Bioquímica

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 wildtype 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.

glutamate uptake in Aβ 1-40 -treated mice. These results reinforce and extend the notion of the potential 43 neuroprotective action of atorvastatin against the neuronal toxicity induced by Aβ 1-40 . In addition, the 44 present findings suggest that the spatial learning and memory deficits induced by Aβ peptides in rodents may 45 not be entirely related to neuronal damage. 46

Ethnopharmacological relevance: Aloysia gratissima (Gill. et Hook) Tronc. (Verbenaceae) is used traditionally for the treatment of headache, bronchitis, and nervous systems disorders including depression. Aim of the study: To investigate the antidepressant-like and neuroprotective effects of Aloysia gratissima aqueous extract (AE) and the involvement of l-arginine-nitric oxide (NO)-cyclic guanosine monophosphate (cGMP) pathway. Materials and methods: The antidepressant-like effect of AE was evaluated through behavioral despair in forced swimming test (FST) and tail suspension test (TST). Swiss albino mice were treated by oral route and after 1 h were analyzed the time of immobility in the FST and TST. In addition, the neuroprotective effect of AE against glutamate excitotoxicity was evaluate through cell viability of hippocampal slices, phosphorylation of Akt, and the immunocontent of inducible oxide nitric synthase (iNOS) were investigated by western blotting. Results: The immobility time in the FST and TST were reduced by AE (100-1000 and 10-300 mg/kg, respectively). The antidepressant-like effect of AE in the TST was prevented by the pretreatment with N-methyl-d-aspartate (NMDA), l-arginine or sildenafil. The subeffective dose of AE produced a synergistic antidepressant-like effect with MK-801 (an antagonist of NMDA receptor), methylene blue, l-NNA (an inhibitor of NO synthase) or ODQ (an inhibitor of soluble guanylate cyclase). In ex vivo experiments, pretreatment with AE prevented the loss of cell viability induced by glutamate, thus affording neuroprotection. Glutamate toxicity caused a decreased Akt phosphorylation and an increased iNOS expression. Conclusions: The present study provides convincing evidence of neuroprotection and the involvement of the l-arginine-NO-cGMP pathway in the antidepressant-like effect of AE. Therefore, AE could be of potential interest for the treatment of depressive disorders and neurological conditions associated with glutamate excitotoxicity.

Thyroid hormone (T 3 ) regulates the growth and differentiation of rat cerebellar astrocytes. Previously, we have demonstrated that these effects are due, at least in part, to the increased expression of extracellular matrix molecules and growth factors, such as fibroblast growth factor-2. T 3 also modulates neuronal development in an astrocytemediated manner. In the mammalian central nervous system, excitatory neurotransmission is mediated mainly by glutamate. However, excessive stimulation of glutamate receptors can lead to excitotoxicity and cell death. Astrocytic glutamate transporters, GLT-1 and GLAST, play an essential role in the clearance of the neuronal-released glutamate from the extracellular space and are essential for maintaining physiological extracellular glutamate levels in the brain. In the present study, we showed that T 3 significantly increased glutamate uptake by cerebellar astrocytes compared with control cultures. Inhibitors of glutamate uptake, such as L-PDC and DL-TBOA, abolished glutamate uptake on control or T 3 -treated astrocytes. T 3 treatment of astrocytes increased both mRNA levels and protein expression of GLAST and GLT-1, although no significant changes on the distribution of these transporters were observed. The gliotoxic effect of glutamate on cultured cerebellar astrocytes was abolished by T 3 treatment of astrocytes. In addition, the neuronal viability against glutamate challenge was enhanced on T 3 -treated astrocytes, showing a putative neuroprotective effect of T 3 . In conclusion, our results showed that T 3 regulates extracellular glutamate levels by modulating the astrocytic glutamate transporters. This represents an important mechanism mediated by T 3 on the improvement of astrocytic microenvironment in order to promote neuronal development and neuroprotection. V V C 2008 Wiley-Liss, Inc.

Glutamate excitotoxicity may culminate with neuronal and glial cell death. Glutamate induces apoptosis in vivo and in cell cultures. However, glutamate-induced apoptosis and the signaling pathways related to glutamateinduced cell death in acute hippocampal slices remain elusive. Hippocampal slices exposed to 1 or 10 mM glutamate for 1 h and evaluated after 6 h, showed reduced cell viability, without altering membrane permeability. This action of glutamate was accompanied by cytochrome c release, caspase-3 activation and DNA fragmentation. Glutamate at low concentration (10 lM) induced caspase-3 activation and DNA fragmentation, but it did not cause cytochrome c release and, it did not alter the viability of slices. Glutamate-induced impairment of hippocampal cell viability was completely blocked by MK-801 (non-competitive antagonist of NMDA receptors) and GAMS (antagonist of KA/AMPA glutamate receptors). Regarding intracellular signaling pathways, glutamate-induced cell death was not altered by a MEK1 inhibitor, PD98059. However, the p38 MAPK inhibitor, SB203580, prevented glutamate-induced cell damage. In the present study we have shown that glutamate induces apoptosis in hippocampal slices and it causes an impairment of cell viability that was dependent of ionotropic and metabotropic receptors activation and, may involve the activation of p38 MAPK pathway.

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 10 mM) had no effect on synaptosomal l-[ 3 H]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 10 mM 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 M) did not alter glutamate release per se, but completely abolished the QA-induced glutamate release. NMDA (50 M) also stimulated glutamate release, without altering QA-induced glutamate release, suggesting that QA effects were exerted via NMDA receptors. QA (5 and 10 mM) 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.