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R Balázs

Publications and source records attributed to R Balázs.

At least 19 recordsLinked to original sources
PubMed2000

Expression of metabotropic glutamate receptor 5 is increased in astrocytes after kainate-induced epileptic seizures.

In the CNS there is a differential distribution of the metabotropic glutamate receptor 5 (mGluR5) in neurons and glia. Hippocampal nerve cells contain large amounts of the receptor transcript and protein that are expressed at very low levels in astrocytes. This is unexpected, as mGluR-induced phosphoinositide hydrolysis is substantial in cultured astrocytes and is mediated only by mGluR5 in these cells. In order to detect mGluR5 in astrocytes in vivo, we destroyed in a circumscribed part of the hippocampus nerve cells that have high level of receptor expression. Unilateral injection of kainate into the right amygdala produced epileptic seizures, as well as selective degeneration of nerve cells restricted to the ipsilateral CA3 and CA4 regions of the hippocampus, followed by the development of gliosis. In the affected fields only, mGluR5 immunoreactivity was severely reduced 3 days after kainate injection, followed by a progressive reappearance and lasting presence of the receptor protein. Receptor mRNA virtually disappeared from the pyramidal cell layer of the lesioned CA3/4 region. On the other hand, the message level increased persistently in the CA3 stratum lucidum and radiatum, the site of massive astrogliosis. In these sites, mGluR5 mRNA became detectable in double labeling studies in glial fibrillary acidic protein-positive astrocytes. We showed previously that growth factors induce a pronounced elevation of mGluR5 expression in cultured astrocytes (Miller et al. J Neurosci 15:-6109, 1995). The well-documented increase in the level of growth factors in the damaged brain may underlie the induction of the receptor expression in astrocytes in vivo, which may also be involved in repair processes in the injured nervous tissue.

Amygdala↗
PubMed1998

Receptor-coupled phospholipase C and adenylyl cyclase function with different calcium pools in astrocytes.

Astrocytes express phospholipase C (PLC)-coupled metabotropic glutamate receptors (only mGluR5 is detectable) and adenylyl cyclase (AC)-linked beta-adrenergic receptors. Calcium-sensitive effector enzymes are associated with these signal transduction pathways, but the relevant calcium compartments involved were found to be different. mGluR5-linked PLC responded primarily to extracellular Ca2+, suggesting a close spatial relation between the enzyme and Ca2+ entry channels. On the other hand, the calcium-inhibited AC associated with beta-adrenergic receptors was sensitive to intracellular Ca2+ selectively accessible to intracellular Ca2+ chelation. Furthermore, cAMP formation induced by direct activation of AC by forskolin was less responsive to intracellular Ca2+ chelation than that evoked by the receptor-activated AC, raising the possibility of selective access of the receptor to a pool of calcium-inhibited AC and/or the calcium modulation of some components of the coupling pathway.

Adenylyl Cyclases↗
PubMed1998

Calcium influx via ionotropic glutamate receptors causes long lasting inhibition of metabotropic glutamate receptor-coupled phosphoinositide hydrolysis.

Functional interaction between ionotropic and metabotropic glutamate receptors (iGluR and mGluR respectively) was studied in cerebellar granule cell cultures using quisqualate (QA), the most potent agonist of phosphoinositide hydrolysis coupled mGluR, and N-methyl-D-aspartate (NMDA) or kainate (KA) that activate different classes of iGluR. Two h exposure to NMDA or KA resulted in a marked reduction (about 75%) of QA-evoked PI hydrolysis. The efficacy of the two agonists was about the same, but the potencies were different (IC50 for NMDA about 35 microM and for KA about 70 microM). NMDA-induced depression of QA-stimulated PI hydrolysis was relatively long lasting but reversible. Recovery required protein synthesis. In nominally Ca2+-free medium both NMDA and KA failed to attenuate QA-stimulated PI hydrolysis. The effect of NMDA was prevented by the NMDA receptor antagonist MK801, but not by the wide spectrum protein kinase inhibitor staurosporin nor by the nitric oxide synthase inhibitor N omega-nitro-L-arginine. Cycloheximide and concanavalin A were also ineffective. The effect of KA was prevented by the selective non-NMDA receptor antagonist 2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo(F)quinoxaline (NBQX). Voltage sensitive Ca2+ channel antagonists together with MK801 did not counteract the inhibition by KA of the QA response. Both NMDA and KA attenuated PI hydrolysis evoked by the muscarinic receptor agonist carbachol (about 30%), indicating that the activation of iGluRs exerts a relatively general inhibitory effect on the function of different PLC-coupled metabotropic receptors. Consistent with this observation is that treatments either with KA and NMDA induced an inhibition (about 30%) of NaF-stimulated PI hydrolysis which occurs through the direct activation of G proteins. Our observations show that ionotropic glutamate receptor stimulation induces a long lasting suppression of QA-evoked PI breakdown through a Ca2+ dependent mechanism which seems to involve receptor coupled transduction systems downstream from mGluR. Such a Ca2+-dependent cross-talk involving ionotropic and metabotropic receptors may play a role in certain events of synaptic plasticity.

