In vivo neurochemical profiling of rat brain by 1H-[13C] NMR spectroscopy: cerebral energetics and glutamatergic/GABAergic neurotransmission

The quantification of excitatory and inhibitory neurotransmission and the associated energy metabolism is crucial for a proper understanding of brain function. Although the detection of glutamatergic neurotransmission in vivo by ¹³C NMR spectroscopy is now relatively routine, the detection of GABAergic neurotransmission in vivo has remained elusive because of the low GABA concentration and spectral overlap. Using ¹H-[¹³C] NMR spectroscopy at high magnetic field in combination with robust spectral modeling and the use of different substrates, [U-¹³C₆]-glucose and [2-¹³C]-acetate, it is shown that GABAergic, as well as glutamatergic neurotransmitter fluxes can be detected non-invasively in rat brain in vivo .     

Localized 13C NMR Spectroscopy in the Human Brain of Amino Acid Labeling from d-[1-13C]Glucose

Abstract: Cerebral metabolism of d [1-¹³C]glucose was studied with localized ¹³C NMR spectroscopy during intravenous infusion of enriched [1-¹³C]glucose in four healthy subjects. The use of three-dimensional localization resulted in the complete elimination of triacylglycerol resonance that originated in scalp and subcutaneous fat. The sensitivity and resolution were sufficient to allow 4 min of time-resolved observation of label incorporation into the C3 and C4 resonances of glutamate and C4 of glutamine, as well as C3 of aspartate with lower time resolution. [4-¹³C]Glutamate labeled rapidly reaching close to maximum labeling at 60 min. The label flow into [3-¹³C]glutamate clearly lagged behind that of [4-¹³C]glutamate and peaked at t = 110–140 min. Multiplets due to homonuclear ¹³C-¹³C coupling between the C3 and C4 peaks of the glutamate molecule were observed in vivo. Isotopomer analysis of spectra acquired between 120 and 180 min yielded a ¹³C isotopic fraction at C4 glutamate of 27 ± 2% (n = 4), which was slightly less than one-half the enrichment of the C1 position of plasma glucose (63 ± 1%), p < 0.05. By comparison with an external standard the total amount of [4-¹³C]glutamate was directly quantified to be 2.4 ± 0.1 µmol/ml-brain. Together with the isotopomer data this gave a calculated brain glutamate concentration of 9.1 ± 0.7 µmol/ml, which agrees with previous estimates of total brain glutamate concentrations. The agreement suggests that essentially all of the brain glutamate is derived from glucose in healthy human brain.     

Stable Isotopes and Carbon Cycle Processes in Forests and Grasslands

Abstract: Scaling and partitioning are frequently two difficult challenges facing ecology today. With regard to ecosystem carbon balance studies, ecologists and atmospheric scientists are often interested in asking how fluxes of carbon dioxide scale across the landscape, region and continent. Yet at the same time, physiological ecologists and ecosystem ecologists are interested in dissecting the net ecosystem CO₂ exchange between the biosphere and the atmosphere to achieve a better understanding of the balance between photosynthesis and respiration within a forest. In both of these multiple-scale ecological questions, stable isotope analyses of carbon dioxide can play a central role in influencing our understanding of the extent to which terrestrial ecosystems are carbon sinks. In this synthesis, we review the theory and present field evidence to address isotopic scaling of CO₂ fluxes. We first show that the ¹³C isotopic signal which ecosystems impart to the atmosphere does not remain constant over time at either temporal or spatial scales. The relative balances of different biological activities and plant responses to stress result in dynamic changes in the ¹³C isotopic exchange between the biosphere and atmosphere, with both seasonal and stand-age factors playing major roles influencing the ¹³C biosphere-atmosphere exchange. We then examine how stable isotopes are used to partition net ecosystem exchange fluxes in order to calculate shifts in the balance of photosynthesis and respiration. Lastly, we explore how fundamental differences in the ¹⁸O isotopic gas exchange of forest and grassland ecosystems can be used to further partition terrestrial fluxes.     

