Investigations of non-NMDA receptor-induced toxicity in serum-free antioxidant-rich primary cultures of murine cerebellar granule cells

A culture system was developed whereby murine cerebellar granule cells were grown under serum-free conditions in chemically defined B27-supplemented neurobasal medium plus depolarizing K+ levels, to allow the investigation of the role of agonists at the kainate and alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionate (AMPA) receptors in glutamate-mediated neurotoxicity. Neurones were killed in a concentration-dependent manner by L-glutamate, kainate and its analogues, domoate and 4-(2-methoxyphenyl)-2-carboxy-3-pyrrolidineacetic acid, but not by (S)-AMPA or (S)-5-fluorowillardiine. Kainate (60% maximal cell death at 1mM) was markedly more toxic than NMDA (40% maximal cell death at 1mM) and was shown to be the predominant cause of excitatory amino acid-induced toxicity in these cells as the neuronal death induced by KA was attenuated by the non-NMDA antagonist CNQX, but not the AMPA antagonist LY293558. This study suggests that serum-free cultures of cerebellar granule cells in B27-supplemented neurobasal medium provide a valuable model system for investigations of the role of the kainate receptor in excitatory amino acid-induced neurodegeneration.