Prostaglandin F2 alpha inhibition of epinephrine stimulated cyclic AMP and progesterone production by rat corpora lutea of various ages

Epinephrine can mimic the stimulatory effects of LH in vitro on cyclic AMP (cAMP) and progesterone production by isolated rat corpora lutea. The aim of the present study was to test whether the effects of epinephrine in vitro on the rat corpus luteum, as with LH, can be inhibited by prostaglandin F2 alpha (PGF2 alpha). The stimulatory effect of epinephrine on tissue levels of cAMP in 1-day-old corpora lutea was not inhibited by PGF2 alpha. A dose-dependent inhibition by PGF2 alpha (0.5-50 microM) was seen for 3-day-old corpora lutea and this inhibition could not be overcome by higher concentrations of epinephrine (0.165-165 microM). The stimulation by epinephrine on progesterone production was inhibited by PGF2 alpha (5 microM) in 3- and 5-day-old, but not in 1-day-old corpora lutea. Thus, PGF2 alpha can inhibit the stimulatory effect of epinephrine in 3- and 5-day-old corpora lutea, but not in the newly formed corpora lutea (1-day-old) and PGF2 alpha shows in this respect the same age dependent inhibitory pattern as in relation to LH stimulation.     

Occurrence and properties of a prostaglandin F2alpha receptor in bovine corpora lutea.

Prostaglandin F2alpha was specifically bound by a particulate fraction from bovine corpora lutea. The rate constants for the association (7.5 X 10(3) M-1 S-1) and dissociation (2.1 X 10-4 S-1) reactions gave a dissociation constant of 2.8 X 10(-8) M which is similar to that determined from a Scatchard plot of binding data at equilibrium (5 X 10(-8) M). The receptor was stable for several hours at 23 degrees C but was rapidly destroyed at 37 degrees C. The pH optimum for the binding reaction was 6.3. The receptor had high specificity for prostaglandin F2alpha and had much lower affinities for other prostaglandins. Luteinizing and follicle-stimulating hormones had no effect on the prostaglandin F2alpha-receptor interaction.     

Inhibition by vanadate of cyclic AMP production in rat corpora lutea incubated in vitro

Vanadate, a normal constituent of cells, has been reported to affect a variety of enzymes involved in phosphate transfer; the findings regarding adenylate cycle vary with the tissue and experimental system. In the corpus luteum, cyclic AMP (cAMP) stimulates steroidogenesis; and prostaglandin F2 alpha, which induces luteal regression, inhibits luteinizing hormone (LH)-induced cAMP accumulation. We examined the influence of orthovanadate on cAMP concentration in isolated corpora lutea from pseudopregnant rats. With 2 mM vanadate, basal cAMP level was unaffected, but LH-induced cAMP accumulation was inhibited by 45-68%. Lower doses of vanadate (0.2-1 mM) were almost as effective. When added simultaneously with LH, vanadate was inhibitory within 25 min, but no inhibition occurred when vanadate was added for 30 min to tissue pretreated with LH for 60 min. The decrease in cAMP accumulation was observed also when corpora lutea were exposed to vanadate in the presence of the phosphodiesterase inhibitor 3-isobutyl-1-methylxanthine (0.5 mM), indicating that vanadate inhibits cAMP synthesis. Vanadate may increase cytosolic calcium by inhibiting ion pumps in cell membranes. Thus, we examined the effect of vanadate in corpora lutea incubated in calcium-depleted medium and found that vanadate still inhibited cAMP formation. Vanadyl sulfate (0.4 and 2 mM) reduced the LH-induced cAMP accumulation as effectively as vanadate. Thus, the use of vanadate as a tool for exploring physiological regulators of luteal adenylate cyclase should be considered.     

Effect of prostaglandin F2 alpha on the ultrastructure and function of sheep corpora lutea.

Early morphological changes in the ultrastructure of CL of ewes treated with prostaglandin F2alpha were examined in relation to luteal function as judged by plasma progesterone concentration. The luteolytic effect of prostaglandin F2alpha was confirmed, but there was little synchrony between morphological and functional luteolysis. Significant changes included a decrease in the amount of smooth endoplasmic reticulum, a change in the shape of mitochondria and a decrease in the number of membrane-bound granules. There was also an accumulation of lipids.     

Luteinizing hormone receptors and progesterone content in porcine corpora lutea after prostaglandin F2 alpha

The effect of prostaglandin F2 alpha (PGF2 alpha) on luteinizing hormone (LH) receptors, weight and progesterone content of corpora lutea (CL), and serum progesterone concentrations was studied in gilts. Fifteen gilts were hysterectomized between Days 9 to 11 of the estrous cycle. Twelve gilts were injected i.m. with 10 mg of PGF2 alpha and 3 with saline on Day 20. Ovaries were surgically removed from each of 3 gilts at 4, 8, 12 and 24 h following PGF2 alpha treatment and from the 3 control gilts 12 h following saline injection. Jugular blood samples for progesterone analysis were collected from all gilts at 0, 2 and 4 h following treatment and at 8, 12 and 24 h for gilts from which ovaries were removed at 8, 12 and 24 h, respectively. Mean serum progesterone and CL progesterone concentrations decreased within 4 h after PGF2 alpha treatment (P less than 0.05) and remained low through 24 h after treatment. The number of unoccupied LH receptors decreased by 4 h (P less than 0.05) and this trend continued through 24 h. There were no differences in luteal weight or affinity of unoccupied LH receptors of luteal tissue at 4, 8 12 and 24 h after PGF2 alpha when compared to luteal tissue from controls. These data indicate that during PGF2 alpha-induced luteolysis in the pig, luteal progesterone, serum progesterone concentrations and the number of LH receptors decrease simultaneously.     

