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Recent studies have demonstrated that astrocytes have much greater abilities to produce and respond to signalling molecules in the CNS than had been previously estimated. We now report a dramatic enhancement in the ability of a glutamate metabotropic receptor agonist, 1-aminocyclopentane-trans-(+/-)-1,3-dicarboxylic acid (trans-(+/-)-ACPD, to stimulate phosphoinositide hydrolysis in astrocytes cultured in a serum-free defined medium compared with astrocytes cultured in conventional serum-containing medium (43.2 +/- 3.6 vs. 3.2 +/- 0.48-fold of basal, respectively). This enhancement was selective to trans-(+/-)-ACPD as little or no difference in the response to carbachol or norepinephrine was seen between the two culture conditions. These results indicate a great potential for the phosphoinositide pathway in astrocyte glutamatergic signal transduction.

High affinity transport of excitatory amino acids such as L-glutamate into astrocytes is necessary for the termination of its excitatory signal and the prevention of its excitotoxic effects. The removal of glutamate from the synaptic cleft is carried out by both sodium- and chloride-dependent systems. Both sodium-dependent D-[3H]aspartate and chloride-dependent L-[3H]glutamate binding were found to increase in the dentate gyrus molecular layer of rats following an entorhinal lesion. The increased binding reached a maximum at 5 and 7 days postlesion and returned to normal by 12 days postlesion. No changes in binding were observed at long time points postlesion. This increased ability to transport glutamate may be a compensatory response to protect the remaining neurons from the excitotoxic conditions that accompany neuronal degeneration.

beta-N-Oxalyl-L-alpha,beta-diaminopropionic acid (beta-L-ODAP) is an excitatory amino acid agonist found in the seeds of Lathyrus sativus that is believed to be the major causative agent in the pathology of human lathyrism. We have found that in addition to its previously recognized neurotoxic properties, beta-L-ODAP is also gliotoxic. When added to cultures of neonatal rat astrocytes, beta-L-ODAP induced a series of morphological changes (e.g., extensive vacuole formation, pale and swollen nuclei with obvious nucleoli, and cellular swelling) that led to the eventual lysis of the glial cells. If the beta-L-ODAP was removed prior to the lysis of the astrocytes, many of the early morphological changes appeared to be reversible. When quantitated by a loss of the lactate dehydrogenase activity, beta-L-ODAP lysed the astrocytes with an LD50 of 2.1 +/- 0.2 mM following 48 h of exposure. Lower concentrations of beta-L-ODAP were found to be more toxic if the duration of the exposure was increased. The results suggest that the overall impact of the toxin on the CNS may represent the cumulative action of beta-L-ODAP at a number of distinct points on both neurons and astrocytes. The potential that these multiple sites of action may affect the normal regulation of extracellular glutamate and, consequently, disturb the balance between its normal and pathological roles is discussed.

Olfactory receptor neurons are a CNS entry point for a wide variety of airborne substances. Therefore, it is probable that detoxification mechanisms are present in these neurons to neutralize such agents. Glutathione (GSH) is an essential component of several detoxification schemes, and in this study we examined the distribution and levels of GSH in the olfactory epithelium, olfactory bulb, cortex, hippocampus and cerebellum in neonatal, weanling, adult and aged rats. We report that GSH is primarily localized to the olfactory receptor neurons and their oxons within the olfactory epithelium. It is also localized within the glomerular neuropil and granule cells of the olfactory bulb. Levels of GSH in the olfactory epithelium and hippocampus do not change as a function of age, although GSH levels decrease in several brain regions, including the olfactory bulb, cerebellum and cortex.

In the present study cultured astrocytes were implanted into the inferior colliculus of rats to create an astrocyte-enriched field that could be examined autoradiographically. The presence of the astrocytes was confirmed with anti-glial fibrillary acidic protein (GFA) immunocytochemistry. We report the presence of a chloride-dependent glutamate binding site on the implanted astrocytes. In the presence of chloride, the specific glutamate binding detected in the implant area was 5-fold greater than that found in a corresponding contralateral region. When the chloride was replaced with acetate, glutamate binding to the astrocytes decreased by more than 80%. The chloride-dependent binding to the astrocytes was insensitive to inhibition by kainic acid (KA) and N-methyl-D-aspartate (NMDA) and sensitive to quisqualate, L-aspartate, L-2-amino-4-phosphonobutyrate, and L-alpha-aminoadipate. The pharmacology of the binding was very similar to that of the in vitro glutamate binding to membranes from cultured astrocytes and to that of a chloride-dependent transport system identified in a glioma cell line. We conclude that the interaction of glutamate with astrocytes is an important component of the total glutamate binding observed in brain slices.

The glutamate analogue 2-amino-4-phosphonobutyric acid (AP4) is a potent antagonist at several synapses where an excitatory amino acid appears to be the neurotransmitter. Previous studies identified a Cl-/Ca2+ dependent [3H]glutamate binding site in synaptic plasma membrane (SPM) preparations that was also labeled by [3H]AP4 and exhibited a pharmacology similar to the AP4 receptor. This report examines the pharmacological specificity in both biochemical and electrophysiological preparations in greater detail. Several compounds are identified which readily interact with the apparent binding site in membranes, but neither mimic nor inhibit the action of AP4 in electrophysiological studies. The rate of dissociation of [3H]AP4 from SPMs is shown to increase in the presence of added AP4 and increasing the osmolarity in the SPM binding assay decreases the level of observed [3H]AP4 binding. These findings indicate both a heterogeneous population of binding sites and the occurrence of transport. It is concluded that much of the AP4 binding observed in SPM preparations is to a site other than the AP4 receptor. The results provide a further pharmacological description of AP4 receptors which should facilitate the identification of the receptor in biochemical preparations.