EVIDENCE THAT GLYCERALDEHYDE-3-PHOSPHATE DEHYDROGENASE IS INVOLVED INAGE-INDUCED APOPTOSIS IN MATURE CEREBELLAR NEURONS IN CULTURE

Under typical culture conditions, cerebellar granule cells die abruptl y after 17 days in vitro. This burst of neuronal death involves ultras tructural changes and internucleosomal DNA fragmentations characterist ic of apoptosis and is effectively arrested by pretreatment with actin omycin-D and cycloheximide. The level of a 38-kDa protein in the parti culate fraction is markedly increased during age-induced cell death an d by pretreatment with NMDA, which potentiates this cell death. Conver sely, the age-induced increment of the 38-kDa particulate protein is s uppressed by actinomycin-D and cycloheximide. N-terminal microsequenci ng of the 38-kDa protein revealed sequence identity with glyceraldehyd e-3-phosphate dehydrogenase (GAPDH). A GAPDH antisense oligodeoxyribon ucleotide blocks age-induced expression of the particulate 38-kDa prot ein and effectively inhibits neuronal apoptosis. In contrast, the corr esponding sense oligonucleotide of GAPDH was completely ineffective in preventing the age-induced neuronal death and the 38-kDa protein over expression. Moreover, the age-induced expression of the 38-kDa protein is preceded by a pronounced increase in the GAPDH mRNA level, which i s abolished by actinomycin-D, cycloheximide, or the GAPDH antisense, b ut not sense, oligonucleotide, Thus, our results suggest that overexpr ession of GAPDH in the particulate fraction has a direct role in age-i nduced apoptosis of cerebellar neurons.