Convergence in the relationship of CO2 and N2O exchanges between soil and atmosphere within terrestrial ecosystems

Ecosystem CO₂ and N₂O exchanges between soils and the atmosphere play an important role in climate warming and global carbon and nitrogen cycling; however, it is still not clear whether the fluxes of these two greenhouse gases are correlated at the ecosystem scale. We collected 143 pairs of ecosystem CO₂ and N₂O exchanges between soils and the atmosphere measured simultaneously in eight ecosystems around the world and developed relationships between soil CO₂ and N₂O fluxes. Significant linear regressions of soil CO₂ and N₂O fluxes were found for all eight ecosystems; the highest slope occurred in rice paddies and the lowest in temperate grasslands. We also found the dominant role of growing season on the relationship of annual CO₂ and N₂O fluxes. No significant relationship between soil CO₂ and N₂O fluxes was found across all eight ecosystem types. The estimated annual global N₂O emission based on our findings is 13.31 Tg N yr⁻¹ with a range of 8.19–18.43 Tg N yr⁻¹ for 1980–2000, of which cropland contributes nearly 30%. Our findings demonstrated that stoichiometric relationships may work on ecological functions at the ecosystem level. The relationship of soil N₂O and CO₂ fluxes developed here could be helpful in biogeochemical modeling and large-scale estimations of soil CO₂ and N₂O fluxes.     

Root ammonium transport efficiency as a determinant in forest colonization patterns: an hypothesis

Ratios of ammonium (NH₄⁺) to nitrate (NO₃^–) in soils are known to increase during forest succession. Using evidence from several previous studies, we hypothesize that a malfunction in NH₄⁺ transport at the membrane level might limit the persistence of early successional tree species in later seral stages. In those studies, ¹³N radiotracing was used to determine unidirectional fluxes and pool sizes of NH₄⁺ and NO₃^– in seedlings of the late-successional species white spruce ( Picea glauca ) and in the early successional species Douglas-fir ( Pseudotsuga menziesii var. glauca ) and trembling aspen ( Populus tremuloides ). At high external NH₄⁺, the two early successional species accumulated excessive NH₄⁺ in the root cytosol, and exhibited high-velocity, low-efficiency (15% to 22%), membrane fluxes of NH₄⁺. In sharp contrast, white spruce had low cytosolic NH₄⁺ accumulation, and lower-velocity but much higher-efficiency (65%), NH₄⁺ fluxes. Because these divergent responses parallel known differences in tolerance and toxicity to NH₄⁺ amongst these species, we propose that they constitute a significant driving force in forest succession, complementing the discrimination against NO₃^– documented in white spruce (Kronzucker et al. 1997).     

Snow depth, soil freezing, and fluxes of carbon dioxide, nitrous oxide and methane in a northern hardwood forest

Soil–atmosphere fluxes of trace gases (especially nitrous oxide (N₂O)) can be significant during winter and at snowmelt. We investigated the effects of decreases in snow cover on soil freezing and trace gas fluxes at the Hubbard Brook Experimental Forest, a northern hardwood forest in New Hampshire, USA. We manipulated snow depth by shoveling to induce soil freezing, and measured fluxes of N₂O, methane (CH₄) and carbon dioxide (CO₂) in field chambers monthly (bi-weekly at snowmelt) in stands dominated by sugar maple or yellow birch. The snow manipulation and measurements were carried out in two winters (1997/1998 and 1998/1999) and measurements continued through 2000. Fluxes of CO₂ and CH₄ showed a strong seasonal pattern, with low rates in winter, but N₂O fluxes did not show strong seasonal variation. The snow manipulation induced soil freezing, increased N₂O flux and decreased CH₄ uptake in both treatment years, especially during winter. Annual N₂O fluxes in sugar maple treatment plots were 207 and 99 mg N m⁻² yr⁻¹ in 1998 and 1999 vs. 105 and 42 in reference plots. Tree species had no effect on N₂O or CO₂ fluxes, but CH₄ uptake was higher in plots dominated by yellow birch than in plots dominated by sugar maple. Our results suggest that winter fluxes of N₂O are important and that winter climate change that decreases snow cover will increase soil:atmosphere N₂O fluxes from northern hardwood forests.     

