共查询到20条相似文献,搜索用时 16 毫秒
1.
Robert A. Waniewski 《Journal of neurochemistry》1992,58(1):167-174
The effect of ammonia on glutamate accumulation and metabolism was examined in astrocyte cultures prepared from neonatal rat cortices. Intact astrocytes were incubated with 70 microM L-[14C(U)]glutamate and varying amounts of ammonium chloride. The media and cells were analyzed separately by HPLC for amino acids and labelled metabolites. Extracellular glutamate was reduced to 8 microM by 60 min. Removal of glutamate from the extracellular space was not altered by addition of ammonia. The rate of glutamine synthesis was increased from 3.6 to 9.3 nmol/mg of protein/min by addition of 100 microM ammonia, and intracellular glutamate was reduced from 262 to 86 nmol/mg of protein after 30 min. The metabolism of accumulated glutamate was matched nearly perfectly by the synthesis of glutamine, and both processes were proportional to the amount of added ammonia. The transamination and deamination products of glutamate were minor metabolites that either decreased or remained unchanged with increasing ammonia. Thus, ammonia addition stimulates the conversion of glutamate to glutamine in intact astrocyte cultures. At physiological concentrations of ammonia, glutamine synthesis appears to be limited by the rate of glutamate accumulation and the activity of competing reactions and not by the activity of glutamine synthetase. 相似文献
2.
Activation of Glutamate Dehydrogenase by Leucine and Its Nonmetabolizable Analogue in Rat Brain Synaptosomes 总被引:1,自引:3,他引:1
Leucine and beta-(+/-)-2-aminobicyclo[2.2.1]heptane-2-carboxylic acid (BCH) stimulated, in a dose-dependent manner, reductive amination of 2-oxoglutarate in rat brain synaptosomes treated with Triton X-100. The concentration dependence curves were sigmoid, with 10-15-fold stimulations at 15 mM leucine (or BCH); oxidative deamination of glutamate also was enhanced, albeit less. In intact synaptosomes, leucine and BCH elevated oxygen uptake and increased ammonia formation, consistent with stimulation of glutamate dehydrogenase (GDH). Enhancement of oxidative deamination was seen with endogenous as well as exogenous glutamate and with glutamate generated inside synaptosomes from added glutamine. With endogenous glutamate, the stimulation of oxidative deamination was accompanied by a decrease in aspartate formation, which suggests a concomitant reduction in flux through aspartate aminotransferase. Activation of reductive amination of 2-oxoglutarate by BCH or leucine could not be demonstrated even in synaptosomes depleted of internal glutamate. It is suggested that GDH in synaptosomes functions in the direction of glutamate oxidation, and that leucine may act as an endogenous activator of GDH in brain in vivo. 相似文献
3.
Exogenous Glutamate Is Metabolized to Glutamine and Exported by Rat Primary Astrocyte Cultures 总被引:13,自引:13,他引:0
Rat cortical astrocytes in primary culture were examined for their capacity to transport and metabolize exogenous L-[U-14C]glutamate. After incubation for time periods up to 120 min, cells and incubation media were analyzed for labelled and endogenous glutamate and its metabolic products by HPLC coupled with fluorescence detection and liquid scintillation counting. Glutamine was the major labelled metabolite after 120 min, accounted for 38% of the original glutamate label, and was found primarily in the incubation medium. A further 13.5% of the label was recovered in deaminated metabolites of glutamate, 1.2% was associated with aspartate, 23% remained in glutamate, and 10.2% was found in an acid-precipitated cell fraction. More than 84% of the label was recovered in these fraction. suggesting that the maximum possible formation and loss of 14CO2 was 16%. The rate of total glutamine synthesis was 1.1 nmol X mg protein-1 X min-1 when 9 microM exogenous glutamate was present. The total amount of glutamine synthesized greatly exceeded the consumption of glutamate, indicating that a substantial proportion of glutamine was synthesized from other carbon sources. Almost all of the newly formed glutamine was exported into the medium. These results indicate that astrocytes in primary culture, by accumulating glutamate, producing glutamine, and exporting it, are capable of carrying out the glial component of the glutamine cycle. 相似文献
4.
