An enhanced expression of the immediate early gene, Egr-1, is associated with neuronal apoptosis in culture

Cultured cerebellar granule cells grown in medium containing 10 mM K+ (K10) underwent apoptosis after four to five days in vitro, unless they were res cued by the addition of insulin-like growth factor-I. The few GABAergic neu rons present in the cultures were more resistant to apoptotic degeneration, as indicated by double fluorescent staining with the chromatin dye Hoechst 33258 and with glutamate decarboxylase-67 antibodies. As compared with sis ter cultures grown in 25 mM K+, K10 cultures showed an increased expression of the Egr-1 protein and a reduced expression of the Fos protein, The incr ease in Egr-1 was more substantial in granule cells than in GABAergic neuro ns, and was not oberved in K10 cultures chronically exposed to insulin-like growth factor-I. To examine the temporal relationship between the increase in Egr-1 and the development of programmed cell death, we induced apoptosi s in K25 cultures at six days in vitro by replacing their medium with serum -free K10 medium. A substantial, but transient, increase in Egr-1 expressio n was observed in granule cells 6 h after switching the medium, a time that preceded the appearance of the phoenotypical markers of apoptotic death. A n early reduction in the Fos protein was observed after switching the mediu m from K25 into serum-free K10, but also after switching the medium into se rum-free K25, a condition which was not associated with the development of apoptosis nor with the increase in Egr-1. We suggest that a transient induction of Egr-1 contributes to the chain of events leading to the execution phase of neuronal apoptosis in culture. (C) 1999 IBRO. Published by Elsevier Science Ltd.