GLT-1, GLAST, and EAAC1 are high-affinity, Na(+)-dependent glutamate transporters identified in rat forebrain. The expression of these transporter subtypes was characterized in three preparations: ...undifferentiated rat cortical astrocyte cultures, astrocytes cocultured with cortical neurons, and astrocyte cultures differentiated with dibutyryl cyclic AMP (dBcAMP). The undifferentiated astrocyte monocultures expressed only the GLAST subtype. Astrocytes cocultured with neurons developed a stellate morphology and expressed both GLAST and GLT-1; neurons expressed only the EAAC1 transporter, and rare microglia in these cultures expressed GLT-1. Treatment of astrocyte cultures with dBcAMP induced expression of GLT-1 and increased expression of GLAST. These effects of dBcAMP on transporter expression were qualitatively similar to those resulting from coculture with neurons, but immunocytochemistry showed the pattern of transporter expression to be more complex in the coculture preparations. Compared with astrocytes expressing only GLAST, the dBcAMP-treated cultures expressing both GLAST and GLT-1 showed an increase in glutamate uptake Vmax, but no change in the glutamate K(m) and no increased sensitivity to inhibition by dihydrokainate. Pyrrolidine-2,4-dicarboxylic acid and threo-beta-hydroxyaspartic acid caused relatively less inhibition of transport in cultures expressing both GLAST and GLT-1, suggesting a weaker effect at GLT-1 than at GLAST. These studies show that astrocyte expression of glutamate transporter subtypes is influenced by neurons, and that dBcAMP can partially mimic this influence. Manipulation of transporter expression in astrocyte cultures may permit identification of factors regulating the expression and function of GLAST and GLT-1 in their native cell type.
Glutamate uptake is coupled to counter-transport of K
+, and high external K
+ concentrations can induce reversal of glutamate uptake in whole-cell patch-clamp and isolated membrane preparations. ...However, high external K
+ causes little or no reversal of glutamate uptake in intact astrocytes, suggesting a regulatory mechanism not evident in membrane preparations. One mechanism by which intact cells could limit the effects of altered extracellular ion concentrations on glutamate transport is by compensatory changes in intracellular Na
+ concentrations. This possibility was examined using astrocyte cultures treated in two ways to reduce the driving force for glutamate uptake: incubation in high K
+ (with reciprocal reduction in Na
+), and incubation with metabolic inhibitors to induce ATP depletion. ATP depletion produced a rise in intracellular Na
+, a collapse of the membrane sodium gradient and a massive reversal of glutamate uptake. By contrast, incubation in high K
+/low Na
+ medium did not significantly alter the sodium gradient and did not induce glutamate uptake reversal. The sodium gradient was shown to be maintained under these conditions by compensatory reductions in intracellular Na
+ that approximately matched the reductions in extracellular Na
+.
These findings suggest a mechanism by which astrocytes may limit reversal of glutamate uptake under high K
+/low Na
+ conditions, and further suggest a general mechanism by which Na
+-dependent transport processes could be shielded from fluctuating extracellular ion concentrations.
Non-synaptic release may be the major route of excitatory amino acid (EAA) efflux during cerebral ischemia. Possible routes of non-synaptic release include non-specific anion channels, reversal of ...Na(+)-, Cl(-)-, or Ca(2+)-dependent uptake, and cell lysis. In the present study we employ a novel approach to show reversal of Na(+)-dependent uptake as a major route of EAA efflux from astrocyte cultures under conditions of energy failure. Primary rat astrocyte cultures were subjected to combined blockade of glycolytic and oxidative metabolism after incubation with 3H-D-aspartate (D-ASP). Energy failure produced an efflux of D-ASP that was maximal by 90 minutes. The efflux over this period was reduced by more than 50% in cells that had been pre-loaded with PDC (L-transpyrrolidine-2,4-dicarboxylic acid) or TBHA (threo-beta-hydroxyaspartic acid), compounds that are competitive inhibitors of Na(+)-dependent glutamate uptake. The effect of pre-loading with the inhibitors was concentration dependent. No effect was seen if the inhibitors were added after induction of energy failure, suggesting that the attenuation of D-ASP efflux resulted from binding of the inhibitors to an intracellular site. These results provide strong evidence that EAA efflux from astrocytes under conditions of energy failure occurs largely through reversal of Na(+)-dependent uptake.
: Elevated extracellular potassium concentration (K+e) has been shown to induce reversal of glial Na+‐dependent glutamate uptake in whole‐cell patch clamp preparations. It is uncertain, however, ...whether elevated K+e similarly induces a net glutamate efflux from intact cells with a physiological intracellular milieu. To answer this question, astrocyte cultures prepared from rat and mouse cortices were incubated in medium with elevated K+e (by equimolar substitution of K+ for Na+), and glutamate accumulation was measured by HPLC. With K+e elevations to 60 mM, medium glutamate concentrations did not increase during incubation periods of 5–120 min. By contrast, 45 min of combined inhibition of glycolytic and oxidative ATP production increased medium glutamate concentrations 50–100‐fold. Similar results were obtained in both rat and mouse cultures. Studies were also performed using astrocytes loaded with the nonmetabolized glutamate tracer d‐aspartate, and parallel results were obtained; no increase in medium d‐aspartate content resulted from K+e elevation up to 90 mM, whereas a large increase occurred during inhibition of energy metabolism. These results suggest that a net efflux of glutamate from intact astrocytes is not induced by any K+e attainable in brain.
