes did not consistently produce detectable nitrite levels in response to any stimulus examined. Human macrophages were viable and metabolically active as indicated by the MTT assay, and their respiratory burst response to phorbol myristate acetate was increased following incubation with Interferon-gamma, as expected for typical macrophages. In contrast, rat or mouse peritoneal macrophages produced nitrite concentrations of approximately 20-100 microM in response to lipopolysaccharide, Interferon-gamma, or both. These results demonstrate substantial differences in the production of nitrites by rodent and human macrophages. Because of the heterogeneity among macrophage populations, these findings may not be applicable to all human macrophage populations, but they suggest a need for caution in extrapolating from rodent studies regarding the role of reactive nitrogen intermediates in anti-tumor or anti-microbial functions of human macrophages.

Animals↗

Production of nitrite by neonatal rat microglial cells/brain macrophages.

Microglial cells/brain macrophages from neonatal rats were examined for their capacity to generate nitrite, a product of the NO pathway. Upon incubation with bacterial lipopolysaccharide (LPS) or rat interferon-gamma (IFN-gamma), cells from the microglia-enriched fraction released measurable amounts of nitrite into the supernatant within 24-48 hr. The production of nitrite was dependent on the cell number and the dose of IFN-gamma and LPS. It could be inhibited by NG-monomethylarginine. We conclude that activated microglial cells can secrete nitrite. Stimulation of the NO pathway in microglial cells may be relevant to the pathogenesis of inflammatory and autoimmune demyelinating diseases of the brain.

Animals↗