Biomedical subjects

Carla I Tasca

Publications and source records attributed to Carla I Tasca.

9 recordsLinked to original sources

PubMed2006

Oxygen-glucose deprivation decreases glutathione levels and glutamate uptake in rat hippocampal slices.

Ischemia is a transitory or permanent reduction of blood flow that may provoke an excessive release of glutamate. In that condition, increased reactive oxygen species generation and/or decreased cerebral antioxidant capacity may induce cell death. Antioxidant enzymes and thiols play an important role in the cellular defenses against oxidative stress. The purpose of this study was to evaluate cell viability, glutamate uptake and antioxidant status in rat hippocampal slices exposed to oxygen-glucose deprivation (OGD), an in vitro model of ischemia. After 15 min or 1 h of OGD, hippocampal slices showed a significant reduction of cell viability. Reperfusion during 1 or 2 h did not increase cell death. In this condition, the activities of antioxidant enzymes catalase, glutathione reductase, and peroxidase did not change. However, slices exposed to 15 min OGD and reperfused for 1 or 2 h showed higher superoxide dismutase activity. A significant reduction of glutathione levels was observed after 1 or 2 h of reperfusion in slices previously exposed to 1 h of OGD, although the protein-thiol content was unchanged. Slices exposed to 1 h of OGD and reperfused for 2 h showed reduced sodium-dependent l-[(3)H]glutamate uptake. The reduction of glutamate uptake was partially reversed by dl-dithiothreitol (DTT), a thiol-reducing agent, which may reduce thiol groups in glutamate transporters. Therefore, higher glutamate levels in the synaptic cleft could promote transporter reversal and impair glutamate uptake. Increased extracellular glutamate levels associated with decreased glutathione levels might exacerbate cell damage induced by oxygen and glucose deprivation.

Amino Acid Transport System X-AG↗

PubMed2006

Synaptosomal glutamate release and uptake in mice lacking the cellular prion protein.

Glutamate plays a central role in the fast excitatory synaptic transmission and is a key neurotransmitter involved in several neurophysiological processes. Glutamate levels on the synaptic cleft are related to neural excitability, neuroplasticity, and neuronal damage associated with excitotoxicity. Mice lacking the cellular prion protein (PrP(c)) gene (Prnp) present a decreased astrocytic glutamate uptake in cultures, higher neuronal excitability in vitro and sensitivity to pro-convulsant drugs in vivo, and age-dependent memory impairment. Here, we investigate if PrP(c) might be involved in neuronal uptake and release of glutamate. For this purpose, we compared synaptosomal preparations from the cerebral cortex, entorhinal cortex, hippocampus, cerebellum, and olfactory bulb of 3- or 9-month-old PrP(c) null mice and with respective wild-type controls. Although we observed differences in synaptosomal glutamate release and uptake regarding the age of mice and the brain structure studied, these differences were similar for PrP(c) null mice and their respective wild-type controls. Therefore, despite a possible correlation between neuronal glutamate transporters, excitability, and neuronal damage, our results suggest that PrP(c) expression is not critical for neuronal glutamate transport.

Animals↗

PubMed2006

GTP uptake into rat brain synaptic vesicles.

Uptake of neurotransmitters into synaptic vesicles is driven by an electrochemical gradient generated by a vacuolar-type proton pump ATPase. This uptake implies a key role for synaptic vesicles in the regulation of neurotransmitter systems. Guanine nucleoside and nucleotides are involved in the inhibition of glutamate-induced cellular responses via an extracellular action and diverse trophic, proliferative, and modulatory effects of guanine nucleotides on neural cells have been shown. Here, we characterized the uptake of GTP into synaptic vesicles isolated from whole rat brain, by using a tritiated poorly-hydrolyzable GTP analog, 5'-guanylylimidodiphosphate ([3H]GppNHp). Uptake of GTP into synaptic vesicles is saturable, time- and temperature-dependent, and relies on a proton-eletrochemical gradient. However, [3H]GMP and [3H]GDP radioactive labeling in synaptic vesicles is not dependent on temperature and vesicular ATPase activity, which indicates that these nucleotides only bind to and are not taken up into synaptic vesicles. GTP is taken up by the same eletrochemical gradient-dependent transport system, as are neurotransmitters storage, which indicates that this guanine nucleotide may also function as a neurotransmitter.