Resveratrol Increases Glutamate Uptake, Glutathione Content, and S100B Secretion in Cortical Astrocyte Cultures 总被引:3,自引:0,他引:3
de Almeida LM Piñeiro CC Leite MC Brolese G Tramontina F Feoli AM Gottfried C Gonçalves CA 《Cellular and molecular neurobiology》2007,27(5):661-668
Resveratrol (3,5,4′-trihydroxy-trans-stilbene) is a polyphenol present in grapes and red wine, which has antioxidant properties and a wide range of other biological
effects. In this study, we investigated the effect of resveratrol, in a concentration range of 10–250 μM, on primary cortical
astrocytes; evaluating cell morphology, parameters of glutamate metabolism such as glutamate uptake, glutamine synthetase
activity and glutathione total content, and S100B secretion. Astrocyte cultures were prepared of cerebral cortex from neonate
Wistar rats. Morphology was evaluated by phase-contrast microscopy and immunocytochemistry for glial fibrillary acidic protein
(GFAP). Glutamate uptake was measured using l-[2,3-3H]glutamate. Glutamine synthetase and content of glutathione were measured by enzymatic colorimetric assays. S100B content
was determined by ELISA. Typical polygonal morphology becomes stellated when astrocyte cultures were exposed to 250 μM resveratrol
for 24 h. At concentration of 25 μM, resveratrol was able to increase glutamate uptake and glutathione content. Conversely,
at 250 μM, resveratrol decreased glutamate uptake. Unexpectedly, resveratrol at this high concentration increased glutamine
synthetase activity. Extracellular S100B increased from 50 μM upwards. Our findings reinforce the protective role of this
compound in some brain disorders, particularly those involving glutamate toxicity. However, the underlying mechanisms of these
changes are not clear at the moment and it is necessary caution with its administration because elevated levels of this compound
could contribute to aggravate these conditions. 相似文献
5.
Abstract: The present study sought to investigate the presence and distribution of some enzymatic activities involved in the metabolism of glutamate in the giant nerve fiber of the tropical squid Sepioteuthis sepioidea . Specific activities of aspartate aminotransferase and glutamate dehydrogenase were evaluated in homogenates of the isolated giant fiber, extruded axoplasm, and axoplasm-free giant nerve fiber sheaths. The activities of both enzymes were present in the tissue. The specific activity of aspartate aminotransferase was similar in axoplasm and sheaths. However, the specific activity of glutamate dehydrogenase was an order of magnitude higher in the sheaths. This finding is discussed in the framework of the hypothesis that proposes that a differential distribution of the enzymes of the glutamatergic system between the axonal and neuroglial compartments forms part of a system of communication between these cells whose neuronal signal may be glutamate. 相似文献
6.
Riluzole Enhances Glutamate Uptake in Rat Astrocyte Cultures 总被引:2,自引:0,他引:2
1. Riluzole is used for the treatment of amyotrophic lateral sclerosis and reported to have neuroprotective effects in animal models of Parkinson's disease, Huntington's disease, and brain ischemia. The neuroprotective action of riluzole has been attributed to its ability to inhibit glutamate release (A. Doble, Neurology 47(4):233S-241S, 1996). 2. The effect of riluzole on L-[2,3-3H] glutamate uptake was investigated in rat cortical astrocyte cultures. 3. Riluzole showed a biphasic concentration-dependent effect on basal glutamate uptake. At low concentrations (1 and 10 microM) riluzole significantly increased glutamate uptake, whereas from 100 microM promoted a slight reduction. 4. Considering the large range of glutamate levels in the synaptic cleft, we studied the 1 microM riluzole effect on uptake of glutamate at different concentrations (1-1000 microM). Riluzole was more effective at low glutamate concentrations (10 microM), enhancing the basal glutamate uptake up to 42%. 5. The action of riluzole on astrocytic glutamate uptake could be an additional mechanism to its neuroprotective role, perhaps suggesting a modulatory action on glutamatergic system involving glutamate clearance from synaptic cleft. 相似文献
7.