Seasonal metabolism of a small, arboreal monitor lizard, Varanus scalaris, in tropical Australia

The field metabolic rates (FMR) and water fluxes of Varanus scalaris were measured during the wet and dry seasons by the doubly-labelled water technique. Seasonal measurements of standard (night-time) metabolism (SMR) and resting (daytime) metabolism (RMR) were made in the laboratory at 18, 24, 30 and 36°C, and maximal oxygen consumption was measured at 36°C on a motorized treadmill. This population was active throughout the year. In the wet season, the mean FMR was 7.8 kJ day⁻¹ (128.0 kJkg⁻¹ day⁻¹; mean mass = 66.4 g, n = 13), and during the dry season the mean was 5.0 kJ day⁻¹ (67.6 kJ kg⁻¹ day⁻¹; mean mass = 77.4 g, n = 17). The mean water flux rates for these animals were 3.6 and 1.2 ml day⁻¹, respectively (60.4 and 16.6 ml kg⁻¹ day⁻¹). The seasonal means of FMR and water flux were significantly different by ANCOVA ( P < 0.0001). Measurements of SMR and RMR were significantly higher in the wet season (ANCOVA: P < 0.0001), but we found no difference in the maximal oxygen consumption between seasons (ANCOVA: P = 0.6). The maximal oxygen consumption of the lizards on the treadmill (2.9 ml min⁻¹= 1.8 ml g⁻¹ h⁻¹), mean mass = 97.4 g, n = 16) was 20 times that of the SMR at the same temperature during the dry season, and 11 times that of the SMR during the wet season. The seasonal differences in FMR were attributable to: changes in SMR (12.2%) and RMR (16.4%); differences in night-time body temperatures (11.3) and daytime body temperatures (16.4%); and activity (broadly defined to include locomotion, digestion, and reproductive costs (43.7%).     

Impairment of Na+,K+-ATPase activity following enterotoxigenic Campylobacter jejuni infection: changes in Na+, Cl− and 3-O-methyl-D-glucose transport in vitro, in rat ileum

Abstract Unidirectional fluxes of Na⁺, Cl⁻ and 3-O-methyl-D-glucose (3-MG) were measured in vitro across Campylobacter jejuni live culture-infected and control rat ileal short-circuited tissues by the Using Chamber technique. Net secretion of Na⁺ and enhanced secretion of Cl⁻ ions was observed in the infected animals ( P < 0.001, n =6) as compared to the net absorption of Na⁺ and marginal secretion of Cl⁻ ions in the control animals. There was a significant decrease in the mucosal-to-serosal fluxes of 3-MG in C. jejuni -infected rat ileum. The specific Na⁺,K⁺-ATPase activity when measured biochemically in the membrane-rich fraction of enterocytes was found to be significantly lower (58%) in the infected group as compared to the control group ( P < 0.001). Our results therefore suggest that infection with an enterotoxigenic C. jejuni inhibits the Na⁺,K⁺-ATPase activity in rat enterocytes. The impairment of Na⁺,K⁺-ATPase activity thus appears to induce a secondary change in Na⁺,Cl⁻ and 3-MG transport in vitro in rat ileum.     

Portacaval Anastomosis Results in Altered Neuron-Astrocytic Metabolic Trafficking of Amino Acids: Evidence from 13C-NMR Studies

Abstract: ¹³C-NMR spectroscopy was used to evaluate the dynamic consequences of portacaval anastomosis on neuronal and astrocytic metabolism and metabolic trafficking between neurons and astrocytes. Glutamate is predominantly labeled from [1-¹³C]glucose, whereas [2-¹³C]acetate is more efficient in labeling glutamine, in accordance with its primary metabolism in astrocytes. Alanine and succinate labeling was only observed with [1-¹³C]glucose as precursor. Brain [1-¹³C]glucose metabolism in portacaval-shunted rats was similar to that in sham-operated controls with the exception of labeled glutamine and succinate formation, which was increased in shunted rats. The ¹³C enrichment was, however, decreased owing to an increase in total glutamine and succinate. Using [2-¹³C]acetate, on the other hand, flux of astrocytic label to neurons was severely decreased because label incorporation into glutamate, aspartate, and GABA was decreased following portacaval shunting. The latter amino acids are predominantly localized in neurons. These findings demonstrate that metabolic trafficking of amino acids from astrocytes to neurons is impaired in portacaval-shunted rats.     

