title: Maintenance of prolactin (PRL) binding sites in rat liver cells in suspension culture: effect of PRL and of inhibitors of various cellular functions.
abstract: An in vitro method to study the regulation of PRL receptors has been established using adult rat liver cells cultured in a continuous suspension in L-15 medium. PRL binding averaged 28.2 +/- 1.8% of the added labeled hormone per 10(6) cells in freshly isolated liver cells prepared from female rats treated with 17 beta-estradiol. When these cells were incubated at 37 C, binding rapidly declined by 50% at 10 h and 90% at 48 h. This rapid decline could be counteracted by the inclusion of ovine PRL (50 nM), which maintained initial PRL receptor levels up to 48 h of culture. Higher concentrations of PRL (2.5 microM) induced a rapid down-regulation, apparent at 2 and 10 h of culture. Cycloheximide (50 micrograms/ml) induced a slight diminution of control PRL receptor levels and partially reversed the effect of 50 nM PRL. Approximately 60% of the PRL receptors were resistant to the effect of cycloheximide. On the other hand, actinomycin D (10 micrograms/ml) had no effect on PRL receptor levels in control and only a very slight effect in PRL-treated cells. Dinitrophenol, which blocks metabolic oxidation, also partially reversed the effect of 50 nM PRL although it was without any significant effect on control levels. Chloroquine (100 microM) and colchicine (1 microM) failed to alter PRL binding either in the absence or presence of 50 nM PRL. Our results suggest that the existence of regulatory factors occurring in vivo, which are absent in the culture medium, could be responsible for the decline in PRL receptor levels in the control hepatocytes. PRL itself could be one of these factors. On the other hand, and in agreement with the putative actions of the drugs utilized, the mechanism of the PRL-induced maintenance of receptor levels appears to lie in part with an effect on receptor synthesis at the translational (ribsomal) level but to be independent of the internalization or of lysosomal degradation.
mesh_terms: D000818:Animals; D001665:Binding Sites; D002478:Cells, Cultured; D002738:Chloroquine / Q000494:pharmacology; D003078:Colchicine / Q000494:pharmacology; D003513:Cycloheximide / Q000494:pharmacology; D003609:Dactinomycin / Q000494:pharmacology; D004140:Dinitrophenols / Q000494:pharmacology; D005260:Female; D006652:Histological Techniques; D008099:Liver / Q000166:cytology / Q000378:metabolism*; D011388:Prolactin / Q000378:metabolism* / Q000494:pharmacology; D051381:Rats
chemical_list: D004140:Dinitrophenols; D003609:Dactinomycin; D002738:Chloroquine; D011388:Prolactin; D003513:Cycloheximide; D003078:Colchicine