Glutamate-mediated neuroprotection against N-methyl-D-aspartate toxicity:: A role for metabotropic glutamate receptors

We studied N-methyl-D-aspartate-induced cell death in organotypic hippocampal slices from seven-day-old Wistar rat pups cultured for 12-14 days in a medium containing no added glutamate. Propidium iodide fluorescence intensity was used as an indicator of cell death measured with the help of confocal microscopy. Exposure of slices for 2 h to L-glutamate (1-500 mu M) prior to the N-methyl-D-aspartate challenge significantly reduced N-methyl-D-aspartate-induced cell death. Glutamate at 10 and 500 mu M concentrations was highly protective against N-methyl-D-aspartate-induced cell death, but was less protective at the 1 mu M concentration. The protection was not blocked by the Na+ channel blocker tetrodotoxin (1 mu M): the N-methyl-D-aspartate receptor antagonist D-2-amino-5-phosphonopentanoic acid (20 mu M) or the alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate receptor antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (20 mu M). 1S,3R 1-Aminocyclopentane-trans-1,3-dicarboxylic acid, an agonist at metabotropic glutamate receptor types 1, 2/3 and 5, was protective at 100 mu M but not at 50 mu M. In contrast, the ionotropic glutamate receptor agonist aspartate (250 mu M) facilitated N-methyl-D-aspartate toxicity. Treatment of slices with the protein kinase C inhibitor staurosporine (0.2 mu M) or antisense oligonucleotide (10 nM, 72 h) that selectively inhibits metabotropic glutamate receptor type 5 synthesis significantly reduced glutamate protection. These results suggest that ambient glutamate may reduce nerve cell susceptibility to injury caused by excessive N-methyl-D-aspartate receptor activation by acting at metabotropic glutamate receptors linked to protein kinase C. (C) 2000 IBRO. Published by Elsevier Science Ltd. All rights reserved.