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PubMed1998

Dual spillover problem in the myocardial septum with nitrogen-13-ammonia flow quantitation.

UNLABELLED: Conventional cardiac PET modeling techniques for [13N]ammonia flow determination do not fully account for the effects of spillover of activity from the right ventricle (RV) onto the activity in the myocardial septum. The purpose of this study was to investigate and to quantitatively account and correct for this effect. METHODS: Simulations were performed to determine the error introduced by conventional quantitation using septal time-activity curves, which only account for left ventricle (LV) spillover. Furthermore, we explored two separate methods to account for the dual spillover problem: direct estimation of the RV and LV spillover fractions incorporated into the [13N]ammonia model by using the LV and RV input functions in the fit and estimation of the relative dispersion and time shift between the LV and RV input functions by fitting using only the LV input function. The simulated curves were fitted using a two-compartment [13N]ammonia model. Flow estimates from the conventional model and the models including either of the two correction procedures were compared with canine microsphere data. RESULTS: The influence of RV spillover on flow estimation in the septum is determined by several parameters (e.g., dispersion between the RV and LV input function). Depending on the value of these parameters, the septal flow may be underestimated by 0%-30%. The applied methods for correction of the dual spillover problem were comparable and allow for more accurate quantitation in the septum. The canine microsphere data revealed that flow underestimation in the septum is small but significant. CONCLUSION: Dual spillover in the myocardial septum can introduce significant errors in the estimation of flow by the conventional [13N]ammonia model fitting method, which does not properly account for the RV spillover. Adjusting for the RV spillover in one of the two proposed methods allows for more accurate quantitation of myocardial septal flow with [13N]ammonia PET data.

Ammonia↗
PubMedDate not supplied

Neurotoxicity of ammonia and glutamate: molecular mechanisms and prevention.

Ammonia is a main factor in the pathogenesis of hepatic encephalopathy. We found that acute ammonia toxicity is mediated by activation of NMDA receptors. Chronic moderate hyperammonemia prevents acute ammonia toxicity in rats. Chronic exposure of cultured neurons to 1 mM ammonia leads to impaired response of the NMDA receptor to activation by its agonists (due to decreased protein kinase C-mediated phosphorylation) and prevents glutamate (Glu) neurotoxicity. Compounds that prevent ammonia toxicity in mice (e.g. carnitine) also prevent Glu toxicity in cultured neurons. These compounds did not prevent activation of NMDA receptor or the rise of Ca2+. They interfered with subsequent steps in the toxic process. The protective effect of carnitine is mediated by activation of metabotropic Glu receptors. Agonists of mGluRs, especially of mGluR5, prevent Glu toxicity. Agonists of muscarinic receptors also prevent Glu toxicity and there seems to be an interplay between muscarinic and metabotropic Glu receptors in the protective effect. We have tried to identify intracellular events involved in the process of neuronal death. It is known that the rise of Ca2+ is an essential step. Glu leads to depletion of ATP; some compounds (e.g. carnitine) prevent Glu-induced neuronal death without preventing ATP depletion: additional events are required for neuronal death. Glu induces activation of Na+/K+-ATPase, which could be involved in the toxic process. Inhibitors of protein kinase C, calcineurin or nitric oxide synthase prevent Glu toxicity. Our results indicate that Glu toxicity can be prevented at different steps or by activating receptors coupled to the transduction pathways interfering with the toxic process. Agents acting on these steps could prevent excitotoxicity in vivo in animals.

Adenosine Triphosphate↗
PubMed2001

Spatiotemporal Stability of an Ammonia-Oxidizing Community in a Nitrogen-Saturated Forest Soil.

