Stimulation by phorbol ester and diacylglycerol of luteinizing hormone glycosylation and release by rat anterior pituitary cells.

We studied the effects of protein kinase C (PKC) activators on LH glycosylation and release and the effect of 17 beta-estradiol on PKC activator-induced LH release. Rat anterior pituitary cells were incubated for 4 h with diluent, GnRH, and the PKC activators, phorbol 12-myristate 13-acetate (PMA), L-alpha-1,2-dioctanoyl glycerol (C8), and 1-oleoyl-2-acetyl-glycerol. LH translation and glycosylation were monitored by measuring incorporation of [14C]alanine ([14C]A) and [3H]glucosamine ([3H]GA), respectively, into total (medium + cell) immunoprecipitable LH. Immunoreactive LH (IRLH) was measured by RIA. PMA (10(-9) M) and 1-oleoyl-2-acetyl-glycerol (50-200 microM) had no significant effects. PMA at 10(-7) M elevated (P less than 0.01) medium IRLH, medium and total [3H]GA-LH, and medium but not total [14C]A-LH. PMA at 10(-7) M increased (P less than 0.01) uptake and incorporation of [3H]GA, but not [14C]A, into total pituitary protein. C8 increased both medium IRLH and total [3H]GA-LH (P less than 0.01) without altering total [14C]A-LH. Two hundred micromolar C8 increased medium concentrations of [3H]GA-LH (P less than 0.01) and [14C]A-LH (P less than 0.05). C8 (50-200 microM) had no detectable effects on uptake and incorporation of precursors into protein. GnRH (1 nM) enhanced (P less than 0.01) both medium IRLH and total [3H]GA-LH, but had no effect on total [14C]A-LH. Pretreatment of pituitary cells with 17 beta-estradiol (6 X 10(-10) M) greatly enhanced LH release induced by C8. In conclusion, PMA and C8, like GnRH, stimulated both LH glycosylation and release. These results suggest that PKC may regulate both LH release and glycosylation and may be important in estrogen modulation of LH release.