Adenosine Triphosphate↗

PubMed2005

Impaired exercise capacity, but unaltered mitochondrial respiration in skeletal or cardiac muscle of mice lacking cellular prion protein.

The studies of physiological roles for cellular prion protein (PrP(c)) have focused on possible functions of this protein in the CNS, where it is largely expressed. However, the observation that PrP(c) is expressed also in muscle tissue suggests that the physiological role of PrP(c) might not be limited to the central nervous system. In the present study, we investigated possible functions of PrP(c) in muscle using PrP(c) gene (Prnp) null mice (Prnp(0/0)). For this purpose, we submitted Prnp(0/0) animals to different protocols of exercise, and compared their performance to that of their respective wild-type controls. Prnp(0/0) mice showed an exercise-dependent impairment of locomotor activity. In searching for possible mechanisms associated with the impairment observed, we evaluated mitochondrial respiration (MR) in skeletal or cardiac muscle from these mice during resting or after different intensities of exercise. Baseline MR (states 3 and 4), respiratory control ratio (RCR) and mitochondrial membrane potential (DeltaPsi) were evaluated and were not different in skeletal or cardiac muscle tissue of Prnp(0/0) mice when compared with wild-type animals. We concluded that Prnp(0/0) mice show impairment of swimming capacity, perhaps reflecting impairment of muscular activity under more extreme exercise conditions. In spite of the mitochondrial abnormalities reported in Prnp(0/0) mice, our observation seems not to be related to MR. Our results indicate that further investigations should be conducted in order to improve our knowledge about the function of PrP(c) in muscle physiology and its possible role in several different neuromuscular pathologies.

Animals↗

PubMed2005

Neurotoxicity induced by glutamate in glucose-deprived rat hippocampal slices is prevented by GMP.

Guanosine-5'-monophosphate (GMP) was evaluated as a neuroprotective agent against the damage induced by glutamate in rat hippocampal slices submitted to glucose deprivation. In slices maintained under physiological conditions, glutamate (0.01 to 10 mM), Kainate, alpha-amino-3-hydroxi-5-methylisoxazole-propionic acid (AMPA), N-methyl-D-aspartate (NMDA), 1S,3R-aminocyclopentane-1,3-dicarboxylic acid (1S,3R-ACPD), or L-2-amino-4-phosphonobutanoic acid (L-AP4) (100 microM) did not alter cell membrane permeability, as evaluated by lactate dehydrogenase (LDH) release assay. In slices submitted to glucose deprivation, GMP (from 0.5 mM) prevented LDH leakage and the loss of cell viability induced by 10 mM glutamate. LDH leakage induced by Kainate, AMPA, NMDA or 1S,3R-ACPD was fully prevented by 1 mM GMP. However, glutamate uptake was not altered in slices submitted to glucose deprivation and glutamate analogues. Glucose deprivation induced a significant decrease in ATP levels which was unchanged by addition of glutamate or GMP. Our results show that glucose deprivation decreases the energetic charge of cells, making hippocampal slices more susceptible to excitotoxicity and point to GMP as a neuroprotective agent acting as a glutamatergic antagonist.

Animals↗

PubMed2005

In vivo quinolinic acid increases synaptosomal glutamate release in rats: reversal by guanosine.

Glutamate, the main excitatory neurotransmitter in the mammalian central nervous system (CNS), plays important role in brain physiological and pathological events. Quinolinic acid (QA) is a glutamatergic agent that induces seizures and is involved in the etiology of epilepsy. Guanine-based purines (GBPs) (guanosine and GMP) have been shown to exert neuroprotective effects against glutamatergic excitotoxic events. In this study, the influence of QA and GBPs on synaptosomal glutamate release and uptake in rats was investigated. We had previously demonstrated that QA "in vitro" stimulates synaptosomal L-[3H]glutamate release. In this work, we show that i.c.v. QA administration induced seizures in rats and was able to stimulate synaptosomal L-[3H]glutamate release. This in vivo neurochemical effect was prevented by i.p. guanosine only when this nucleoside prevented QA-induced seizures. I.c.v. QA did not affect synaptosomal L-[3H]glutamate uptake. These data provided new evidence on the role of QA and GBPs on glutamatergic system in rat brain.

Animals↗

PubMed2004

Guanine derivatives modulate L-glutamate uptake into rat brain synaptic vesicles.