Animals↗
PubMed1998

Metabotropic glutamate receptor agonists potentiate cyclic AMP formation induced by forskolin or beta-adrenergic receptor activation in cerebral cortical astrocytes in culture.

The metabotropic glutamate receptor (mGluR) agonist 1-aminocyclopentane-1S,3R-dicarboxylic acid (ACPD) potentiated the accumulation of cyclic AMP induced by either beta-adrenergic receptor stimulation (isoproterenol) or direct activation of adenylyl cyclase (AC) with forskolin in rat cerebral cortical astrocytes grown in a defined medium. In contrast, ACPD inhibits the cyclic AMP response in astrocytes cultured in a serum-containing medium. Pharmacological characterization indicated that a group I mGluR, of which only mGluR5 is detectable in these cells, is involved in the potentiation of cyclic AMP accumulation. Potentiation was elicited by mGluR I agonists [e.g., (R,S)-3,5-dihydroxyphenylglycine (DHPG)], but not by mGluR II or III agonists; it was pertussis toxin resistant and abolished by procedures suppressing mGluR5 function (phorbol ester pretreatment or DHPG-induced receptor down-regulation). Nevertheless, it appears that products generated through the mGluR5 transduction pathway, such as elevated [Ca2+]i or activated protein kinase C (PKC), are not involved in the potentiation as it was not influenced by either the intracellular calcium chelator BAPTA-AM or the PKC inhibitor Ro 31-8220. An inhibitor of phospholipase C, U-73122, markedly attenuated mGluR5-activated phosphoinositide hydrolysis but did not significantly affect the DHPG potentiation of the cyclic AMP response. A mechanism is proposed in which the potentiating effect on AC could be mediated by free betagamma complex that is liberated after the agonist-bound mGluR5 interacts with its coupled G protein.

Adenylyl Cyclases↗
PubMed1997

Weaver mutant mouse cerebellar granule cells respond normally to chronic depolarization.

We studied the effects of chronic K(+)-induced membrane depolarization and treatment with N-methyl-D-aspartate (NMDA) on cerebellar granule cells (CGCs) from weaver mutant mice and non-weaver litter-mates. The weaver mutation is a Gly-to-Ser substitution in a conserved region of the Girk2 G protein-coupled inward rectifying potassium channel [Patil N., Cox D. R., Bhat D., Faham M., Myers R. M. and Peterson A. S. (1995) Nature Genet. 11, 126-129] which induces early death of CGCs. The biochemical differentiation of CGCs was estimated as the rate of 2-deoxy-D-glucose accumulation and the expression of neural cell adhesion molecule (NCAM). High (25 mM) K+ ion concentration or treatment with NMDA greatly promoted the biochemical differentiation of both weaver mutant and non-weaver litter-mate mouse CGCs. In contrast to the marked effect on biochemical differentiation in both weaver and non-weaver mice CGSs, chronic high K+ treatment only had limited effect on survival. The survival of weaver mutant mouse CGCs in medium containing 5 mM K+ ions was very low, only 20% of the plated cells surviving at 7 days after plating, as opposed to the 50% for non-weaver CGCs. Chronic high K+ treatment improved the relative survival of weaver mutant mouse CGCs 1.6 2.2-fold and that of non-weaver CGCs 1.2-1.4-fold; the same number of CGCs (about 20% of the plated cells) were rescued by high K+ in both types of culture. The findings indicate that, in culture weaver mutant mouse, CGCs have a normal response to membrane depolarization and that the normal function of the Girk2 potassium channel is not critical for the survival of differentiated CGCs.