Using 9.4 g of
96Zr isotope and 1221 days of data from the NEMO-3 detector corresponding to 0.031 kg y, the obtained
2
ν
β
β
decay half-life measurement is
T
1
/
2
2
ν
=
2.35
±
0.14
(
stat
)
±
0.16
...(
syst
)
×
10
19
yr
. Different characteristics of the final state electrons have been studied, such as the energy sum, individual electron energy, and angular distribution. The 2
ν nuclear matrix element is extracted using the measured
2
ν
β
β
half-life and is
M
2
ν
=
0.049
±
0.002
. Constraints on
0
ν
β
β
decay have also been set.
The NEMO 3 detector, which has been operating in the Fréjus underground laboratory since February 2003, is devoted to the search for neutrinoless double-beta decay (beta beta 0v). The half-lives of ...the two neutrino double-beta decay (beta beta 2v) have been measured for 100Mo and 82Se. After 389 effective days of data collection from February 2003 until September 2004 (phase I), no evidence for neutrinoless double-beta decay was found from approximately 7 kg of 100Mo and approximately 1 kg of 82Se. The corresponding limits are T1/2(beta beta0v) > 4.6 x 10(23) yr for 100Mo and T1/2(beta beta 0v) > 1.0 x 10(23) yr for 82Se (90% C.L.). Depending on the nuclear matrix element calculation, the limits for the effective Majorana neutrino mass are < 0.7-2.8 e/v for 100Mo and < 1.7-4.9 eV for 82Se.
Le couplage des méthodes de fractionnement granulométrique organo-minéral d'un sédiment et de l'extraction alcaline de ses composés humiques a permis d'étudier la répartition à la fois qualitative et ...quantitative du carbone dans les différentes fractions de la vase de l'étang des Noës (Yvelines) et de la rivière le Réveillon (Val de Marne). La comparaison des deux milieux sédimentaires, biologiquement et texturalement dissemblables, a mis en évidence que les processus d'humification et de minéralisation évoluent spatialement de façon différente, comme en témoigne la dégradation différenciée des matières organiques. Les sédiments de l'étang des Noës présentent une évolution de l'humification vers la profondeur, où l'on constate une forte production d'acides humiques, composés fortement polymérisés. En revanche, dans les sédiments du Réveillon, l'humification, prépondérante dès la strate superficielle, subit un blocage dans la strate profonde qui se traduit par une faible production de composés humiques. La méthodologie employée permet de constater une distribution des substances humiques et de formuler des hypothèses quant à l'évolution des sédiments étudiés.
The coupling of the methods of granulometric analysis of the organo-mineral textural fractions and the alkaline extraction of humic compounds was used to study the distribution, both qualitatively and quantitatively, of carbon in different mud fractions from the pond Noës (Yvelines) and the river Réveillon (Val de Marne). A comparison of the two sediment environments, biologically and texturally different, showed that the processes of humification and mineralisation evolved spatially in different ways, because of the different breakdown processes of the organic material. The pond sediments from Noës provide a change in humification with depth, where there is a strong production of humic acids that are chiefly polymers. On the other hand, in the sediments of Réveillon, humification is chiefly in the superficial strata and is blocked in the deeper strata with a low production of humic compounds. The methodology has elucidated the distribution of humic substances and has led to the development of hypotheses for changes in the sediments that were studied.
Technical design and performance of the NEMO 3 detector Arnold, R.; Augier, C.; Bakalyarov, A.M. ...
Nuclear instruments & methods in physics research. Section A, Accelerators, spectrometers, detectors and associated equipment,
2005, 2005-1-00, Letnik:
536, Številka:
1
Journal Article
Recenzirano
Odprti dostop
The development of the Neutrino Ettore Majorana Observatory (NEMO
∼
3
) detector, which is now running in the Fréjus Underground Laboratory (L.S.M. Laboratoire Souterrain de Modane), was begun more ...than ten years ago. The NEMO 3 detector uses a tracking-calorimeter technique in order to investigate double beta decay processes for several isotopes. The technical description of the detector is followed by the presentation of its performance.
Force and intracellular calcium signals were monitored in whole bullfrog semitendinosus muscles during fatigue produced by intermittent tetanic stimulation. Intracellular calcium signals were ...monitored using the fluorescent calcium-sensitive indicator indo-1 from the ratio of fluorescence intensities (R) at 400 and 470 nm. Fatiguing stimulation caused 1) proportional decreases of tetanic force and R, suggesting a component of the decreased force during fatigue of whole muscle may be due to insufficient calcium to activate contraction; 2) a progressive slowing of the relaxation of both force and R, suggesting slowed force relaxation may be mediated by slowed calcium removal from the myoplasm; 3) an increase of resting level R, suggesting impaired calcium removal from, or increased leakage to the cytosol; 4) prolongation of the twitch contraction, which was paralleled by changes in R. These findings are consistent with previous single fiber studies and suggest that changes in whole muscle contractility with fatigue may be partially mediated by changes in calcium handling by the cell.
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