Cardiorespiratory status of triploid brown trout during swimming at two acclimation temperatures

At 14° C, standard metabolic rate (75·1 mg O₂ h⁻¹ kg⁻¹), routine metabolic rate (108.8 mg O₂ h⁻¹ kg⁻¹), active metabolic rate ( c . 380 mg O₂ h⁻¹ kg⁻¹), critical swimming speed (U_crit 1·7 BL s⁻¹), heart rate 47 min⁻¹), dorsal aortic pressure (3·2 kPa) and ventilation frequency (63 min⁻¹) for triploid brown trout Salmo trutta were within the ranges reported for diploid brown trout and other salmonids at the same temperature. During prolonged swimming ( c . 80% U _crit), cardiac output increased by 2·3-fold due to increases in heart rate (1·8-fold) and stroke volume (1·2-fold). At 18° C, although standard and routine metabolic rates, as well as resting heart rate and ventilation frequency increased significantly, active metabolic rate and certain cardiorespiratory variables during exercise did not differ from those values for fish acclimated to 14° C. As a result, factorial metabolic scope was reduced (2·93-fold at 18° C v . 5·13-fold at 14° C). Therefore, it is concluded that cardiorespiratory performance in triploid brown trout was not unusual at 18° C, but that reduced factorial metabolic scope may be a contributing factor to the mortality observed in triploid brown trout at temperatures near 18° C.     

Effects of spring flood and water level draw-down on methane dynamics in the littoral zone of boreal lakes

1. The annual dynamics of methane (CH₄) in a temporarily flooded meadow, mire bank, lacustrine sedge fen, temporarily and continuously inundated sedge ( Carex sp.) and reed ( Phragmites australis ) marshes were studied from June to November in the humic mesoeutrophic Lake Mekrijärvi and in eutrophicated parts of the mesotrophic Lake Heposelkä in the southern part of East Finland. The effects of water level and temperature on littoral CH₄ fluxes were determined. Vegetation zonation along the moisture gradient, and associated CH₄ fluxes, were evaluated.
2. The CH₄ flux increased along the moisture gradient from –0.2 to 14.2 mg CH₄ m^–2 h^–1, and was highest in the permanently inundated marshes. The duration of anoxia in the sediment caused differences in the CH₄ flux. Estimated emissions for the period 1 June – 30 September in continuously inundated sparse reed and sedge marshes, drying sedge marsh, and lacustrine sedge fen were 13, 11 and 6 g CH₄ m^–2, respectively.
3. In continuously inundated vegetation, the fluxes were highest in late July/early August. The seasonal CH₄ flux pattern suggested that the fluxes were regulated by the supply of organic matter during the course of the summer and the water level. In the temporarily flooded zone, the seasonal CH₄ flux dynamics was greatly affected by changes in the lake water level, the fluxes being highest during the spring flood in early June.     

Quantification of the GABA Shunt and the Importance of the GABA Shunt Versus the 2-Oxoglutarate Dehydrogenase Pathway in GABAergic Neurons