Animals↗
PubMed1997

Metabotropic glutamate receptor mGluR5 in astrocytes: pharmacological properties and agonist regulation.

Metabotropic glutamate receptor (mGluR) agonists induce extensive phosphoinositide (PI) hydrolysis in astrocytes grown in a chemically defined medium with select growth factors. These astrocytes express mGluR5 transcripts, but none of the splice variants of mGluR1, thus permitting the characterization of mGluR5 in a native CNS cell without interference from mGluR1 activity. mGluR5 activation (1) was not associated with stimulation of cyclic AMP formation, (2) showed high sensitivity to the removal of extracellular versus intracellular Ca2+, (3) displayed high coupling efficiency relative to receptor density, and (4) induced PI hydrolysis that was suppressed by phorbol esters with low potency. The rank order of agonist potency was similar to that observed in mGluR1 and mGluR5 transfected cells. The phenylglycine antagonists tested were effective in blocking responses to 1-aminocyclopentane-1S,3R-dicarboxylic acid, but not to glutamate. Prolonged exposure to agonists induced a two-phase desensitization of mGluR5 function, an initial phase (completed by 1 h and plateaus for another 3 h) and a late phase (progressive decrease to approximately 30% of control levels by 24 h). Only the latter phase was associated with receptor down-regulation. Desensitization of mGluR5 function did not involve receptor internalization or phosphorylation mediated by protein kinase C or A; it was purely homologous, and reversible. Resensitization after short agonist treatment did not require prior receptor sequestration. Recovery after prolonged agonist exposure required new protein synthesis, but the restoration of function was more rapid than normalization of receptor protein levels, indicating that regulation also involves other components of the transduction system. The protracted desensitization of mGluR5 in astrocytes suggests that the functions mediated by this receptor are maintained under a variety of conditions ranging from repetitive stimulation to injury responses.

Animals↗
PubMed1996

Developmental changes in NMDA-induced cell swelling and its transition to necrosis measured with 1H magnetic resonance imaging, impedance and histology.

The vulnerability of the rat brain to intracerebrally injected N-methyl-D-aspartate (NMDA) drastically changes with age. We evaluated the developmental changes in the early and late responses to NMDA using 1H magnetic resonance imaging (MRI), cortical impedance and histology. NMDA, injected in the striatum of rats at postnatal days (P) 4, 7, 10, 14 and 21, induced a significant age-dependent reduction in the apparent diffusion coefficient (ADC) of tissue water in the striatum and the cerebral cortex monitored 1 h later using diffusion-weighted MRI. The reduction in ADC amounted 65% at P4 with lower values thereafter and was about 30% at P21. NMDA similarly induced a reduction in the cortical extracellular space (by 50% at P7 and 10% at P16) as measured with impedance recordings. The progressive decrease in the effect of NMDA with brain development was also indicated by a decrease in the volume of tissue in which the changes in ADC occurred (50 mm3 at P4 and 8 mm3 at P21). The diffusion of extracellular tracer molecules Mn2+ or [3H]-(R)-alpha-amino-3-hydroxy-5-methyl-4-isoxazole proprionic acid (AMPA) injected in the striatum and monitored with T1-weighted MRI and autoradiography respectively showed a similar age dependence with the diffusion volume being twofold larger in P7 than in P21 brain. Thus restriction in diffusion during brain development may contribute to the decrease in NMDA-induced injury with age. The volume of tissue necrosis and gliosis, measured with T2-weighted MRI and histology 5 days after NMDA injection, was similar to that outlined by the ADC reduction detected soon after the insult at P4, P7 and P21. However, at P10 and P14 only 50% of the tissue showing a hyperintense signal in DW images displayed necrosis and gliosis 5 days later. This study shows that during development the early response to NMDA in terms of cytotoxic cell swelling (indicated both with impedance recordings and diffusion-weighted MRI) decreases with age. In addition, with maturation only part of the brain tissue acutely affected by NMDA does proceed into necrosis and gliosis, indicating an increased capacity of cells in the developing rat brain to survive NMDA-induced cell swelling.

