Development of excitatory amino acid induced cytotoxicity in cultured neurons

The neurotoxicity of the excitatory amino acids (EAAs) l ‐glutamate ( l ‐glu), l ‐aspartate ( l ‐asp), N ‐methyl‐ d ‐aspartate (NMDA), kainate (KA), quisqualate (QA) and RS ‐α‐amino‐3‐hydroxy‐5‐methyl‐4‐isoxazolopropionate (AMPA) was followed as a function of development in primary cultures of cerebral cortex neurons and cerebellar granule cells. These two types of neurons express, respectively, glutamate receptor subtypes with sensitivity to all of these excitatory amino acids or only to glutamate and aspartate. None of the EAAs were toxic in cerebral cortex neurons at 2 days in culture, whereas at culture day 4 the neurons became sensitive to glutamate, at day 5 to KA followed by sensitivity to QA at day 6, and finally to NMDA, l ‐asp and AMPA at day 7. The rank order of potency of the EAAs was in cerebral cortex neurons cultured for 12 days: l ‐asp ( ed 50 = 0.5 μM) = l ‐glu ( ed 50 = 1 μM) > AMPA( ed 50 = 10 μM) > NMDA ( ed 50 = 65 μM) > QA = KA ( ed 50 = 100 μM). Cerebellar granule cells were insensitive to all of the EAAs at 3 and 5 days in culture but at day 8 the cells became sensitive to toxicity induced by l ‐glu ( ed 50 = 70 μM) and l ‐asp ( ed 50 = 30 μM). In order to determine Ed 50 values for l ‐asp and l ‐glu accurately, media in these experiments also contained 500 μM of the glutamate uptake inhibitor l ‐aspartate‐β‐hydroxamate. It is concluded that the EAA‐induced cytotoxicity is strictly related to the glutamate receptor subtype expression in the neurons. Furthermore, based on the differences in temporal development of toxicity of the different EAAs in cortex neurons, it is evident that all glutamate receptor subtypes independently mediate the cytotoxic action of EAAs. Finally, it seems justified to conclude that QA and AMPA do not have strictly analogous actions.