Phospholipase C-beta and ovarian sex steroids in pig granulosa cells.
We compared the membrane effects of estradiol, progesterone, and androstenedione in a single experimental model, the ovarian granulosa cells collected from immature Large White sows. We measured changes in cytosolic free calcium concentration ([Ca2+]i) in confluent Fura-2 loaded cells. We used pharmacological tools and polyclonal phospholipase C-beta (PLC-beta) antibodies. Each steroid (0.1 pM to 1 nM) transiently increased intracellular calcium concentration ([Ca2+]i) within 5 sec. They mobilized Ca2+ from the endoplasmic reticulum as shown by using two phospholipase C inhibitors, neomycin and U-73122. Ca2+ mobilization involved PLC-beta1 for progesterone, PLC-beta2 for estradiol and PLC-beta4 for androstenedione. A pertussis toxin-insensitive G protein was involved in the effects of progesterone on Ca2+ mobilization whereas estradiol and androstenedione effects were mediated via a pertussis toxin-sensitive G-protein. Ca2+ influx from the extracellular milieu was involved in the increase in [Ca2+]i induced by progesterone and estradiol, but not by androstenedione. Influx of Ca2+ was independent of Ca2+ mobilization from calcium stores, and it was suggested that L-type Ca2+ channels for estradiol and T-type Ca2+ channels for progesterone were involved. The three steroids had no effect on cAMP. Rapid effects of progesterone, estradiol, and androstenedione involved a direct action on cell membrane elements such as PLC-beta, G-proteins, and calcium channels, and these mechanisms were hormone-specific.
Lieberherr
M
M
The Centre National de la Recherche Scientifique, Jouy en Josas, France.
Grosse
B
B
Machelon
V
V
D000818:Animals; D015153:Blotting, Western; D002118:Calcium / Q000378:metabolism; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism; D000242:Cyclic AMP / Q000378:metabolism; D004721:Endoplasmic Reticulum / Q000378:metabolism; D005260:Female; D012739:Gonadal Steroid Hormones / Q000494:pharmacology*; D006107:Granulosa Cells / Q000378:metabolism*; D007527:Isoenzymes / Q000378:metabolism*; D010053:Ovary / Q000378:metabolism*; D037342:Pertussis Toxin; D043265:Phosphatidylinositol Diacylglycerol-Lyase; D054799:Phospholipase C beta; D013552:Swine; D010738:Type C Phospholipases / Q000378:metabolism*; D010566:Virulence Factors, Bordetella / Q000494:pharmacology
D012739:Gonadal Steroid Hormones; D007527:Isoenzymes; D010566:Virulence Factors, Bordetella; D000242:Cyclic AMP; D037342:Pertussis Toxin; D010738:Type C Phospholipases; D054799:Phospholipase C beta; D043265:Phosphatidylinositol Diacylglycerol-Lyase; D002118:Calcium