Age Factors↗
PubMed1996

Characterization of agonist-induced down-regulation of NMDA receptors in cerebellar granule cell cultures.

Exposure of cerebellar granule cells to NMDA in culture at 5 days in vitro, when cells are not yet vulnerable to NMDA, evoked a pronounced reduction in NMDA receptor activity, measured by NMDA-induced 45Ca2+ influx, and counteracted the normal developmental increase in NMDA receptors. The effect was concentration and time dependent, the half-maximal effect being reached at about 45 microM and by 4-5 h. The decrease in NMDA receptor function was accompanied by a significant reduction in the protein level of the obligatory NMDA receptor subunit (NR) NR1. Both parameters remained at a low level as long as the agonist was present. However, receptor down-regulation was reversible, as receptor protein levels and NMDA responses were restored to control values upon NMDA removal, this process requiring protein synthesis. NMDA treatment also elicited a decrease in NR1, NR2A, and NR2B subunit messenger RNA (mRNA) levels. However, in comparison with NMDA receptor proteins, the decrease was faster, and NMDA receptor mRNA content recovered to control levels within 24 h in spite of the presence of NMDA. Concerning the mechanisms of agonist-induced regulation of NMDA receptor expression, it seems that protein kinase C-mediated protein phosphorylation is not involved, whereas inhibition of Ca2+/calmodulin-dependent kinase II/IV by KN-62 does depress NMDA receptor expression even in the absence of NMDA.

Animals↗
PubMed1996

Attenuation of beta-amyloid neurotoxicity in vitro by potassium-induced depolarization.

The cell death of cultured neurons triggered by beta-amyloid peptides has been theorized to model, at least in part, the neurodegeneration associated with Alzheimer's disease. To investigate potential strategies to interrupt beta-amyloid neurotoxicity in vitro, we examined the effects of potassium-induced membrane depolarization, a treatment previously demonstrated to reduce development-related apoptosis in cultured neurons. We report here that cultured rat hippocampal neurons pretreated for several hours with 30 mM KCl exhibit significantly reduced vulnerability to aggregated beta-amyloid peptides. The potassium-mediated neuroprotection was mimicked by activation of voltage-sensitive calcium channels using S(-)-Bay K8644 and was attenuated by R(+)-Bay K 8644, a blocker of voltage-dependent calcium channels, and KN-82, an inhibitor of calcium/calmodulin-dependent protein kinase II. The protein synthesis inhibitor cycloheximide also attenuated beta-amyloid neurotoxicity. Addition of cycloheximide following 30 mM KCl significantly increased protection offered by membrane depolarization, whereas cycloheximide addition during membrane depolarization blocked the protective effect. These data suggest that one cellular pathway that can inhibit neuronal death induced by beta-amyloid involves calcium influx through voltage-sensitive channels followed by stimulation of calcium/calmodulin-dependent protein kinase activity and synthesis of new protein(s).

Alzheimer Disease↗
PubMed1995

Agonist-induced down-regulation of NMDA receptors in cerebellar granule cells in culture.

In contrast to the acute toxic effect of NMDA on mature cerebellar granule cells, chronic treatment with NMDA (140 microM from 1 to 9 days in vitro) did not compromise cell survival. Such treatment markedly suppressed NMDA receptor activity: at 8 days in vitro NMDA-induced 45Ca2+ influx was reduced by approximately 60% and acute exposure to NMDA (highest concentration tested, 1 mM) at 9 days in vitro did not cause detectable toxicity. The reduction in NMDA receptor activity was accompanied by a significant decrease (approximately 80% at 9 days in vitro) in the level of the NR1 and the NR2A NMDA receptor subunit protein, detected using the selective photoaffinity ligand [125I]CGP55802A. It seems, therefore, that the agonist-induced decrease in NMDA receptor activity is due to receptor down-regulation. In contrast to the marked influence of chronic NMDA exposure on the cellular content of the NMDA receptor subunit proteins, mRNA levels of the different subunits (NR1, NR2A, NR2B and NR2C) were not significantly affected. It seems, therefore, that agonist-induced down-regulation of the NMDA receptor involves critically mRNA translation and/or post-translational regulation.