† D. Hantaz-Ambroise ‡D. Cambier A. Aït-Ikhlef §Ph. Parvy M. Murawsky F. Rieger 《Journal of neurochemistry》1995,65(3):1199-1204
Abstract: The wobbler mouse develops an inherited motoneuronal degeneration of unknown origin in the spinal cord. Primary cultures of adult wobbler spinal cord astrocytes display abnormal morphological characteristics with fewer processes and paucity of cell-cell contacts. We have searched for a possible involvement of glutamate and glutamine intra- and extracellular accumulations in vitro in the abnormal differentiation of mutant astrocytes. We have found significantly higher glutamate and glutamine concentrations in the culture media of mutant astrocytes over a 3-day period compared with normal control astrocytes. Moreover, intracellular glutamate concentrations decreased substantially in mutant astrocytes, but intracellular glutamine concentrations remained unchanged. Furthermore, decreasing initial glutamine concentrations in the culture medium (glutamine-depleted medium) led to the recovery of normal extra- and intracellular concentrations of glutamate and recovery of quasi-normal morphological differentiation and increased cell-cell contacts, leading to an essentially normal looking astrocyte network after 3 days of culture. Under these conditions, which lead to recovery, the only remaining abnormality was the higher glutamine extracellular concentration attained in the originally depleted glutamine media. These findings suggest that mechanisms regulating glutamate/glutamine synthesis and/or influx/efflux are defective in wobbler astrocytes, leading to metabolic imbalance and possible cytotoxic effects characterized by disturbed intercellular networks and poor differentiation. 相似文献
8.
Burbaeva GSh Boksha IS Tereshkina EB Savushkina OK Starodubtseva LI Turishcheva MS Mukaetova-Ladinska E 《Neurochemical research》2007,32(9):1434-1444
We have used a systemic approach to establish a relationship between enzyme measures of glial glutamate and energy metabolism
(glutamine synthetase and glutamine synthetase-like protein, glutamate dehydrogenase isoenzymes, brain isoform creatine phosphokinase)
and two major glial proteins (glial fibrillary acidic protein and myelin basic protein) in autopsied brain samples taken from
patients with schizophrenia (SCH) and mentally healthy subjects (23 and 22 cases, respectively). These biochemical parameters
were measured in tissue extracts in three brain areas (prefrontal cortex, caudate nucleus, and cerebellum). Significant differences
in the level of at least one of the glutamate metabolizing enzymes were observed between two studied groups in all studied
brain areas. Different patterns of correlative links between the biochemical parameters were found in healthy and schizophrenic
brains. These findings give a new perspective to our understanding of the impaired regulation of enzyme levels in the brain
in SCH. 相似文献
9.
Evidence that Aspartate Aminotransferase Activity and Ketodicarboxylate Carrier Function Are Essential for Biosynthesis of Transmitter Glutamate 总被引:1,自引:5,他引:1
Based on the selective inhibition of glutamate release in cerebellar granule cells in primary cultures by the aspartate aminotransferase inhibitor, aminooxyacetic acid, and by the ketodicarboxylate carrier inhibitor, phenylsuccinate, a novel model for synthesis of transmitter glutamate is suggested: Glutamate is formed from glutamine in the mitochondrial intramembrane space by phosphate-activated glutaminase, transported across the inner membrane in exchange with aspartate, transaminated in the matrix to alpha-ketoglutarate, which via the ketodicarboxylate carrier is transferred to the cytoplasm, and transaminated to form transmitter glutamate. Such a mechanism would explain the functional role of aspartate aminotransferase in glutamatergic neurons. 相似文献
10.