Abstract: We investigated the activity of the cerebral GABA shunt relative to the overall cerebral tricarboxylic acid (TCA) cycle and the importance of the GABA shunt versus 2-oxoglutarate dehydrogenase for the conversion of 2-oxoglutarate into succinate in GABAergic neurons. Awake mice were dosed with [1-¹³C]glucose, and brain extracts were analyzed by ¹³C NMR spectroscopy. The percent enrichments of GABA C-2 and glutamate C-4 were the same: 5.0 ± 1.6 and 5.1 ± 0.2%, respectively (mean ± SD). This, together with previous data, indicates that the flux through the GABA shunt relative to the overall cerebral TCA cycle flux equals the GABA/glutamate pool size ratio, which in the mouse is 17%. It has previously been shown that under the experimental conditions used in this study, the ¹³C labeling of aspartate from [1-¹³C]glucose specifically reflects the metabolic activity of GABAergic neurons. In the present study, the reduction in the formation of [¹³C]aspartate during inhibition of the GABA shunt by γ-vinyl-GABA indicated that not more than half the flux from 2-oxoglutarate to succinate in GABAergic neurons goes via the GABA shunt. Therefore, because fluxes through the GABA shunt and 2-oxoglutarate dehydrogenase in GABAergic neurons are approximately the same, the TCA cycle activity of GABAergic neurons could account for one-third of the overall cerebral TCA cycle activity in the mouse. Treatment with γ-vinyl-GABA, which increased GABA levels dramatically, caused changes in the ¹³C labeling of glutamate and glutamine, which indicated a reduction in the transfer of glutamate from neurons to glia, implying reduced glutamatergic neurotransmission. In the most severely affected animals these alterations were associated with convulsions.     

Tracing carbon and oxygen isotope signals from newly assimilated sugars in the leaves to the tree-ring archive

The analysis of δ ¹³C and δ ¹⁸O in tree-ring archives offers retrospective insights into environmental conditions and ecophysiological processes. While photosynthetic carbon isotope discrimination and evaporative oxygen isotope enrichment are well understood, we lack information on how the isotope signal is altered by downstream metabolic processes.
In Pinus sylvestris , we traced the isotopic signals from their origin in the leaf water ( δ ¹⁸O) or the newly assimilated carbon ( δ ¹³C), via phloem sugars to the tree-ring, over a time-scale that ranges from hours to a growing season.
Seasonally, variable ¹³C enrichment of sugars related to phloem loading and transport did lead to uncoupling between δ ¹³C in the tree-ring, and the c _i/ c _a ratio at the leaf level. In contrast, the oxygen isotope signal was transferred from the leaf water to the tree-ring with an expected enrichment of 27‰, with time-lags of approximately 2 weeks and with a 40% exchange between organic oxygen and xylem water oxygen during cellulose synthesis.
This integrated overview of the fate of carbon and oxygen isotope signals within the model tree species P. sylvestris provides a novel physiological basis for the interpretation of δ ¹³C and δ ¹⁸O in tree-ring ecology.     

AUTORADIOGRAPHIC DETECTION OF DNA POLYMERASE CONTAINING NUCLEI IN SARCOMA 180 ASCITES CELLS

A technique has been developed allowing the autoradiographic detection of incorporation of ³H-thymidine-5'-triphosphate (³H-TTP) into nuclear DNA of smears of Sarcoma-180 (S-180) mouse ascites tumors under the direction of the cell's own nuclear DNA polymerase. Dried smears are dipped into an agar solution, which strips cytoplasm from the nuclei, and are then air dried and incubated with a buffered mixture containing four nucleotide triphosphates (one labeled), Mg⁺⁺, and Ficoll, with the cell's own DNA acting as primer. The incorporation of ³H-TTP into the nuclei, like the cell free DNA polymerase assay, is largely dependent on the presence of all four nucleotide triphosphates and Mg⁺⁺ and produces a product which is DNase sensitive and RNase resistant.
DNA polymerase activity, as studied in a cell free assay, decreases with tumor age. This correlates well with a decreasing ³H-TTP labeling index in autoradiographs of aging tumors. The ³H-TTP labeling index has also been shown to exceed but parallel the in vivo ³H-thymidine (³H-TdR) pulse labeling index for all tumor ages examined.
In at least some cell systems DNA polymerase seems characteristic of cells in cycle. The autoradiographic detection of nuclei containing the enzyme offers a new tool for the study of tumor cytokinetics.     

