Enhanced Glutamate Uptake Research Articles

Guanosine enhances glutamate uptake in brain cortical slices at normal and excitotoxic conditions.

1. The effect of guanosine on L-[2,3-3H]glutamate uptake was investigated in brain cortical slices under normal or oxygen-glucose deprivation (OGD) conditions. 2. In slices exposed to physiological conditions, guanosine (1-100 microM) stimulated glutamate uptake (up to 100%) in a concentration-dependent manner when a high (100 microM) but not a low (1 microM) concentration of glutamate was used. 3. In slices submitted to OGD, guanosine 1 and 100 microM also increased 100 microM glutamate uptake (38 and 70%, respectively). 4. The increasing of glutamate and taurine released to the incubation medium in cortical slices submitted to OGD were significantly attenuated by the presence of guanosine in the incubation medium. 5. Guanosine prevented the increase in propidium iodide incorporation into cortical slices induced by OGD, indicating a protective role against ischemic injury. 6. These results support the hypothesis of a protective role for guanosine during brain ischemia, possibly by activating glutamate uptake into neural cells.

Isoflurane enhances glutamate uptake via glutamate transporters in rat glial cells.

In this study I examined whether isoflurane, an inhalational anesthetic used commonly in clinical practice, affected glutamate uptake via glutamate transporters, proteins expressed in the plasma membrane of cells in the central nervous system. Isoflurane at clinically relevant concentrations (1-3%) caused a time-, sodium- and concentration-dependent increase of glutamate uptake in primary cultures of rat cerebral mixed glial cells. This enhancement was inhibited by a specific glutamate transporter inhibitor. The study also demonstrated that 2.0% isoflurane significantly increased both Vmax and Km of transporter-mediated glutamate uptake. Thus, isoflurane enhances glutamate uptake by a pathway that requires function of glutamate transporters. This represents a novel pharmacological effect of inhalational anesthetics and may contribute to isoflurane-induced anesthesia and neuroprotective effects.

Activation of glutamate uptake by guanosine in primary astrocyte cultures.

Guanine-based purines have been shown to modulate the effects of glutamate, which is essential for brain function and mediates excitotoxicity. In the search for a mechanism involving the interaction between purine nucleoside guanosine and glutamate, we found that guanosine dose-dependently, significantly (63%) and potently (EC50 =2.47 microM) enhanced glutamate uptake in cultured astrocytes. This effect was not inhibited by the blocker of nucleoside transporter dipyridamole nor by the adenosine antagonist theophylline, suggesting an extracellular site of action without the involvement of adenosine receptors. These results indicate a regulatory role of guanosine on extracellular levels of glutamate, possibly contributing for protecting neural cells against glutamate-induced excitotoxicity.

Neuronal Release of Vasoactive Intestinal Peptide Is Important to Astrocytic Protection of Neurons from Glutamate Toxicity

Astrocytes regulate clearance of glutamate from the vicinity of neurons. This helps to protect neurons directly from glutamate toxicity. Recent findings have indicated that a complex molecular interaction between neurons and astrocytes that is necessary for this protection occurs. In the present investigation the role of vasoactive intestinal peptide (VIP) in signaling between neurons and astrocytes was investigated. VIP was found to be necessary for the protective effects of astrocytes in a coculture system. VIP in combination with neuronal-conditioned medium enhanced glutamate uptake by astrocytes. Also, VIP enhanced the expression of the high-affinity VIP receptor, increased astrocytic release of interleukin-6, and indirectly reduced the toxicity of glutamate in neuronal-conditioned astrocyte medium. These results indicate that VIP is essential to the molecular interaction of neurons and astrocytes and is involved in the regulation of the protective effects of astrocytes for neurons.

Increase of glutamate uptake in astrocytes: a possible mechanism of action of volatile anesthetics.

Glutamate is the most ubiquitous excitatory neurotransmitter in the vertebrate central nervous system. Astrocytes play an important role in terminating glutamatergic neurotransmission by removing released glutamate from the synaptic cleft. The authors examined the effects of several anesthetics on the glutamate uptake activity of astrocytes. Cultured astrocytes from hippocampi of rat embryos were incubated with solution containing [3H]glutamate, which was pre-equilibrated with 0-4% halothane at 37 degrees C. The uptake activity was evaluated as the amount of radioactivity per cell of protein. When the reaction solution was equilibrated with 4% halothane, glutamate uptake increased to about 165% of the control. The effect of halothane was dose-dependent, and a significant augmentation (30-50%) of glutamate uptake was observed at a range in clinical use concentrations (1-2%). On the other hand, the uptake of gamma-aminobutyric acid, an inhibitory transmitter, was hardly affected by 1-4% halothane. The effect of halothane on glutamate uptake was also examined in neuron-rich culture, and similar augmentation was observed, although the extent was less than that in astrocyte culture. Biochemical subcellular fractions (i.e., glial plasmalemmal vesicles and synaptosomes) were also examined, however, only slight (not significant) increase was detected in the glutamate uptake activity. Other volatile anesthetics, such as enflurane, isoflurane, and sevoflurane, also enhanced glutamate uptake, whereas the intravenous anesthetics ketamine and pentobarbital showed no effect on glutamate uptake. The increase of glutamate uptake by astrocytes in the presence of volatile anesthetics potentially attenuates excitatory synaptic transmission in the entire central nervous system, a finding that may explain in part the action of volatile anesthetics.

