KAINATE-INDUCED APOPTOSIS CORRELATES WITH C-JUN ACTIVATION IN CULTURED CEREBELLAR GRANULE CELLS

We have investigated the involvement of c-Jun in cell death induced by exposure of primary cultures of murine cerebellar granule cells to th e glutamate receptor agonist kainate (KA) and evaluated its possible u se as a marker for apoptosis, Using cerebellar granule cell neurones f rom postnatal day 7 mice, we found that 1 hr exposure to KA (1-1000 mu M) induced a concentration-dependent neuronal cell death with charact eristic apoptotic morphology, including cell shrinkage, neurite blebbi ng and DNA fragmentation, In addition KA-induced a concentration-depen dent expression of c-Jun mRNA and protein as deter mined by in situ hy bridization and immunocytochemistry respectively, DNA fragmentation wa s detected using terminal transferase-mediated nick-end (TUNEL) labell ing and agarose gel electrophoresis, KA-induced cell death was signifi cantly attenuated by the nonNMDA receptor antagonist 6-cyano-7-nitroqu inoxaline-2,3-dione (CNQX; 50 mu M), which shifted the concentration-r esponse curve significantly rightward, The number of apoptotic cell bo dies, determined by TUNEL, was also reduced by CNQX (50 mu M), with on ly 15-20% of neurones staining positive after exposure to 1mM KA, In a ddition, the number of positively stained cells for c-Jun protein and mRNA was substantially reduced by CNQX (50 mu M) as determined by rand om and representative cell counts, These results show for the first ti me that KA induced apoptotic neuronal death in cultured murine cerebel lar granule cells involves the induction of c-Jun mRNA and protein, su ggesting the involvement of this immediate early gene in excitotoxic r eceptor-mediated apoptosis and its potential use as a marker for apopt otic cell death. (C) 1998 Wiley-Liss, Inc.