We have studied the chronic effect of cholera toxin (CTX) on prolactin synthesis and secretion in GH3 pituitary-tumour cells. Time-course analysis showed that prolactin secretion increased with time of CTX exposure, reached a peak at 3 h, and decreased thereafter. Prolactin synthesis was also shown to be stimulated by CTX. The basic and forskolin-stimulated cyclic AMP levels of the CTX-treated cells followed a biphasic time response similar to that of prolactin secretion. Exposure of cells to CTX for more than 3 h abolished the subsequent CTX-catalysed ADP-ribosylation in vitro. Moreover, a significant decrease in the pertussis-toxin-catalysed ADP-ribosylation was found after cells were exposed to CTX for longer than 6 h. Western-blot analysis indicated that the amount of Gs alpha (alpha-subunit of Gs) protein increased within 3 h, followed by a gradual decrease to 50% of the control level at 24 h. The accumulation of Gs alpha mRNA increased within 6 h of CTX exposure, and decreased thereafter to 40% of the basal level at 48 h. Our findings that prolonged treatment of CTX induced similar patterns of time responses in Gs alpha protein expression, cyclic AMP production and prolactin secretion indicate that CTX-induced changes in Gs alpha protein levels may be responsible for the cellular response leading to prolactin secretion.

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