Activation of nerve growth factor synthesis in primary glial cells by phorbol 12-myristate 13-acetate: role of protein kinase C.

Phorbol 12-myristate 13-acetate (PMA) induces a dramatic production of nerve growth factor (NGF) in primary cultures of newborn mouse astrocytes maintained in a serum-free medium. This stimulation is dose-dependent and a maximal effect on the levels of cell-secreted factor was observed at a concentration of 10 nM. At this concentration, the promoting effect of PMA appears much more important than that elicited by 10% fetal calf serum (FCS) under the same culture conditions. PMA acts primarily on the accumulation of NGF mRNA, which was detected by northern blot analysis after 6 h of treatment. This accumulation may be totally or partially prevented when PMA-treated glial cells are concomitantly exposed to the protein kinase inhibitors H-7, H-9, and to a lesser degree, HA-1004. The known specificity of these inhibitors agrees with the possibility that protein kinase C (PKC), which constitutes so far the sole known target of PMA, represents a key element involved in the stimulation of NGF gene. The role of PKC is further supported by the observation that alpha phorbol didecanoate, which has no activity on PKC, is depleted of effect on the synthesis of NGF. Likewise, 1,2-dioctanoylglycerol (1,2-DOG) has a weak, but significant promoting action on the production of NGF, unlike the 1,3-isomer which is not active on PKC. Finally, a treatment of 15 min with 100 nM PMA is sufficient to stimulate the cells, suggesting that the activation phase of PKC, rather than its down regulation, constitutes an important trigger leading to an increased expression of the NGF gene.(ABSTRACT TRUNCATED AT 250 WORDS)