Elevated levels of nitrogen input into various terrestrial environments in recent decades have led to increases in soil nitrate production and leaching. However, nitrifying potential and nitrifying activity tend to be highly variable over space and time, making broad-scale estimates of nitrate production difficult. This study investigates whether the high spatiotemporal variation in nitrate production might be explained by differences in the structure of ammonia-oxidizing bacterial communities in nitrogen-saturated coniferous forest soils. The diversity of ammonia-oxidizing bacteria of the b-subgroup Proteobacteria was therefore investigated using two different PCR-based approaches. The first targeted the 16S rRNA gene and involved temporal temperature gradient electrophoresis (TTGE) of specifically amplified PCR products, with subsequent band excision and nucleotide sequence determination. The second approach involved the cloning and sequencing of PCR-amplified amoA gene fragments. All recovered 16S rDNA sequences were closely related to the culture strain Nitrosospira sp. AHB1, which was isolated from an acid soil and is affiliated with Nitrosospira cluster 2, a sequence group previously shown to be associated with acid environments. All amoA-like sequences also showed a close affinity with this acid-tolerant Nitrosospira strain, although greater sequence variation could be detected in the amoA analysis. The ammonia-oxidizing bacterial community in the nitrogen-saturated coniferous forest soil was determined to be very stable, showing little variation between different organic layers and throughout the year, despite large differences in the total Bacterial community structure as determined by 16S rDNA DGGE community fingerprinting. These results suggest that environmental heterogeneity affecting ammonia oxidizer numbers and activity, and not ammonia oxidizer community structure, is chiefly responsible for spatial and temporal variation in nitrate production in these acid forest soils.

Journal Article↗
PubMed1984

Inhibition of urea synthesis by pent-4-enoic acid: potentiation by ammonia.

Pent-4-enoic acid inhibited ureagenesis approximatively 90% in rat hepatocytes incubated with pyruvate, ammonia and ornithine. Inhibition of ureagenesis was much less with alanine as substrate (approximatively 10%). The addition of ammonia led to a drastic dose-dependent inhibition of ureagenesis by pent-4-enoate. Half-maximum effect of ammonia was observed at 0.2 mM concentration. Concomitant cellular concentrations of N-acetylglutamate were also drastically modified by the addition of ammonia as was the accumulation of citrulline. These data suggest that ammonia may seriously interfere with the metabolism of pent-4-enoic acid and leads to a dramatic potentiation of its toxicity.

Ammonium Chloride↗
PubMed1985

Visible spectrophotometric determination of cephalosporins and penicillins by indophenol derivatization with and without alkaline degradation to ammonia.

Cephalosporins and penicillins give reproducible yields of ammonia on degradation in 0.5 M sodium hydroxide solution at 100 degrees C: the ammonia formed was determined in the degraded solutions using the indophenol reaction. In another approach the ammonia driven off on refluxing alkaline solutions of the cephalosporin or penicillin was collected in dilute hydrochloric acid solution and determined using the indophenol reaction. For eight of the fourteen cephalosporins and penicillins studied identical yields were recorded using the two procedures: these varied from 29% for penicillin G to 137% for cephalonium based on the production of one ammonia molecule per beta-lactam molecule. For six other cephalosporins the distillation method gave substantially higher yields of ammonia than did the direct determination. Eight cephalosporins and penicillins were found to give substantial indophenol-type reactions without prior hydrolysis of the beta-lactam, but the sensitivities were usually lower than for the hydrolysis method. Manual spectrophotometric procedures for the determination of cephalosporins and penicillins based on these reactions have been developed.

Journal Article↗
PubMed2005

Effect of ozone treatment on ammonia removal of activated carbons.

In this work, activated carbons (ACs) were modified by ozone treatment to enhance the efficiency of removal of ammonia gas over the ACs. Surface properties of the ACs were confirmed by X-ray photoelectron spectroscopy (XPS) analysis and N2 adsorption isotherms at 77 K were investigated by BET and D-A methods to characterize the specific surface area, total pore volume, and micropore volume. The ammonia removal efficiency was confirmed by the gas-detecting tube technique. The results showed that the specific surface area and micropore volume of ACs were slightly destroyed as the ozone treatment time increased. However, the ozone treatment led to an increase in ammonia removal efficiency of ACs, mainly due to an increase of acid functional groups, such as carbonyl and ether groups, on carbon surfaces. It was revealed that the improvement of ammonia removal efficiency of ACs was greatly affected by the interfacial acid-base interactions between modified ACs and basic ammonia adsorbate.

Journal Article↗
PubMed2006

Possible implication of ciliary neurotrophic factor (CNTF) and beta-synuclein in the ammonia effect on cultured rat astroglial cells: a study using DNA and protein microarrays.

