Effects of dexamethosone and glucagon after long-term exposure on cyclic AMP phosphodiesterase 4 in cultured rat hepatocytes.

67% of total cAMP phosphodiesterase activity (PDE) in cultured rat hepatocytes could be detected in the cytosol, 15% in plasma membrane, 15% in 'dense vesicle,' and 3% in endoplasmatic reticulum fractions. Up to 84% of the PDE activity of the cytosol is represented by the rolipram-sensitive PDE 4. ICI 118233-inhibited PDE 3 was found predominantly in membranes. We were able to show that dexamethasone acts on the PDE 4 in cytosolic and plasma membrane fractions whereas glucagon effected the PDE 4 of the cytosol and the PDE 3 in 'dense vesicle' membranes. Primary culture of hepatocytes was used to study long-term effects of dexamethasone and glucagon on PDE 4 activity. Addition of dexamethasone (0.1 microM) at the beginning of cultivation leads to a decrease of total PDE 4 activity whereas after 24 h precultivation no dexamethasone effect could be observed. Glucagon effects on PDE 4 were investigated in 20 h precultured hepatocytes. Maximal stimulation was achieved after 2 h of exposure. PDE 4 subtypes A, B , D and, to a lesser degree, subtype C could be detected by RT-PCR analysis. The results of semiquantitative RT-PCR show that the presence of dexamethasone during the first 24 h of cultivation reduced selectively the transcription of PDE 4D, whereas glucagon was without any effect. Also the translation of PDE 4D was reduced as shown in the Western blot. We would like to discuss the way that dexamethasone influences PDE 4D expression-most likely in combination with other factors such as cytokines--during the time of cell plating, whereas glucagon actions are part of metabolic regulations via phosphorylation reactions.