Animals↗
PubMed1995

NMDA and kainate induce internucleosomal DNA cleavage associated with both apoptotic and necrotic cell death in the neonatal rat brain.

Injection of N-methyl-D-aspartate (NMDA) or kainate in the striatum of 7-day-old rats induced massive cell loss in the ipsilateral striatum, hippocampus and inner cortical layers. In order to examine whether apoptosis contributes to cell death in this model of excitotoxic injury we examined the progression of internucleosomal DNA fragmentation and changes in cellular ultrastructure. Agarose gel electrophoresis of DNA extracted from the ipsilateral striatum, cerebral cortex and hippocampus clearly showed breakdown of DNA into oligonucleosome-sized fragments, indicative of apoptosis, 12 h post-NMDA injection. In addition, an increase between 12 and 24 h was observed as well as a continuous presence 5 days later. Kainate induced a similar time course of oligonucleosomal DNA fragmentation, but the intensity of the ethidium bromide stained bands was less compared with that observed for NMDA. DNA fragmentation was not detected in animals intrastriatally injected with Tris-HCl or in animals treated with MK-801 [(+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohept-5,10-imine hydrogen maleate, 1 mg/kg] 30 min after NMDA injection. MK-801 had no effect on DNA fragmentation induced by kainate. In addition to agarose gel electrophoresis, terminal deoxynucleotidyltransferase-mediated dUTP-biotin nick end labelling (TUNEL) was used for detection of DNA fragmentation in sections. A gradual increase in the density of both apoptotic and non-apoptotic TUNEL nuclei was found in the anterior cingulate (ACC) and retrosplenial (RSC) areas of the cortex, the striatum, and the CA1 area and dentate gyrus of the hippocampus over the first 24 h post-NMDA or kainate injection. In the contralateral hemisphere hardly any TUNEL nuclei were present and their density was comparable with that in animals injected with vehicle only. In the ipsilateral mammillary nucleus (MN), which showed no signs of acute cell swelling after intrastriatal injection with NMDA, internucleosomal DNA fragmentation was found 24 and 48 h after intrastriatal NMDA injection. Here, the density of TUNEL cells with apoptotic morphology was high at 12 and 24 h post-NMDA injection but returned to control levels by 5 days. Electron microscopy showed cells with a clearly apoptotic morphology in the ACC and RSC and in the MN 24 h after NMDA injection. In the CA1 area of the hippocampus a necrotic, rather than an apoptotic, ultrastructure prevailed, indicating that the TUNEL method stained both apoptotic and necrotic cells.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1995

AMPA receptors in cerebellar granule cells during development in culture.

The survival and maturation of differentiating cerebellar granule cells in culture are known to be promoted by excitatory amino acids (EAAs) which, however, compromise the survival of mature cells. In contrast to the trophic effect, the toxic effect of alpha-amino-3-hydroxy-5-methyl-4-isoxasolepropiate (AMPA) could only be elicited when the desensitisation of AMPA receptors was blocked, cyclothiazide being used in this study. Nevertheless, even under these conditions, toxicity induced by AMPA in contrast to kainate was, at 9 DIV, only half of the maximal toxicity attained by 13-16 DIV. Since cellular responses to AMPA depend so dramatically on the maturational stage of granule cells, we examined here whether this characteristic is related to developmental changes in AMPA receptor properties, which may result from changes in the subunit composition of the receptor. In contrast to toxicity, AMPA-induced 45Ca2+ influx (determined in the presence of cyclothiazide and the NMDA receptor blocker MK-801) reached a maximum already at 9 DIV. This also applied to a fraction of the 45Ca2+ uptake which persisted either after Cd2+ application or under Na(+)-free conditions and therefore presumably was mediated directly through AMPA receptor channels. Quantitative analysis of Western blots showed that the amounts of GluR4 and to a lesser extent GluR2/3/4c are substantial already at 2 DIV, remaining fairly constant until 9 DIV, followed by an increase by 16 DIV. However GluR1, which is hardly detectable in granule cells in vivo and is also low early in vitro, increased almost linearly with cultivation time.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1995

Regulation of the expression of NMDA receptor subunits in rat cerebellar granule cells: effect of chronic K(+)-induced depolarization and NMDA exposure.

