The effects and interaction between cAMP-analogue dibutyryl-cAMP and calmodulin antagonists were investigated on de novo synthesis and secretion of lipids in cultures of hepatoma McArdle-RH7777 cells and normal rat hepatocytes. DibutyrylcAMP caused a significant decrease in the secretion of de novo synthesized triacyl[3H]glycerol in both cultures of McArdle cells and rat hepatocytes. The inhibitory effect of dibutyryl-cAMP was concentration-dependent and appeared at the lowest concentration examined, 5 μM. Dibutyryl-cAMP at a concentration of 50 μM suppressed secretion of triacylglycerol by approximately 38% (p<0.05) and secretion of phosphatidylcholine by 30% (p<0.05). Dibutyryl-cAMP did not affect the synthesis of triacylglycerol and phosphatidylcholine, except at the highest concentration tested, 500 μM, where both triacylglycerol and phosphatidylcholine synthesis were suppressed significantly.
Anticalmodulin W-7 also inhibited secretion of newly made triacylglycerol in a concentration-dependent manner in both cultures of McArdle cells and rat hepatocytes. W-7 at a concentration of 20 μM suppressed triacylglycerol secretion by about 37% (p<0.05), while the secretion of phosphatidylcholine and synthesis of triacylglycerol and phosphatidylcholine were not affected, unless at more than 20 μM concentration, at which both triacylglycerol and phosphatidylcholine synthesis were decreased significantly.
The inhibitory effect elicited by dibutyryl-cAMP (100 μM) was not abolished in the presence of calmodulin antagonists, W-7 (20 μM), trifluoperazine (20 μM) and chlorpromazine (20 μM). The simultaneous effects of dibutyryl-cAMP and either calmodulin antagonists were not additive or synergistic. None of the calmodulin antagonists affected the cellular content of de novo synthesized triacylglycerol and phosphatidylcholine significantly. Neither dibutyryl-cAMP nor any calmodulin antagonist or their combination did affect the overall rate of de novo synthesis of triacylglycerol and phosphatidylcholine. All calmodulin antagonists examined alone also had a net significant inhibitory effect on the secretion of newly made triacylglycerol. The results presented here suggest that calmodulin antagonists have net direct effects and hence could not overcome dibutyryl-cAMP-induced suppressive effects on the secretion of newly made triacylglycerol. The cell types, normal hepatocytes ! relative to hepatomas, did not influence the results.
