In the present report we describe the effect of glutamate on respiratory activity in primary cultures of astrocytes, derived from cerebral cortex of newborn rat. Glutamate (100 microM) caused an increased oxygen consumption. This effect could not be inhibited by antagonists to the NMDA or AMPA/kainate receptors. Neither trans-ACPD (an agonist to the metabotropic glutamate receptor) nor the Krebs cycle intermediate alpha-ketoglutarate had any effect on the respiratory rate. An uncontrolled influx of Na+, caused by gramicidin, could mimic the glutamate effect on respiratory activity. In addition, the glutamate effect was abolished by addition of ouabain or replacement of Na+ by Li+ in the perfusion buffer. We conclude that the co-transport of Na+, in the Na(+)-dependent high-affinity glutamate uptake system, mediated the glutamate-induced increase in oxygen consumption through an increased activity of Na+/K(+)-ATPases.

One of the pathological hallmarks of Alzheimer's disease (AD) is the presence of amyloid plaques. The main constituent of the amyloid plaques is the amyloid beta-peptide (A beta) shown to activate glial cells in vitro. A growing body of evidence suggests that these cells contribute to neurotoxicity through production of inflammatory cytokines, chemokines, and neurotoxic substances, such as reactive oxygen species (ROS). In this study, mRNA levels of the inflammatory cytokines interleukin (IL)-1alpha and beta, and IL-6 were analysed by reverse transcriptase-polymerase chain reaction (RT-PCR) in rat primary mixed glial cells after treatment with A beta(25-35), a biologically active fragment of A beta peptide with neurotoxic properties. Clear morphological changes of the astrocytes, as well as proliferation and clustering of microglial cells were observed by light and immunofluorescence microscopy after 24 h treatment. Significant increases in IL-1alpha and IL-6 mRNA levels were detected after 24 and 72 h, whereas significantly increased levels of IL-1beta mRNA could only be detected after 4 h treatment. The most pronounced effect was seen on IL-6 mRNA expression, which increased approx two- to threefold after treatment. In addition, increased secretion of IL-6 was detected after 96 h exposure. Recently, association of IL-1alpha and IL-6 gene polymorphism with AD was reported, suggesting that these cytokines may play an important role in the development of the disease. The increased mRNA levels of IL-1alpha and IL-6 in parallel with the morphological changes in the mixed glial-cell cultures support that these cytokines may be involved in A beta-induced gliosis and in the pathogenesis of AD.

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1. Current opinions on the mechanisms for glutamate-mediated neurotoxicity are reviewed. The protective role of astrocytic high-affinity glutamate transport is also discussed. 2. Low-density seeding of primary astrocytes from rat hemispheres was found to result in the development of reactive-like astrocytes. Typical glial signs of amyotrophic lateral sclerosis (ALS) could not be induced in astrocyte cultures by serum from ALS-patients. 3. Glutamate (100 mumol/L) was found to induce an increase in respiratory activity in primary cultures of astrocytes. This stimulation appeared to be related to the co-transport of Na2+ with glutamate and a resulting activation of Na2+/K(+)-ATPase. Both basal respiration and glutamate-stimulated oxygen consumption was inhibited by organic solvents. 4. Preliminary results show that heavy metals cause an increase in the mitochondrial DNA content at concentrations that have no effect on growth rate or morphology in a glial cell line. This increase was accompanied by an inhibition of oxygen consumption and an increased production of lactate at unaltered ATP levels.

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