Photosynthesis and respiration of a diatom biofilm cultured in a new gradient growth chamber

Abstract A diatom biofilm was grown in a chamber developed for culture of biofilms in chemical gradients. The diatoms grew on a polycarbonate membrane filter which separated a sterile reservoir, with added phosphate, from a reservoir without phosphate. Within 3 weeks of inoculation, a thick biofilm developed on the surface of the filter. The biofilms were homogeneous and therefore suitable for calculations of O₂ diffusion fluxes from concentration profiles of O₂. Profiles of O₂, pH, and gross photosynthesis at different light intensities and liquid medium concentrations of dissolved inorganic carbon and O₂ were measured with microelectrodes. Respiratory activity in a layer of the biofilm was determined as the difference between gross photosynthesis and outflux of O₂ from that layer. The photosynthetic activity in a well-developed biofilm grown at 360 μEinst m⁻² s⁻¹ and 2.4 mM HCO₃⁻ was limited by the supply of inorganic carbon. Exposure to light above 360 μEinst m⁻² s⁻¹ stimulated gross photosynthesis as well as respiratory processes without affecting net outflux of O₂. Higher concentrations of inorganic carbon, on the other hand, enhanced gross photosynthesis without concurrent increase in respiratory rate, resulting in an increased outflux of O₂. High concentrations of O₂ in the liquid medium decreased the net outflux of O₂ with little effect on the gross photosynthesis. The effects of inorganic carbon and O₂ on the metabolic activities of the biofilm were consistent with the presence of photorespiratory activity.     

Metabolic Precursors and Compartmentation of Cerebral GABA in Vigabatrin-Treated Rats

Abstract: The metabolic precursors and cerebral compartmentation of the augmented GABA pool induced by vigabatrin, an irreversible inhibitor of GABA transaminase, have been investigated by ¹³C NMR. Adult rats receiving rat chow ad libitum were given either drinking water only or drinking water containing 2.5 g/L vigabatrin for 7 days. Both groups of animals were infused either with [1,2-¹³C₂]acetate (15 µmol/min/100 g body weight), an exclusive precursor of GABA formation through the glial glutamine pathway, or with [1,2-¹³C₂]glucose (15 µmol/min/100 g body weight), a substrate that can produce GABA through the glial glutamine pathway or by direct metabolism in the neurons. The brains were frozen in situ, extracted with perchloric acid, and analyzed by ¹³C NMR. In vigabatrin-treated animals [¹³C]glutamine, a common intermediate for [¹³C]GABA synthesis from glucose or acetate, was accumulated to similar amounts during infusions with [1,2-¹³C₂]glucose or [1,2-¹³C₂]acetate. However, [¹³C]GABA accumulation was sevenfold higher during [1,2-¹³C₂]glucose infusions or twofold higher during [1,2-¹³C₂]acetate infusions. These results show that the direct pathway of GABA formation by neuronal metabolism of glucose predominates over the alternative pathway through glial glutamine. Near-equilibrium relationships of the aminotransferases of GABA and aspartate imply that the observed [¹³C]GABA accumulation occurs initially in the neuronal compartment.     

A precise potentiometric method for determination of algal activity in an open CO2 system

Abstract. The activity of the green alga Scenedesmus obliquus was studied in simplified nutrient solutions (20 mol m₋₃ NaNO₃, 20 mol m₋₃ NH₄C1, 20 mol m₋₃ NH₄NO₃, and 20 mol m₋₃ NaCl, respectively) at 25 °C. The experiments were performed under welldefined incident photon density fluxes ranging from 10 to 200 μmol m₂ s₋₁, Light-dependent changes in pH and alkalinity (A) were followed by means of a potentiometric method using a glass electrode. In the experiments, carbon dioxide with known partial pressure was bubbled through the algal suspension, and during dark periods ul intervals of 1 h, the solution was allowed to equilibrate with the gas phase. This technique was applied to calculate equilibrium values of pH and alkalinity at regular intervals during a 12-h period. Results obtained in NaNO₃, solution show a linear increase in A with time, at each level of illumination studied. After an initial drop, A also increases in NH₄NO₃, solution in a similar way to that in NaNO₃ solution. The change in A with time was also found to increase linearly with the photon density flux studied and no saturation level could be defined. In experiments in NaCl solution, no changes in A were registered while measurements in NH₄Cl solution showed a decrease in A with time.     