Astrocytes are considered the key cell in hepatic encephalopathy; although their precise role in the disease has not yet been determined, exposure to ammonia appears to have an important pathogenic effect. We exposed confluent cultures of rat astroglial cells to ammonia (5mM NH(4)Cl) for 1, 3, 5 and 7 days, and determined astroglial levels of actin, glial fibrillary acidic protein (GFAP), glutamine synthetase (GS), GLAST glutamate transporter, 25kDa heat-shock protein (HSP25), HSP60 and HSP70 by Western blot; the glutamine content in culture medium was measured by mass spectrometry. Significant increases were observed for GS, HSP60 and glutamine, and significant reductions for actin and GFAP. Astrocytes exposed to ammonia for 4 days were used to analyze the effect of ammonia in protein and DNA microarrays. After protein microarray data filtration by signal intensity, x-fold change and z-score, 11 proteins were selected, among which the significant increase in beta-synuclein was confirmed by Western blot. DNA microarray data filtration by intensity signal, x-fold change and p-value selected almost 600 genes. The significant increase in alpha-synuclein mRNA was confirmed by quantitative RT-PCR, but no change was observed in alpha-synuclein protein levels. A notable decrease in ciliary neurotrophic factor (CNTF) was demonstrated by Western blot after ammonia treatment, concurring with the reduction in CNTF mRNA observed in DNA microarrays. We discuss the possibility of a pathogenic role for CNTF and a protective role for beta-synuclein in experimental hyperammonemia. This study demonstrates the use of microarrays as tools to ascertain the possible implication of previously unidentified proteins in the pathogenesis of hepatic encephalopathy.

Ammonium Chloride↗
PubMedDate not supplied

Effects of ammonia on glutamate transporter (GLAST) protein and mRNA in cultured rat cortical astrocytes.

Ammonia is a neurotoxic substance which accumulates in brain in liver failure and it has been suggested that ammonia plays a key role in contributing to the astrocytic dysfunction characteristic of hepatic encephalopathy. In particular, the effects of ammonia may be responsible for the reduced astrocytic uptake of neuronally-released glutamate and high extracellular glutamate levels consistently seen in experimental models of hepatic encephalopathy. To further address this issue, [(3)H]-D-aspartate uptake was examined in primary rat cortical astrocyte cultures exposed to 5 mM ammonium chloride for a period of 7 days. In addition, reverse transcriptase-polymerase chain reaction (RT-PCR) and Western blot studies were performed to examine the mRNA and protein expression respectively of the glutamate transporter GLAST in ammonia-treated cells. Studies revealed a 57% (p<0.05) decrease in [(3)H]-D-aspartate uptake and a concomitant significant decrease in GLAST transporter protein (43%, p<0.05) and mRNA (32%, p<0.05) expression. The reduced capacity of astrocytes to reuptake glutamate following ammonia exposure may result in compromised neuron-astrocyte trafficking of glutamate and could thus contribute to the pathogenesis of the cerebral dysfunction characteristic of hyperammonemic syndromes such as hepatic encephalopathy.

ATP-Binding Cassette Transporters↗
PubMed2000

Removal of a high load of ammonia gas by a marine bacterium, Vibrio alginolyticus.

A newly isolated marine bacterium, Vibrio alginolyticus was used to remove a high load of ammonia gas. By a stepwise increase in ammonia supply over the concentration range of 120-2000 ppm (v/v), complete removal of ammonia was observed from the start of the experiment in a suspended culture of the bacterium in basal medium containing 3% NaCl. When cells were inoculated onto an inorganic packing material in a biofilter, and a high load of ammonia was introduced continuously under nonsterile conditions, the average percentage of gas removed exceeded 85% for a 61-d operation. The maximum removal capacity and the complete removal capacity were 22.8 g-N/kg-dry packing material/d and 18.6 g-N/kg-dry packing material/d, respectively, which were about four times larger than those obtained in nitrifying sludge inoculated onto the same packing material. During this operation, the nonsterile air supply had no adverse effect on the removability of ammonia by V. alginolyticus.

