Reversal of Glycerol Ether-Stimulated Acetylcholinesterase Activity by c-fos Antisense Oligonucleotide in Primary Neuronal Cultures

Abstract

We have previously reported that dodecylglycerol (DDG), an alkyl glycerol lipid similar to platelet activating factor (PAF), stimulates differentiation in primary neuronal cultures obtained from fetal rat cerebral cortex, hippocampus and cerebellum (Ved et al., 1994). The neuronal differentiation was evident both morphologically as development of axon-like extensions and biochemically as an increase in neuron specific enzyme activity (Ved et al., 1991). We also reported that DDG mediated neuronal differentiation resulted in a transient increase in c-fos mRNA levels (Ved et al., 1993). The objectives of present study were to determine if c-fos antisense oligonucleotide (ASO) would reverse DDG-stimulated acetylcholinesterase (AChE) and choline acetyltransferase (ChAT) activities, and neuronal differentiation. Primary enriched neurons derived from embryonic (E-17) rat cerebral cortex, cerebellum or hippocampus were treated in a serum-free media with either vehicle, DDG (4 μM) or DDG with either c-fos antisense oligonucleotide (ASO) (5–20 μM), c-myc ASO or non-sense ASO for 24 hrs. Treatment with DDG produced a maximal stimulation in AChE and ChAT activities, and outgrowth of neuronal processes in cultures obtained from cerebellum and a minimum effects in those obtained from cerebral cortex. The percent stimulation in AChE activity following DDG treatment in these neuronal cells were approximately 200%, 150% and 65% above the control levels in cerebellum, hippocampus and cortex, respectively. The percent stimulation in ChAT activity following DDG treatment in neuronal cells were approximately 260%, 220% and 45% above the control levels in cerebellum, hippocampus and cortex, respectively. Pretreatment with c-fos ASO partially inhibited DDG-stimulated AChE and ChAT activities and outgrowth of the neuronal processes, however, c-myc ASO and nonsense ASO had no significant effect. The inhibitory effect of c-fos ASO on DDG-stimulated AChE and ChAT activities and neuronal differentiation was greatest in neurons obtained from cerebellum and hippocampus (100%–52% inhibition). These results suggest a causative role of c-fos proto-oncogene in DDG-mediated neuronal differentiation and further suggest a direct role of c-fos gene product on modulation of AChE and ChAT enzyme activities in the neuronal cells.