The influence of K(+)-induced membrane depolarization and NMDA treatment on the regulation of NMDA receptor subunit (NR) expression was investigated during the development of granule cells in culture, as a follow-up of previous work on NMDA receptor activity. In spite of the increase in NMDA receptor activity elicited by these treatments (K25 or K10 + NMDA cultures), the main developmental changes in receptor mRNA levels were similar to those in untreated cells (K10) (a threefold increase in total NMDA receptor mRNA, quantitative dominance of NR1 mRNA, late expression of NR2C, and virtual absence of NR2D. However, high K+ and NMDA treatment resulted in a greater increase of NR2A mRNA levels and a retardation in the developmental changes in the relative amounts of NR2B and NR2C mRNAs. The correspondence between NMDA receptor activity and the amount of NR1 and NR2A subunit proteins was excellent, the rank order being K25 > K10 + NMDA > K10 at 9 days in vitro. Because the increase in subunit mRNA was not always paralleled by an increase in subunit protein, the control of NMDA receptor expression involves critically, in addition to gene transcription, regulation of translational and/or posttranslational events.

Affinity Labels↗
PubMed1995

Properties of AMPA receptors expressed in rat cerebellar granule cell cultures: Ca2+ influx studies.

Cultured cerebellar granule cells become vulnerable to excitatory amino acids, especially to NMDA and kainate, by 9 days in vitro. In the same time, the sensitivity of cells to (RS)-alpha-amino-3-hydroxy-5-methyl-isoxazole-4-propionate (AMPA), in terms of AMPA-induced toxicity or 45Ca2+ uptake, was very low. The low AMPA responsiveness was due to receptor desensitization, because agents known to block desensitization, cyclothiazide and the lectins concanavalin A and wheat germ agglutinin, rendered granule cells vulnerable to AMPA and produced a pronounced stimulation of 45Ca2+ accumulation. 45Ca2+ influx was induced specifically by AMPA-receptor stimulation, because it was blocked virtually completely by 2,3-dihydroxy-6-nitro-7-sulfamoylbenzoquinoxaline (NBQX) and the benzodiazepine GYKI 52466 (selective non-NMDA receptor antagonists). Nevertheless, indirect routes activated by cellular responses to AMPA-receptor stimulation contributed significantly to the overall 45Ca2+ influx. These included Ca2+ uptake through NMDA-receptor channels, voltage-sensitive Ca2+ channels, and via Na+/Ca2+ exchange. However, nearly one-fifth of the total 45Ca2+ influx remained unaccounted for and this estimate was similar to 45Ca2+ influx observed under Na(+)-free conditions. This observation suggested that a significant proportion of the Ca2+ flux passes through the AMPA-receptor channel proper, a view supported by Co2+ uptake into nearly all granule cells on exposure to AMPA in the presence of cyclothiazide. Results are discussed in light of the reported AMPA receptor-subunit composition of cerebellar granule cells in vitro.

Animals↗
PubMed1995

Development of voltage-activated potassium currents in cultured cerebellar granule neurons under different growth conditions.

1. The functional expression of two potassium currents in cultured cerebellar granule cells was investigated with the whole cell patch-clamp technique in relation to development and growth condition. Cells were grown in medium containing different concentrations of potassium: 25 mM (K25) and 40 mM (K40), together referred to as "high K+"; 10 mM (K10) or "low K+"; and K10 with 100 microM N-methyl-D-aspartate (KNMDA). All conditions are known to influence maturation and survival of granule cells in culture. 2. At 2 days in vitro (DIV) the membrane capacitance, taken as index of membrane surface area, was the same for cells grown in each growth condition. At 7-9 DIV it had increased in each condition, but to a substantially larger extent in cells grown in KNMDA, K25, and K40 than in cells grown in K10. During development the input resistance only decreased in cells grown in KNMDA and high K+. 3. A delayed potassium current (IK) and a fast transient potassium current (IA) could both be recorded at 2 DIV in each growth condition, although a few neurons only expressed the IK. The IK was partially suppressed by tetraethylammonium (5 mM), whereas IA was predominantly sensitive to 4-aminopyridine (5 mM). 4. Normalized for cell capacitance, the specific IA conductance hardly changed during development in cells grown in high K+ and KNMDA. Cells in K10, however, displayed an IA with totally different properties in 23 of 24 cells; the specific IA conductance in these cells was considerably smaller at 7-9 DIV, suggesting a deletion of these channels during development.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1994

