Activation of protein kinase C is coupled to prostaglandin E₂ synthesis in swine granulosa cells

We have examined the role of phospholipid-sensitive calcium-dependent protein kinase (protein kinase C) in prostaglandin E₂ synthesis by monolayer cultures of swine granulosa cells. Specific phorbol ester derivatives known to activate protein kinase C significantly augmented the production of prostaglandin E₂. These stimulatory actions were dose and time-dependent, and could be abolished by the cyclooxygenase inhibitor, indomethacin, or the protein synthesis inhibitor, cycloheximide. Moreover, the rank order of potency of phorbol esters in enhancing prostaglandin E₂ production was concordant with that demonstrated for activation of protein kinase C. Phorbol ester in conjunction with the divalent cation ionophore, A23187, increased prostaglandin E₂ production synergistically. In addition, a non-phorbol stimulator of protein kinase C, 1-octanoyl-2-acetylglycerol, also significantly enhanced prostaglandin E₂ biosynthesis. The stimulated synthesis of prostaglandin E₂ was confirmed by high-pressure liquid chromatographic purification of this radiolabeled metabolite of ³H-arachidonic acid, and by capillary gas chromatography high-resolution mass spectrometry. Thus, the present studies indicate that the protein kinase C effector pathway is functionally coupled to prostaglandin E₂ production in the swine granulosa cell.