Protective Effects of Resveratrol on Hydrogen Peroxide Induced Toxicity in Primary Cortical Astrocyte Cultures 总被引:1,自引:0,他引:1
Vieira de Almeida LM Piñeiro CC Leite MC Brolese G Leal RB Gottfried C Gonçalves CA 《Neurochemical research》2008,33(1):8-15
It is well established that the brain is particularly susceptible to oxidative damage due to its high consumption of oxygen
and that astrocytes are involved in a variety of important activities for the nervous system, including a protective role
against damage induced by reactive oxygen species (ROS). The use of antioxidant compounds, such as polyphenol resveratrol
found in red wine, to improve endogenous antioxidant defenses has been proposed for neural protection. The aim of this study
is to evaluate the putative protective effect of resveratrol against acute H2O2-induced oxidative stress in astrocyte cultures, evaluating ROS production, glutamate uptake activity, glutathione content
and S100B secretion. Our results confirm the ability of resveratrol to counteract oxidative damage caused by H2O2, not only by its antioxidant properties, but also through the modulation of important glial functions, particularly improving
glutamate uptake activity, increasing glutathione content and stimulating S100B secretion, which all contribute to the functional
recovery after brain injury. 相似文献
11.
Jon Henrik Laake Tove Anita Slyngstad Finn-Mogens mejda Haug Ole Petter Ottersen 《Journal of neurochemistry》1995,65(2):871-881
Abstract: The immunogold labeling for glutamate and glutamine was studied at the electron microscopic level in hippocampal slice cultures following inhibition of l -glutamine synthetase [ l -glutamate:ammonia ligase (ADP-forming); EC 6.3.1.2]. In control cultures, glutamate-like immunoreactivity was highest in terminals, intermediate in pyramidal cell bodies, and low in glial cells. Glutamine-like immunoreactivity was high in glial cells, intermediate in pyramidal cell bodies, and low in terminals. After inhibition of glutamine synthetase with l -methionine sulfoximine, glutamate-like immunoreactivity was reduced by 52% in terminals and increased nearly fourfold in glia. Glutamine-like immunoreactivity was reduced by 66% in glia following l -methionine sulfoximine, but changed little in other compartments. In cultures that were treated with both l -methionine sulfoximine and glutamine (1.0 m M ), glutamate-like immunoreactivity was maintained at control levels in terminals, whereas in glia glutamate-like immunoreactivity was increased and glutamine-like immunoreactivity was decreased to a similar extent as in cultures treated with l -methionine sulfoximine alone. We conclude that (a) glutamate accumulates in glia when the flux through glutamine synthetase is blocked, emphasizing the importance of this pathway for the handling of glutamate; and (b) glutamine is necessary for the maintenance of a normal level of glutamate in terminals, and neither reuptake nor de novo synthesis through pathways other than the glutaminase reaction is sufficient. 相似文献
12.
Astrocyte and Neuron Intone Through Glutamate 总被引:1,自引:0,他引:1
The unexpected finding of astrocytes to release glutamate as gliotransmitter challenges the traditional concepts on astrocyte
being “passive” in CNS communications. Glutamate is the major excitatory transmitter in transferring information between neurons,
but is now also known to activate astrocyte through transporters and receptors. Together with the sensitive swelling response,
astrocytes could respond directly to glutamate and neuronal activity. Other new functions of astrocytes include modulation
of synaptic plasticity and cerebral blood flow (CBF). The classic glutamate deplenishment through glutamine synthesis and
CO2 production does not account for the total glutamate internalized into astrocytes. This leads us to speculate there are many
hidden functions of glutamate in neurons and astrocytes waiting to be discovered. In this review, we attempted to reexamine
some of these new and older functions of glutamate and to reevaluate the roles of glutamate intoning these two cell types.
Special issue article in honor of Dr. Anna Maria Giuffrida-Stella. 相似文献
13.
The metabolic fate of L-[U-14C]- and L-[1-14C]glutamate was studied in primary cultures of mouse astrocytes. Conversion of the uniformly labeled compound to glutamine and aspartate was followed by determination of specific activities after dansylation with [3H]dansyl chloride and subsequent thin layer chromatography of the dansylated amino acids. Metabolic fluxes were calculated from the alterations of specific activities and the pool sizes, which were likewise measured by a dansylation method. Formation of 14CO2 from [1-14C]glutamate was determined by the trapping of CO2 in hyamine hydroxide in a gas-tight chamber, which is, in the known absence of glutamate decarboxylase activity in the cultured astrocytes, an unequivocal expression of the metabolic flux via alpha-ketoglutarate to CO2 and succinyl-CoA. The metabolic fluxes determined by these procedures amounted to 2.4 nmol/min/mg protein for glutamine synthesis, 1.1 nmol/min/mg protein for aspartate production, and 4.1 nmol/min/mg protein for formation and subsequent decarboxylation of alpha-ketoglutarate. The latter process was unaffected by virtually complete inhibition of glutamate-oxaloacetic transaminase with aminooxyacetic acid, indicating that the formation of alpha-ketoglutarate occurs as an oxidative deamination rather than as a transamination. This suggests that the formation of alpha-ketoglutarate from glutamate represents a net degradation, not an isotopic exchange. 相似文献
14.