Journal Article↗
PubMed2005

Sensitivity of ammonia interaction with single-walled carbon nanotube bundles to the presence of defect sites and functionalities.

Ammonia adsorption on single-walled carbon nanotubes (SWNTs) was studied by means of infrared spectroscopy at both cryogenic (approximately 94 K) and room (approximately 300 K) temperatures. At 94 K, vacuum-annealed SWNTs showed no detectable ammonia uptake. However, the ammonia adsorption was found to be sensitive to the functionalities and defects on the nanotube surfaces. NH3 adsorption was detected on HNO3-treated nanotubes, characterized by significant functionalities and defects, prior to vacuum annealing. NH3 desorbed from those nanotubes above 140 K, indicating a weak adsorbate-nanotube interaction (approximately 30 kJ/mol). Exposure of annealed samples to ambient air, which possibly regenerated functionalities and defects on nanotube surfaces, restored partially the ammonia uptake capacity. No ammonia adsorption on SWNTs was observed by infrared spectroscopy at room temperature with up to 80 Torr dosing pressure. This work suggests the influence of functionalities and/or defect densities on the sensitivity of SWNT chemical gas sensors. Our theoretical studies on NH3 adsorption on pristine and defective tubes, as well as oxidized tubes, corroborate these findings.

Journal Article↗
PubMed2004

Ammonia adsorption on and diffusion into thin ice films grown on Pt(111).

Ammonia adsorption on and diffusion into thin ice films grown on a Pt(111) surface were studied using Fourier transform infrared spectroscopy (FTIR) and thermal desorption spectroscopy. After exposing the crystalline ice film to ammonia molecules at 45 K (ammonia/ice film), we have detected an intriguing feature at 1470 cm(-1) in the FTIR spectra, which is derived from the adsorption of ammonia on the ice with a characteristic structure which appears in thin film range. The peak intensity of this feature decreases gradually as the thickness of the substrate ice increases. In addition, we have detected a feature at 1260 cm(-1) which appears after annealing the ammonia/ice film. The feature corresponds to the ammonia molecules which reach the ice/Pt(111) interface through the ice film. Intriguingly, the intensity of this feature decreases with the ice thickness and there is a linear relation of the peak intensity of the features at 1470 and 1260 cm(-1). We propose a model in which the solubility of the ammonia molecules is much higher for the thin ice film than that for the ideal ice.

Journal Article↗
PubMed2005

Ubiquity and diversity of ammonia-oxidizing archaea in water columns and sediments of the ocean.

Nitrification, the microbial oxidation of ammonia to nitrite and nitrate, occurs in a wide variety of environments and plays a central role in the global nitrogen cycle. Catalyzed by the enzyme ammonia monooxygenase, the ability to oxidize ammonia was previously thought to be restricted to a few groups within the beta- and gamma-Proteobacteria. However, recent metagenomic studies have revealed the existence of unique ammonia monooxygenase alpha-subunit (amoA) genes derived from uncultivated, nonextremophilic Crenarchaeota. Here, we report molecular evidence for the widespread presence of ammonia-oxidizing archaea (AOA) in marine water columns and sediments. Using PCR primers designed to specifically target archaeal amoA, we find AOA to be pervasive in areas of the ocean that are critical for the global nitrogen cycle, including the base of the euphotic zone, suboxic water columns, and estuarine and coastal sediments. Diverse and distinct AOA communities are associated with each of these habitats, with little overlap between water columns and sediments. Within marine sediments, most AOA sequences are unique to individual sampling locations, whereas a small number of sequences are evidently cosmopolitan in distribution. Considering the abundance of nonextremophilic archaea in the ocean, our results suggest that AOA may play a significant, but previously unrecognized, role in the global nitrogen cycle.

Base Sequence↗
PubMed1997

Differential Ammonia-Elicited Changes of Cytosolic pH in Root Hair Cells of Rice and Maize as Monitored by 2[prime],7[prime]-bis-(2-Carboxyethyl)-5 (and -6)-Carboxyfluorescein-Fluorescence Ratio.

