COMPARISON OF THE EFFECTS OF CYCLOHEXIMIDE AND INHIBIN ON THE GONADOTROPIN SUBUNIT MESSENGER RIBONUCLEIC-ACIDS

Cycloheximide (CHX) has been shown to mimic the action of inhibin on gonadotropin secretion by pituitary cell cultures. We showed previously that suppression of FSH secretion by inhibin is associated with a rapid and profound suppression of FSH beta mRNA levels. The present study was designed to examine the mechanism of action of CHX and to determine whether inhibin's actions involve new protein synthesis. Pituitary cell cultures were treated with control medium or medium containing inhibin, CHX, or inhibin plus CHX for 2 or 6 h. At 6 h, secretion of FSH was decreased by inhibin (72% of control), CHX (58% of control), and the combined inhibitors (56% of control). LH secretion was not significantly changed, while that of free alpha-subunit was reduced only by CHX (68% of control). Levels of FSH-beta, LH-beta, and alpha-subunit mRNAs were measured by Northern analysis. At 2 h inhibin decreased FSH-beta mRNA to 49% of the control value. CHX alone had no effect, while CHX plus inhibin produced intermediate levels (77% of control). By 6 h, however, inhibin and CHX each decreased FSH-beta mRNA to very low levels (12% and 15% of control, respectively), and in cultures treated with both inhibin and CHX, this RNA was barely detectable. To determine the reversibility of the effects of these inhibitors, cells were incubated with fresh control medium after 6 h. Secretion of FSH and free alpha-subunit remained suppressed 4 h later; recovery was complete by 16 h in inhibin treated cultures. FSH-beta mRNA returned to control levels by 4 h in inhibin-treated and by 16 h in CHX-treated cultures. Levels of LH-beta and alpha-subunit mRNA were comparable to control values at all times. In conclusion, 1) CHX, like inhibin, suppresses FSH-beta mRNA levels, although its actions are less rapid and less rapidly reversible; 2) inhibin requires ongoing protein synthesis for full expression of its inhibitory effects; 3) the synthesis and secretion of LH are much less sensitive to inhibition by either inhibin or CHX than are the synthesis and secretion of FSH; and 4) secretion of free alpha-subunit involves a labile protein(s).