Glutamate uptake into synaptic vesicles is driven by a proton electrochemical gradient generated by a vacuolar H(+)-ATPase and stimulated by physiological concentrations of chloride. This uptake plays an important role in glutamatergic transmission. We show here that vesicular glutamate uptake is selectively inhibited by guanine derivatives, in a time- and concentration-dependent manner. Guanosine, GMP, GDP, guanosine-5'-O-2-thiodiphosphate, GTP, or 5'-guanylylimidodiphosphate (GppNHp) inhibited glutamate uptake in 1.5 and 3 min incubations, however, when incubating for 10 min, only GTP or GppNHp displayed such inhibition. By increasing ATP concentrations, the inhibitory effect of GTP was no longer observed, but GppNHp still inhibited glutamate uptake. In the absence of ATP, vesicular ATPase can hydrolyze GTP in order to drive glutamate uptake. However, 5mM GppNHp inhibited ATP hydrolysis by synaptic vesicle preparations. GTP or GppNHp decreased the proton electrochemical gradient, whereas the other guanine derivatives did not. Glutamate saturation curves were assayed in order to evaluate the specificity of inhibition of the vesicular glutamate carrier by the guanine derivatives. The maximum velocity of the initial rate of glutamate uptake was decreased by all guanine derivatives. These results indicate that, although GppNHp can inhibit ATPase activity, guanine derivatives are more likely to be acting through interaction with vesicular glutamate carrier.

Adenosine Triphosphatases↗

PubMed2002

Quinolinic acid stimulates synaptosomal glutamate release and inhibits glutamate uptake into astrocytes.

Quinolinic acid (QA) is an endogenous neurotoxin involved in various neurological diseases, whose action seems to be exerted via glutamatergic receptors. However, the exact mechanism responsible for the neurotoxicity of QA is far from being understood. We have previously reported that QA inhibits vesicular glutamate uptake. In this work, investigating the effects of QA on the glutamatergic system from rat brain, we have demonstrated that QA (from 0.1 to 10mM) had no effect on synaptosomal L-[3H]glutamate uptake. The effect of QA on glutamate release in basal (physiological K+ concentration) or depolarized (40 mM KCl) conditions was evaluated. QA did not alter K+-stimulated glutamate release, but 5 and 10mM QA significantly increased basal glutamate release. The effect of dizolcipine (MK-801), a noncompetitive antagonist of N-methyl-D-aspartate (NMDA) receptor on glutamate release was investigated. MK-801 (5 microM) did not alter glutamate release per se, but completely abolished the QA-induced glutamate release. NMDA (50 microM) also stimulated glutamate release, without altering QA-induced glutamate release, suggesting that QA effects were exerted via NMDA receptors. QA (5 and 10mM) decreased glutamate uptake into astrocyte cell cultures. Enhanced synaptosomal glutamate release, associated with inhibition of glutamate uptake into astrocytes induced by QA could contribute to increase extracellular glutamate concentrations which ultimately lead to overstimulation of the glutamatergic system. These data provide additional evidence that neurotoxicity of QA may be also related to disturbances on the glutamatergic transport system, which could result in the neurological manifestations observed when this organic acid accumulates in the brain.

Animals↗

PubMed2002

Neuroprotective effect of GMP in hippocampal slices submitted to an in vitro model of ischemia.

1. Guanosine-5'-monophosphate (GMP) was evaluated as a neuroprotective agent against the damage observed in rat hippocampal slices submitted to an in vitro model of ischemia with or without the presence of the ionotropic glutamate receptor agonist, Kainic acid (KA). 2. Cellular injury was evaluated by MTT reduction, lactate dehydrogenase(LDH) release assay, and measurement of intracellular ATP levels. 3. In slices submitted to ischemic conditions, 1 mM GMP partially prevented the decrease in cell viability induced by glucose and oxygen deprivation and the addition of KA. 4. KA or N-methyl-D-aspartate (NMDA) receptor antagonists, gamma-D-glutamylamino-methylsulfonate (GAMS) or (+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine maleate (MK-801, 20 microM) also prevented toxicity in hippocampal slices under ischemic conditions, respectively. 5. The association of GMP with GAMS or MK-801 did not induce additional protection than that observed with GMP or that classical glutamate receptor antagonists alone. 6. GMP, probably by interacting with ionotropic glutamate receptors, attenuated the damage caused by glucose and oxygen deprivation in hippocampal slices. This neuroprotective action of GMP in this model of excitotoxicity is of outstanding interest in the search for effective therapies against ischemic injury.

Adenosine Triphosphate↗