The exact nature of the interaction(s) between cAMP and calcium-sensitive phospholipid-dependent protein kinase-C effector pathways is not well understood in many tissues, including the ovary. In the present work we have evaluated the ability of protein kinase-C to modulate receptor- and nonreceptor-mediated cAMP generation in acute suspension cultures of swine luteal cells. Cells were exposed to LH (1 μg/ml), forskolin (100 μM), cholera toxin (1 μg/ml), pertussis toxin (100 ng/ml), and/or phorbol ester [12-O-tetradecanoylphorbol-13-acetate (TPA)] for 0-90 min. TPA had no effect on basal cAMP accumulation, but increased (P < 0.05) LH-, forskolin-, and cholera toxin-activated cAMP formation, with maximal facilitation at 30, 45, and 60 min, respectively. This facilitative effect was robust, as it could be demonstrated in both the presence and absence of the phosphodiesterase inhibitor 3-isobutyl-1-methylxanthine (0.5 mM). TPA increased dose-dependent LH (0.1-1 μg/ml)-, forskolin (3-300 μM)-, and cholera toxin (0.3-10 μg/ml)-stimulated cAMP accumulation. TPA induced a dose-dependent (0.3-30 ng/ml) increase in cAMP accumulation when incubated with the half-maximally effective (ED50) and maximally effective doses of LH (0.8 and 1 μg/ml, respectively), forskolin (10 and 300 μM), and cholera toxin (0.2 and 3 μg/ml). TPA had an ED50 for this functional activation of 6.1 (67% confidence interval, 4.4-9.7) nM. The stimulatory effect of TPA could be mimicked by two synthetic diacylglycerols, 1,2-dioctanoylglycerol and 1-oleoyl-2-acetylglycerol, but not by inactive phorbol esters. In addition, TPA augmented the stimulatory effect of pertussis toxin when combined with maximally effective doses of LH, forskolin, and cholera toxin. The stimulatory action of TPA on cAMP production was limited to endogenous cellular adenylyl cyclase. Bacterially derived adenylyl cyclase toxin isolated from Bordetella pertussis resulted in a dose-dependent increase in cAMP formation over 60 min, which was not facilitated by phorbol ester. We conclude that stimulatory coupling exists between the calcium-dependent protein kinase-C and cAMP-generating systems in swine luteal cells. This stimulatory coupling is enacted in part at the levels of both the guanine binding and the catalytic subunits of adenylyl cyclase.
|Original language||English (US)|
|Number of pages||7|
|State||Published - 1989|
ASJC Scopus subject areas
- Endocrinology, Diabetes and Metabolism