Novel neuron-related regulatory mechanisms for astrocytic glutamate and GABA high affinity uptake

The effect of conditioned media from cultures of cerebral cortical neurons and cerebellar granule cells on high affinity uptake of glutamate and GABA into cerebellar astrocytes cultured from 7-day-old rats was studied. Granule cell-conditioned media selectively enhanced GABA uptake whereas cerebral cortex neuron-conditioned media selectively enhanced glutamate uptake into these astrocytes. This selectivity was apparently restricted to cerebellar astrocytes since astrocytes cultured from cerebral cortex of 7-day-old rats exhibited a slight increase in both GABA and glutamate uptake after culture in conditioned media from both types of neurons. The results point to as yet unknown regulatory mechanisms for the development of astrocytic uptake of transmitter amino acids which may be dependent upon the regional origin of the astrocytes as well as the surrounding neuronal cell type.

3H]Glutamate uptake at insect neuromuscular junctions: effect of chlorpromazine

Previous studies have shown that in an isolated cockroach nerve-muscle preparation, nerve stimulation causes an increase in uptake of [ 3H]glutamate during subsequent incubation in a medium containing this amino acid 21,50. The enhanced glutamate uptake is largest in the tracheole cells, sheath cells, especially in the junctional regions, and in the postsynaptic regions of the muscle. This differential effect suggests that nerve stimulation affects more than one glutamate pool, and since glutamate is the presumed transmitter in this system, the uptake at the junction may be involved in transmitter inactivation by re-uptake. In the present study chlorpromazine, which is known to interfere with transmitter re-uptake in a variety of systems, was added to the [ 3H]glutamate incubation medium after prolonged nerve stimulation. It was found that chlorpromazine decreased the stimulation-enhanced uptao a varying to extent in the different muscle and sheath compartments. It did not affect uptake by tracheole cells and had its major effect at the neuromuscular junctions. Chlorpromazine decreased the junctional sheath uptake to the level of the non-junctional sheath, thus eliminating the differential sheath uptake. In the muscle, chlorpromazine did not completely eliminate the difference between junctional and non-junctional uptake, but decreased it greatly. The data suggest that the various pools depleted by nerve stimulation are replenished by different uptake mechanisms. The finding that chlorpromazine has its major effect on the uptake into junctional compartments is consistent with the hypothesis that the junctional sheath and postsynaptic muscle regions are sites of transmitter inactivation by re-uptake.

Glutamate transport in Escherichia coli K-12: nonidentity of carriers mediating entry and exit.

The exit of glutamate from Escherichia coli K-12 cells preloaded with the radioactive amino acid and its relation to the reaction of entry were studied. Experiments with cells preloaded to different intracellular concentrations of radioactive glutamate confirmed our earlier conclusion that glutamate exit was a first-order reaction. l-Glutamate, competitive inhibitors of glutamate uptake (d-glutamate and l-glutamate-gamma-methyl ester), noncompetitive inhibitors of glutamate uptake (l-serine and l-alanine), and the energy poison NaN(3) all accelerated glutamate exit 2.8-fold. No additive effect was observed in the presence of NaN(3) together with l-glutamate. Preloading with cold l-glutamate did not increase the rate of uptake of radioactive glutamate. It is concluded that the acceleration of glutamate exit in the presence of l-glutamate in the medium is not due to exchange diffusion and that l-glutamate and azide affect exit indirectly by preventing recapture. Sucrose, 25%, slowed down glutamate exit by a factor of about 4.7 and increased the steady-state level of glutamate accumulation to about the same extent. Increasing the intracellular K(+) concentration enhanced glutamate uptake but did not affect the half-time of exit. It is concluded that separate carriers are most probably involved in mediating the entry and exit reactions.

Glutamate uptake by a stimulated insect nerve muscle preparation.

Recent reports suggest that glutamate may be the excitatory neuromuscular transmitter in insects. In this study, glutamate uptake by isolated cockroach nerve muscle preparations was investigated by means of chemical and electron microscope radioautographic techniques. We found that the preparation had a high affinity for glutamate and that nerve stimulation enhanced glutamate uptake. Chemical studies showed that the average tissue concentration of glutamate bound during a 1 hr incubation period in 10(-5)M glutamate-(3)H after nerve stimulation was 2.8 x 10(-5)M. Less than 1% of the radioactivity was present in the perchloric acid-precipitated protein fraction. Using electron microscope radioautography, we observed that sheath cells showed the highest glutamate concentration of all cellular compartments. Uptake was greater at neuromuscular junctions than in other regions of the tissue. The data suggest a possible mechanism for transmitter inactivation and protection of synapses from high blood glutamate.