The effect of the glutamine synthetase (GS) inhibitor, methionine sulfoximine (MSO), on glutamate levels in, and glutamate release from, rat striatal tissue was examined. Tissue levels of glutamate were unchanged 24 h after an intraventricular injection of MSO, but tissue glutamine levels were decreased 50%. Calcium-dependent, potassium-stimulated glutamate release was diminished in tissue prisms from animals pretreated with MSO compared to controls. The decreased release of glutamate correlated over time with the inhibition of GS following an intraventricular injection of MSO. The maximum diminution of calcium-dependent, potassium-stimulated glutamate release (50%) and the maximum inhibition of GS activity (51%) were observed 24 h after MSO. The addition of 0.5 mM glutamine to the perfusion medium completely reversed the effects of MSO pretreatment on calcium-dependent, potassium-stimulated glutamate release. Since GS is localized in glial cells and the measured glutamate release is presumed to occur from neurons, the data support the contention that astroglial glutamine synthesis is an important contributor to normal neuronal neurotransmitter release. 相似文献
15.
Triptolide (T10), an extract from the traditional Chinese herb, Tripterygium wilfordii Hook F (TWHF), has been shown to attenuate
the rotational behavior induced by d-amphetamine and prevent the loss of dopaminergic neurons in the substantia nigra in rat models of Parkinson’s disease. To
examine if the neuroprotective effect is mediated by its stimulation of production of neurotrophic factors from astrocytes,
we investigated the effect of T10 on synthesis and release of nerve growth factor (NGF), brain-derived neurotrophic factor
(BDNF) and glial cell line-derived neurotrophic factor (GDNF) in rat astrocyte cultures. T10 did not affect the synthesis
and release of either BDNF or GDNF. However, it significantly increased NGF mRNA expression. It also increased both intracellular
NGF and NGF level in culture medium. These results indicate that the neuroprotective effect of T10 might be mediated, at least
in part, via a stimulation of the production and release of NGF in astrocytes.
Authors Bing Xue and Jian Jiao contributed equally to this work. 相似文献
16.
Aspartate Aminotransferase for Synthesis of Transmitter Glutamate in the Medulla Oblongata: Effect of Aminooxyacetic Acid and 2-Oxoglutarate 总被引:1,自引:1,他引:0
The effects of aminooxyacetic acid (AOAA), a transaminase inhibitor, and 2-oxoglutarate, a precursor to glutamate by the activity of aspartate aminotransferase (AAT), on slices of rat medulla oblongata, cerebellum, cerebral cortex, and hippocampus were studied. The slices were superfused and electrically stimulated. There was a Ca2+-dependent stimulus-evoked release of endogenous glutamate, gamma-aminobutyric acid (GABA), and beta-alanine in all regions examined. AOAA (10(-4) and 10(-3) M) decreased the release of glutamate in the medulla oblongata and cerebellum but not in the hippocampus. L-Canaline, a specific inhibitor of ornithine aminotransferase, did not affect the glutamate release in the medulla. 2-Oxoglutarate (10(-3) M) increased the release of glutamate in the medulla oblongata and cerebellum but not in the cerebral cortex and hippocampus. Treatment with AOAA (10(-4) M) almost abolished the activities of AAT in all regions studied. AOAA (10(-4) and 10(-3) M) increased the stimulus-evoked release of GABA in the cerebellum, cerebral cortex, and hippocampus, whereas the stimulus-evoked release of beta-alanine was decreased by this agent in all regions studied. These results suggest the participation of AAT in the synthesis of the transmitter glutamate in the medulla oblongata and cerebellum of the rat. 相似文献
17.