Intact hair cells of young rice (Oryza sativa L.) and maize roots (Zea mays L.), grown without external nitrogen, were specifically loaded with 2[prime],7[prime]-bis-(2-carboxyethyl)-5 (and -6)-carboxyfluorescein acetoxymethyl ester to monitor fluorescence ratio cytosolic pH changes in response to external ammonia (NH4+/NH3) application. In neutral media, cytosolic pH of root hairs was 7.15 [plus or minus] 0.13 (O. sativa) and 7.08 [plus or minus] 0.11 (Z. mays). Application of 2 mM ammonia at external pH 7.0 caused a transient cytosolic alkalization (7.5 [plus or minus] 0.15 in rice; 7.23 [plus or minus] 0.13 in maize). Alkalization increased with an increase of external pH; no pH changes occurred at external pH 5.0. The influx of 13N-labeled ammonia in both plant species did not differ between external pH 5.0 and 7.0 but increased significantly with higher pH. Pretreatment with 1 mM 1-methionine sulfoximine significantly reduced the ammonia-elicited pH increase in rice but not in maize. Application of 2 mM methylammonia only caused a cytosolic pH increase at high external pH; the increase in both species compared with the ammonia-elicited alkalization in 1-methionine sulfoximine-treated roots. The differential effects indicate that cytosolic alkalization derived from (a) NH3 protonation after passive permeation of the plasma membrane and, particularly in rice, (b) additional proton consumption via the glutamine synthetase/glutamate synthase cycle.

Journal Article↗
PubMed1968

Inhibition of water uptake in sugar beet roots by ammonia.

Ammonium sulfate, ammonium carbonate or ammonia gas inhibited water uptake in sugar beet roots whenever the pH was sufficiently high to cause the production of ammonia. When ammonia was removed by aeration, inhibition of the water uptake by roots was rapidly reversed. ATP at 0.2 mm appeared to either wholly or partially prevent the ammonia-induced inhibition of water uptake by roots. ATP may be involved in maintaining the structure of water pathways through the root. In roots lacking epidermis, ammonia did not inhibit water uptake by the roots. This may indicate that the site of the inhibition lies within the root epidermis.

Journal Article↗
PubMed1978

Circadian Rhythmicity in the Activities of Phenylalanine Ammonia-Lyase from Lemna perpusilla and Spirodela polyrhiza.

The oscillations in phenylalanine ammonia-lyase activity from Spirodela polyrhiza and phenylalanine ammonia-lyase and tyrosine ammonia-lyase activities from Lemna perpusilla displayed a circadian rhythm under continuous light. Rhythmicity in enzymic activity could not be detected in continuous darkness since under this condition phenylalanine ammonia-lyase activity remains at a fairly constantly low level. Results from our studies of the oscillatory pattern of the respective activities of phenylalanine and tyrosine ammonia-lyase support their "inseparability."

Journal Article↗
PubMed1981

Ammonia accumulation and inhibition of photosynthesis in methionine sulfoximine treated spinach.

Ammonia accumulation and photosynthetic rate inhibition took place when spinach leaf tissue was supplied with methionine sulfoximine (MSO), an inhibitor of glutamine synthetase. This effect was observed in the absence of significant inorganic nitrogen reduction or an exogenous source of ammonia. Both the time lag prior to the initial photosynthetic rate decrease and the rate of that decrease depend on the O(2) and MSO concentrations supplied to the leaf tissue. However, the total rate of ammonia accumulation was similar at both 20% and 2.2% O(2). The decline in photosynthetic rate was not caused by stomatal closure but may be a result of ammonia toxicity. The data point out the importance of glutamine synthetase in preventing the poisoning of leaf metabolism by ammonia generated internally through processes not involved in net nitrogen assimilation. The rapidity of the action of MSO in suppressing photosynthesis was unexpected and should not be overlooked in interpreting data from other experiments involving that inhibitor. MSO shows promise as a tool for investigating C-N flow, particularly during photorespiration.

Journal Article↗
PubMed1981

Effects of Light and of Fusarium solani on Synthesis and Activity of Phenylalanine Ammonia-Lyase in Peas.