Growth conditions differentially modulate the vulnerability of developing cerebellar granule cells to excitatory amino acids.

The survival of immature nerve cells in a cerebellar culture, predominantly excitatory granule cells, is known to be promoted by chronic exposure to high K+ (> 20 mM) or glutamate (Glu) receptor agonists. These treatments are believed to mimic the in vivo effect of the incoming glutamatergic afferents, the mossy fibres. Here we report that with maturation the cells become vulnerable to excitatory amino acids (EAAs) and that the characteristics of EAA sensitivity are dependent on the environmental influences being either "trophic" (25 mM K+ or 140 microM NMDA, K25 or K10 + NMDA) or "non-trophic" (10 mM K+, K10). Toxicity was assayed routinely at 9 days in vitro (DIV) after 24 h exposure to EAAs. Under all the tested conditions, the effect of Glu was mediated exclusively through NMDA receptors. However, the efficacy and potency of Glu were high in K25- and K10 + NMDA-grown cells compared with K10-grown cells. Growth conditions had the same influence on NMDA as on Glu-induced toxicity, but with the following special features: (1) in comparison with K25 cells, the potency of NMDA was significantly lower in K10 + NMDA cells. The K10 + NMDA cultures behaved as if they were completely insensitive to the NMDA which is present in their growth medium. (2) The K10-grown cells were not vulnerable to NMDA, unless the cell membrane was depolarised by shifting the cells into K25 medium. The efficacy of NMDA became then similar to that in K25 cultures, although the potency was about 7-fold less. Thus NMDA receptors can be activated by the depolarisation of K10 cells, implying the operation of Mg2+ blockade of the channel at normal resting membrane potential. Although non-NMDA receptors did not seem to be involved in Glu toxicity, cells were vulnerable to kainate, which killed significantly more cells than Glu (about 80% vs 70%). This was partly due to the resistance of GABAergic interneurons present in the cultures to Glu- or NMDA-induced toxicity. In contrast to the effects of Glu or NMDA, KA vulnerability was lower in cells grown in K25 or K40 than K10 medium (rank order K10 > K25 > K40). Under our experimental conditions, cultured cells were resistant to AMPA, quisqualate and the selective metabotropic Glu receptor agonist 1S,3R-ACPD. Collectively, the observations indicated that EAA sensitivity of cultured cerebellar interneurons is significantly and differentially influenced by environmental factors, believed to mimic in vivo trophic influences on these cells.

Animals↗
PubMed1994

Comparison of diffusion-weighted MRI with changes in cell volume in a rat model of brain injury.

The precise mechanisms that underlie acute changes in tissue water diffusion following cerebral ischemia or related insults such as glutamate exposure remain unexplained, but it has been suggested that these may be caused by cell swelling due to water uptake. This study was undertaken to compare the changes observed in diffusion-weighted MR images with changes in the cellular volume measured by electrical impedance in a model of N-methyl-D-aspartate-induced brain injury in perinatal rats. The results show that the temporal course of the intensity changes in the diffusion-weighted images parallelled the progressive shrinkage of the extracellular space measured from the electrical impedance. After administration of the N-methyl-D-aspartate antagonist MK-801 the signal enhancement in the images was reversed, which paralleled the normalization of the extracellular space observed by the impedance measurements. It was estimated that the extracellular space decreased from 24 to 12% while the apparent diffusion coefficient of water decreased from 0.89 x 10(-9) in normal tissue to 0.42 x 10(-9) m2/s in tissue exposed to N-methyl-D-aspartate. These data indicate that changes in tissue water diffusion are related to changes in cell volume.

Animals↗