We have been interested in elucidating the cell-specific transcriptional factors that regulate activation of the proenkephalin (PENK) gene in bovine adrenal medullary chromaffin (BAMC) cells and C-6 rat glioma cells. Previous studies indicate that PKC pathway regulates the expression of PENK in BAMC cells. In order to compare the cell-specific expression of this opioid peptide gene, we have used C-6 glioma cells as a model to compare differences in the cellular factors which are involved in the transcriptional regulation. The expression of proenkephalin (PENK) mRNA in C-6 rat glioma cells was stimulated by norepinephrine ( a beta-adrenergic agonist) and markedly enhanced by the addition of dexamethasone (a glucocorticoid agonist) to the culture medium, although no induction of glucocorticoid-response-element (GRE)-binding proteins was detectable. In contrast, the stimulation of PENK mRNA expression was not observed with a protein kinase C activator, 12-tetradecanoyl phorbol 13-acetate (TPA), which stimulated the expression of c-fos and c-jun mRNA and their proto- oncoproteins (c-Fos and c-Jun). In addition, an AP-1 activity was induced by TPA and an induction of a kappaB-like binding activity was found with TPA plus cycloheximide treated cells. Together, it suggests that activation of PENK gene in C-6 cells is mainly mediated through beta- adrenergic agonist-elicited cyclic AMP signal pathway, and induction of AP- 1 and kappaB-like binding activities appear not to participate in gene activation. Interestingly, the Western blot data showed no increase in intracellular levels of proenkephalin between control and treated cells. However, a marked increase in immunoreactivities for proenkephalin and its derivative, [Met5]enkephalin was detected in medium and a lesser elevation in cells from modulator-treated cell culture through the time-course. These results indicated that there was an association between an increase in PENK mRNA expression and an elevation of proenkephalins and, subsequently, the synthesized proenkephalins were released into the medium.
