ASSESSMENT OF THE MECHANISM BY WHICH PROLACTIN STIMULATES PROGESTERONE PRODUCTION BY EARLY CORPORA-LUTEA OF PIGS

Previously, we reported that administration of prolactin (PRL) during the early luteal phase ill sows increases plasma progesterone concentr ations. In the current study, we searched for the mechanisms by which PRL exerts this luteotrophic effect. The objectives of the study were (1) to examine the effect of PRL and/or low-density lipoproteins (LDL) on progesterone production by porcine luteal cells derived from early corpora lutea, and (2) to assess the ability of PRL to activate phosp hoinositide-specific phospholipase C (PI-PLC) and protein kinase C (PK C) in these luteal cells. Ovaries with early corpora lutea (day 1-2 of the oestrous cycle) were obtained from the slaughter-house. Progester one production by dispersed luteal cells was measured after treatment with PRL, phorbol 12-myristate 13-acetate or inhibitors of PKC in the presence or absence of LDL. LDL increased progesterone concentration i n the incubation medium (304.5 vs 178.6 ng/ml in control, P<0.05). PRL augmented LDL-stimulated progesterone secretion by luteal cells (to 4 16 ng/ml, P<0.05), but PRL alone did not affect progesterone productio n (209.6 ng/ml, P>0.05). Staurosporine, a PKC inhibitor, inhibited pro gesterone secretion stimulated by the combined action of LDL and PRL; however, such inhibition was not demonstrated when cells were treated with the PKC inhibitor, H-7. PKC activation was assessed by measuring the specific association of [H-3]phorbol dibutyrate (H-3-PDBu) with lu teal cells after treatment with PRL or ionomycin (a positive control). PRL and ionomycin increased H-3-PDBu-specific binding in early luteal cells by 28 +/- 5.5% (within 5 min) and 70.2 +/- 19.3% (within 2 min) over control binding respectively (P<0.05). In addition, PRL did not augment the LDL-stimulated progesterone production in PKC-deficient ce lls. In contrast with PKC, total inositol phosphate accumulation, as w ell as intracellular free calcium concentrations, were not affected by PRL in the current study. We conclude that PRL, in the presence of LD L, stimulates progesterone production by early corpora lutea in vitro. Moreover, PRL appears to activate PKC, but not PI-PLC, in these cells . Thus intracellular transduction of the PRL signal may involve activa tion of PKC that is not dependent on PI-PLC.