CELLULAR AND MOLECULAR CORRELATES OF GLUTAMATE-EVOKED NEURONAL PROGRAMMED CELL-DEATH IN THE IN-VITRO CULTURES OF RAT HIPPOCAMPAL DENTATE GYRUS

An excessive neuronal stimulation through glutamate receptors is known to result in excitotoxic cell death of apoptotic (programmed) charact er. Granule cells of hippocampal dentate gyrus are believed to be part icularly resistant to excitotoxic insults, despite the fact that pyram idal neurons of the hippocampus propel are apparently the most vulnera ble brain cells. In this study, we report that neurons derived from th e rat 5-day-old dentate gyrus, and maintained in vitro for 6 days, may undergo apoptosis after treatment with L-glutamate, in a dose-depende nt manner - with up to 80% of neurons displaying features of programme d cell death after 24 h exposure to 0.5 mM glutamate. This conclusion is based on morphological evaluation of the cultures, nuclear staining with Hoechst 33258 and acridine orange revealing chromatin abnormalit ies, as well as terminal transferase labeling of DNA fragmentation. Si nce apoptosis is believed to be an active process involving gene expre ssion, immunocytochemical investigation of c-Fos and c-Jun transcripti on factor proteins was performed. Elevated expression of both proteins was found to follow quickly (within 1 h) after addition of glutamate. However, this effect was not close-dependent, thus it does not provid e clear correlations to the programmed cell death. In conclusion, this study reports on the establishment of a novel apoptotic model of exci totoxicity, and invites further efforts to investigate a basis for in vitro susceptibility and in vivo resistance of centate gyrus granule c ells to excitotoxic insult evoking apoptosis. (C) 1997 Elsevier Scienc e Ltd.