Phenylalanine ammonia-lyase was purified from peas, and a specific antiserum against the enzyme was produced in rabbits. The antiserum was used to study the first 8 hours of the phenylalanine ammonia-lyase activity response in two different organs of the pea from different developmental stages and in response to two different stimuli. Etiolated seedlings were pulse-labeled with l-[(35)S]methionine after either no light exposure or after specific periods of irradiation with blue light. Immature pods were pulse labeled with mixed l-[(3)H]amino acids after specific time periods following inoculation of the pod endocarp surfaces with macroconidia of Fusarium solani. Immunoprecipitates isolated from extracts of each group were analyzed with sodium dodecyl sulfate disc gel electrophoresis and were found to contain a radioactive protein with an electrophoretic mobility identical to that of the phenylalanine ammonia-lyase subunit (M(r) 81,000). The radioactivity contained in the subunit band was interpreted as being due to de novo synthesis of the enzyme. The net rate of phenylalanine ammonia-lyase labeling, found to be initially low in both tissue types, rose dramatically, peaking at approximately a six- to ten-fold greater level at 4 hours after the beginning of the stimulus. Thereafter, the rate of labeling declined slowly. Inoculation with F. solani f. sp. pisi, a true pathogen of peas, caused a fifty per cent greater rate of peak labeling than did inoculation with a nonpathogen, F. solani f. sp. phaseoli. The time profile of the changing rate of labeling correlates with the changing activity level of the enzyme which peaks at 12 hours after the onset of the stimulus. The data presented favor a model which explains the changing activity of phenylalanine ammonia-lyase as being due to a changing rate of synthesis or degradation (or both) of the enzyme rather than due to the activation of a preformed zymogen.

Journal Article↗
PubMed1982

Stimulation of ammonia and 2-oxoglutarate-dependent o(2) evolution in isolated chloroplasts by dicarboxylates and the role of the chloroplast in photorespiratory nitrogen recycling.

Intact chloroplasts isolated from spinach (Spinacia oleracea L.) leaves showed a light-dependent O(2) evolution (5.5 +/- 0.75 micromoles per milligram chlorophyll per hour) when supplied with ammonia and 2-oxoglutarate. This (ammonia, 2-oxoglutarate)-dependent O(2) evolution was stimulated 2- to 4-fold by the dicarboxylates, malate, succinate, fumarate, glutarate, and l-tartarate. Evolution of O(2) in the presence of malate was dependent on the presence of both 2-oxoglutarate and NH(4)Cl; malate with only either 2-oxoglutarate and NH(4)Cl alone did not support O(2) evolution. Furthermore, in the presence of malate, the amount of O(2) evolved was solely dependent on the amount of NH(4)Cl or 2-oxoglutarate added and malate did not affect the ratio of O(2) evolved to NH(4)Cl or 2-oxoglutarate consumed. Studies with inhibitors (2-(3,4-dichlorophenyl)-1,1-dimethyl urea, methionine sulfoximine, and azaserine) indicated that the above activity was directly linked to glutamine synthetase and glutamate synthase activity in the chloroplast and was not caused by the metabolism of malate. The V(max)/2 of (ammonia, 2-oxoglutarate)-dependent O(2) evolution was reached at 32 micromolar NH(4)Cl and 6 millimolar (approximately) 2-oxoglutarate in the absence of malate, and at 22 micromolar NH(4)Cl and 73 micromolar 2-oxoglutarate when malate (3 millimolar) was present.Intact chloroplasts isolated from pea (Pisum sativum) leaves also showed a stimulation of (ammonia, 2-oxoglutarate)-dependent O(2) evolution by malate. However glutamine was required for this activity even though glutamine with only either NH(4)Cl or 2-oxoglutarate did not respond to malate stimulation.The measured rates of (ammonia, 2-oxoglutarate)-dependent O(2) evolution in isolated spinach chloroplasts in the presence of malate were about 19.5 +/- 4.5 micromoles O(2) evolved per milligram chlorophyll per hour. This is adequate to sustain photorespiratory NH(3) recycling and the refixation of NH(3) arising from NO(3) under ambient conditions in the light. The role of the chloroplast in photorespiratory NH(3) recycling and the nature of the associated transport of 2-oxoglutarate into the chloroplast is discussed.

Journal Article↗