Exogenous Glutamate Concentration Regulates the Metabolic Fate of Glutamate in Astrocytes

Abstract: The metabolic fate of glutamate in astrocytes has been controversial since several studies reported >80% of glutamate was metabolized to glutamine; however, other studies have shown that half of the glutamate was metabolized via the tricarboxylic acid (TCA) cycle and half converted to glutamine. Studies were initiated to determine the metabolic fate of increasing concentrations of [U-¹³C]glutamate in primary cultures of cerebral cortical astrocytes from rat brain. When astrocytes from rat brain were incubated with 0.1 m M [U-¹³C]glutamate 85% of the ¹³C metabolized was converted to glutamine. The formation of [1,2,3-¹³C₃]glutamate demonstrated metabolism of the labeled glutamate via the TCA cycle. When astrocytes were incubated with 0.2–0.5 m M glutamate, ¹³C from glutamate was also incorporated into intracellular aspartate and into lactate that was released into the media. The amount of [¹³C]lactate was essentially unchanged within the range of 0.2–0.5 m M glutamate, whereas the amount of [¹³C]aspartate continued to increase in parallel with the increase in glutamate concentration. The amount of glutamate metabolized via the TCA cycle progressively increased from 15.3 to 42.7% as the extracellular glutamate concentration increased from 0.1 to 0.5 m M , suggesting that the concentration of glutamate is a major factor determining the metabolic fate of glutamate in astrocytes. Previous studies using glutamate concentrations from 0.01 to 0.5 m M and astrocytes from both rat and mouse brain are consistent with these findings.     

Effects of temperature and fertilization on nitrogen cycling and community composition of an urban lawn

We examined the influence of temperature and management practices on the nitrogen (N) cycling of turfgrass, the largest irrigated crop in the United States. We measured nitrous oxide (N₂O) fluxes, and plant and soil N content and isotopic composition with a manipulative experiment of temperature and fertilizer application. Infrared lamps were used to increase surface temperature by 3.5±1.3 °C on average and control and heated plots were split into high and low fertilizer treatments. The N₂O fluxes increased following fertilizer application and were also directly related to soil moisture. There was a positive effect of warming on N₂O fluxes. Soils in the heated plots were enriched in nitrogen isotope ratio ( δ ¹⁵N) relative to control plots, consistent with greater gaseous losses of N. For all treatments, C₄ plant C/N ratio was negatively correlated with plant δ ¹⁵N, suggesting that low leaf N was associated with the use of isotopically depleted N sources such as mineralized organic matter. A significant and unexpected result was a large, rapid increase in the proportion of C₄ plants in the heated plots relative to control plots, as measured by the carbon isotope ratio ( δ ¹³C) of total harvested aboveground biomass. The C₄ plant biomass was dominated by crabgrass, a common weed in C₃ fescue lawns. Our results suggest that an increase in temperature caused by climate change as well as the urban heat island effect may result in increases in N₂O emissions from fertilized urban lawns. In addition, warming may exacerbate weed invasions, which may require more intensive management, e.g. herbicide application, to manage species composition.     

Evidence for the direct 2beta- and 3beta-hydroxylation of [2H2]GA20-13-O-[6'-2H2]glucoside in seedlings of Phaseolus coccineus

[17-²H₂]GA₂₀-13-O-[6'-²H₂]glucoside was synthesized and applied to seedlings of Phaseolus coccineus L. After incubation for 72 h the conjugate metabolites were purified and shown by LC-ESI-tandem-MS and GC-MS to be [17-²H₂]GA₁-13-O-[6'-²H₂]glucoside and [17-²H₂]GA₂₉-13-O-[6'-²H₂]glucoside. This is the first evidence for the conversion of intact GA-O-glucosides, and represents an additional metabolic pathway of the gibberellin metabolism in P. coccineus L. The results indicate that intact GA-O-glucosides are accepted by 2- and 3-oxidases in the plant.