Aspartate levels and release from rat striatal slices following the inhibition of glutamine synthetase (GS) by methionine sulfoximine (MSO) were studied. Striatal levels of aspartate and glutamine were decreased over time in a manner that correlated with GS inhibition. Ca2+-dependent, K+-stimulated aspartate release was diminished in striatal tissue slices from animals pretreated with MSO. The decreased release of aspartate correlated over time with the inhibition of GS. The addition of glutamine to the perfusion medium completely reversed the effects of MSO on calcium-dependent aspartate release. It is suggested that glutamine is a major precursor for transmitter aspartate. 相似文献
18.
Cellular Location of Glutamine Synthetase and Lactate Dehydrogenase in Oligodendrocyte-Enriched Cultures from Rat Brain 总被引:3,自引:3,他引:0
Ruud A. J. Warringa Mario F. van Berlo Wil Klein Matthijs Lopes-Cardozo 《Journal of neurochemistry》1988,50(5):1461-1468
Glial cells were isolated from 1-week-old rat brain and cultured in a serum-free medium supplemented with the hormones insulin, hydrocortisone, and triiodothyronine. After 1 week in culture the cell population consisted mainly of galactocerebroside-positive cells (GC+; oligodendrocytes), the remainder of the cells being positive for glial fibrillary acidic protein (GFAP+; astrocytes). Oligodendrocytes were selectively removed from the cultures by complement-mediated cytolysis. The activities of glutamine synthetase and of various marker enzymes were measured in the nonlysed cells remaining after complement treatment of the cultures and in the culture medium containing proteins of the lysed cells. We found that the cellular activity of glutamine synthetase decreased in parallel with the lysis of GC+ cells and that the activity of glutamine synthetase in the supernatant increased. The activity of glycerol-3-phosphate dehydrogenase, a marker enzyme for oligodendrocytes, was no longer detectable in complement-treated cultures and the activity of glutamine synthetase was markedly lowered, whereas the activity of lactate dehydrogenase was as high as in untreated cultures. The location of glutamine synthetase both in oligodendrocytes and in astrocytes was confirmed by double-label immunocytochemistry with antisera against glutamine synthetase, GC, and GFAP. We conclude that in this culture system glutamine synthetase is expressed in both types of glial cells and that the activity of lactate dehydrogenase is at least one order of magnitude higher in astrocytes than in oligodendrocytes. 相似文献
19.
Wang Pengpeng 《Animal biotechnology》2013,24(2):107-128
In the rumen bacteria, ammonia as the end product of nitrogen is incorporated into carbon skeleton (α-ketoglutarate) to yield glutamine and glutamate which are important nitrogen donors in nitrogenous compounds metabolism in cells. The enzymes glutamine synthetase, glutamate synthetase, and glutamate dehydrogenase are involved in these processes. Some experimental results have proven that the global nitrogen regulation system may participate in the regulation of assimilation of ammonia in rumen bacteria. This review offers a current perspective on the pathways and key enzymes of ammonia assimilation in rumen bacteria with the possible molecular regulation strategy, while points out the further research direction. 相似文献
20.
A survey of the possible nitrogen assimilation pathways in Hyphomicrobium X showed that when the nitrogen source was satisfied by ammonium sulphate or methylamine and the supply was in excess, NADPH-dependent glutamate dehydrogenase was used to assimilate nitrogen. When the nitrogen supply was limited the cells expressed high levels of glutamine synthetase and NADH-dependent glutamine:2-oxoglutamate aminotransferase activity whilst the activity of the glutamate dehydrogenase was lower. When nitrate was the N-source, the glutamine synthetase/glutamine oxoglutamate aminotransferase pathway was utilised irrespective of the nitrogen concentration in the medium. Evidence was obtained to suggest that the glutamine synthetase activity was regulated by adenylylation/deadenylylation. Carbon-limited chemostat cultures showed low glutamine synthetase activity levels but the synthesis of the enzyme was derepressed when the cultures became N-limited. 相似文献