Continuous labeling of pituitary cells with [3H]-leucine resulted in a linear accumulation of [3H] prolactin in control cultures, but in bromoergocryptine (CB-154) treated cultures the rate of [3H]prolactin accumulation decreased with time. The possibility that this decreased accumulation of labeled prolactin was due to CB-154-induced prolactin degradation was examined by incubating cells for 30 min with [3H]leucine followed by incubation in a chase medium containing a 400-fold excess of unlabeled leucine. in control cultures, there was little degradation of [3H] prolactin over a 24-h period. in cultures containing CB-154 in the chase incubation medium, there was a 22% decrease in labeled prolactin after 8 h and a 50% decrease after 24 h. Pretreatment of cells with CB-154 for 24 h before pulse-chase analysis resulted in a greater rate of prolactin degradation than was observed in cells treated with CB-154 during the chase incubation only. CB-154 treatment did not affect the degradation of nonprolactin proteins, demonstrating the specificity of its effects. Cycloheximide did not affect prolactin degradation in CB-154-pretreated cells; however, cycloheximide blocked the ability of CB-154 to induce prolactin degradation when the two drugs were added simultaneously. The relationship between prolactin synthesis and degradation was examined in cells treated for varying times with CB-154 and then pulsed for 30 min with [3H] leucine followed by a 4-h chase incubation. Prolactin synthesis declined sharply after 1-2 days of CB-154 treatment and reached a new plateau of 22% of control values after 4 days of treatment. Prolactin degradation was maximal after 1 day of CB-154 treatment and returned toward control values after 3-4 days of treatment. Lysosomes are likely involved in CB-154-induced prolactin degradation as chloroquine is able to partially block CB-154 effects. These studies suggest that CB-154 is able to induce substantial prolactin degradation. Thus, prolactin degradation is involved in removal of excess prolactin which accumulates in the pituitary when prolactin secretion is inhibited.
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