NEURODEGENERATION MEDIATED BY GLUTAMATE AND BETA-AMYLOID PEPTIDE - A COMPARISON AND POSSIBLE INTERACTION

In Alzheimer's disease, abnormal extracellular accumulations of P-amyloid (a major component of the senile plaques) and of the excitatory amino acid glutamate are both thought to be associated with degeneration of nerve cells. In the present study, using cultured cortical or hippocampal neurons as an in vitro model, we compared the effects of various factors influencing neurodegeneration mediated by glutamate or by beta-amyloid peptide (A beta). We also asked the question: does long-term treatment with sublethal doses of A beta-(25-35) potentiate glutamate-mediated excitotoxicity? Neuronal cell death was quantified using the lactate dehydrogenase (LDH) method. Since extracellular LDH remains stable for days, the magnitude of relative afflux of LDH correlates in a linear fashion with the number of damaged neurons in cultures. When applied singly, both glutamate (for 15 min) and A beta-(25-35) or its parent peptide A beta-(1-40) (continuously) produced a dose-dependent neuronal degeneration. In the case of glutamate, the half-maximal effects were observed at about 0.08 mM glutamate for both cerebral cortical and hippocampal neurons (cultured for 13 days in vitro, DIV). The effect of A beta-(25-35) was also time-dependent, while neurons grown in a chemically defined medium showed relatively greater susceptibility to A beta-(25-35) than those cultured in a serum-containing medium. These differential effects were not related to the presence of different numbers of glial cells in the cultures. Treatment with different doses of the antimitotic inhibitor, cytosine arabinoside, for 24 h (6-7 DIV) produced at 13 DIV cortical neuronal cultures with varying numbers of astrocytes, as determined by the astrocyte-specific enzyme glutamine synthetase. The presence of astrocytes decreased the toxicity of glutamate for neurons. The modulation was due to uptake of glutamate by astrocytes, thereby reducing its effective concentration, as the effect was seen at 0.1 mM and not at 10 mM glutamate. Incorporation of an NMDA receptor mediated Ca2+ ion channel blocker, MK-801, together with glutamate completely inhibited degeneration of cortical neurons, and pretreatment of cultures with basic fibroblast growth factor for 2 days did so partially. However, these compounds had no effect on neurotoxicity mediated by A beta-(25-35). Lastly, the effect of glutamate interacted with that of A beta-(25-35). Pretreatment of cortical neurons for 2 days with 10 mu M A beta-(25-35) by itself had no appreciable effect, but it potentiated significantly the degeneration of these neurons mediated by glutamate. On the basis of these findings, we propose that a subtle abnormality in glutamate metabolism in asymptomatic individuals may potentiate the pathophysiological consequences related to the accumulation of beta-amyloid in selectively vulnerable neurons during the course of Alzheimer's disease.