Abstract
Effects of calcium agonists (A23187, caffein, ryanodine, and thapsigargin) on estradiol-induced vitellogenin (VTG) induction were examined in primary hepatocyte culture in the rainbow trout Oncorhynchus mykiss. These agonists are effective in depleting calcium in various calcium stores. Hepatocytes were precultured for 2 days and then estradiol-17β (E2, 2×10-6M) and the agonists were simultaneously added to the incubation medium. The hepatocytes were cultured for 7 more days and then spent medium was analyzed by SDS-PAGE. The relative rate of VTG production was evaluated by measuring the integrated optical density of the main VTG band and was expressed as the percentage of VTG to total proteins including the VTG. The addition of A23187 (10-7_??_10-6M), caffein (10-4_??_10-2M) or thapsigargin (10-7_??_10-6M) to the incubation medium specifically reduced VTG production by hepatocytes in a concentration-dependent way. Ryanodine also reduced the production to about 60% of the control at a concentration of 10-4M. These results suggest that the synthesis of VTG is more susceptible to intracellular sequestered calcium, at least to calcium in the endoplasmic reticulum, than are other hepatocyte-derived proteins in rainbow trout.