Cyclic AMP formation of isolated human corpora lutea in response to HCG - interference by PGF2 alpha.

Human corpora lutea of defined ages were excised at operation, cut into pieces and incubated in the presence of HCG, PGF2 alpha and PGE2 alone or in combination. Following incubation cAMP formation in tissue and medium was determined. HCG-stimulated tissue cAMP content was most pronounced at a corpus luteum age of 7-10 days after ovulation. This stimulation was antagonized by PGF2 alpha in corpora lutea older than 6 days. PGE2 stimulated cAMP formation per se and this effect was more pronounced when HCG and PGE2 were combined. A possible role for PGF2 alpha as a luteolytic substance in the human is suggested.     

Synthesis of prostaglandin F2 alpha, E2 and prostacyclin in isolated corpora lutea of adult pseudopregnant rats throughout the luteal life-span.

The ability of de novo biosynthesis of prostaglandins (PGs) in individual whole corpora lutea (CL) obtained from sterile-mated adult pseudopregnant rats on different days of the luteal phase and the post-luteolytic period was evaluated. Production of PGs, progesterone and 20 alpha-dihydroprogesterone were determined after in vitro incubation of CL extirpated from Day 2 to Day 19 after mating. A time-relationship with increased accumulation of PGs in the medium was demonstrated from 18 s to 5 h, with large increments during the first 30 min. Basal accumulation of PGs in the incubation medium was highest for 6-keto-PGF1 alpha (the stable metabolite of prostacyclin) greater than PGE2 greater than PGF2 alpha greater than thromboxane B2 (TXB2) and basal accumulation of PGF2 alpha and PGE2 measured in the medium was maximal on Day 10-11 of pseudopregnancy, concomitantly with a decline in secretion of progesterone. Addition of arachidonic acid (AA) dose-dependently increased synthesis of PGs, with absolute amounts of PGE2 greater than 6-keto-PGF1 alpha greater than PGF2 alpha greater than TXB2 and addition of 14 microM indomethacin markedly inhibited accumulation of all PGs measured. Luteinizing hormone (LH, 10 micrograms/ml) stimulated progesterone secretion on all days during pseudopregnancy, but not on the post-luteolytic Day 19. LH increased PGF2 alpha, PGE2 and 6-keto-PGF1 alpha secretion on Day 13 of pseudopregnancy by 76%, 91% and 28%, respectively, but not on the other days tested. Furthermore, stimulation of PG-synthesis by addition of AA abrogated the LH-induced progesterone accumulation markedly, but only on Day 13 of pseudopregnancy. Epinephrine (5 micrograms/ml) increased production of progesterone and also PGs, but only on Day 2 of pseudopregnancy, whereas oxytocin (100 mIU/ml) was found to be without effect on progesterone as well as PG secretion on all days tested. The results of the present study demonstrates the independent ability of the rat CL to synthesize PGG/PGH2-derived prostaglandins, including the putative luteolysin PGF2 alpha. Secondly, we demonstrate that LH and AA-induced increases in PGF2 alpha and PGE2 production during the luteolytic period, may be an autocrine or paracrine mechanism involved in luteolysis.     

Prostaglandin F2alpha specific binding in equine corpora lutea

Preliminary studies indicate the presence of PGF2alpha specific binding sites in membrane fractions prepared from equine corpora lutea. The equilibrium binding data indicate an apparent dissociation constant of 3.2 X 10(-9)M and the concentration of binding sites of -0.1 pmoles/mg membrane protein. Competition of several natural prostaglandins for equine luteal PGF2alpha specific binding sites indicates specificity for the 9alpha-hydroxyl moiety and the 5,6-cis doublebond. Significant increases in relative binding affinities were demonstrated for PGF2alpha analogs with a phenyl ring introduced at carbons 16 or 17. Specific PGF2alpha binding was demonstrated in corpora lutea collected at known stages of the estrous cycle. There was no pattern in these values based on the stage of the cycle. While specific 3H-PGE1 binding could be demonstrated, no high affinity sites could be quantitated. 3H-PGE1 binding appeared unaffected by changes in temperature or time of incubation, whereas PGF2alpha specific binding was significantly modified by both these factors.     

The presence of discrete receptors for prostaglandin F2 alpha in the cell membranes of bovine corpora lutea.

The cell membranes isolated from bovine corpora lutea bound ³H-prostaglandin (PG) F₂α with high affinity and specificity. The specific binding of ³H-PGF₂α was detectable at 10^?10M added ³H-PGF₂α and reached saturation at 10^?7M to 10^?6M. Unlabeled PGF₂α, as low as 10^?9M, inhibited the binding of ³H-PGF₂α with complete inhibition occurring at 10^?6M. The Scatchard analysis of equilibrium binding data revealed that the PGF₂α receptors are heterogeneous: Kd₁?5.1 × 10^?9M, n?289 fmoles/mg protein; Kd₂?1.8 × 10^?8M, n?780 fmoles/mg protein. The relative affinities of various other PGs for binding to PGF₂α receptors were (PGF₂α?100%): PGF₁α?17.5; PGE₁?0.8; PGE₂?22.4; PGA₁?0.007; PGB₁?0.01. The specificity and affinity of ³H-PGF₂α binding is consistent with the possibility that this receptor interaction may reflect an initial event in the action of PGF₂α as a luteolytic agent.     

Luteolytic effects of prostaglandin F2alpha on day 8 to 19 corpora lutea in the bitch

Luteolysis was induced in 5 experimental Beagle (8 cycles) and 7 client-owned bitches treated with 150 to 200 microg/kg, sc of prostaglandin F2alpha administered twice daily for 4 d, starting on Days 8 to 19 after the onset of cytological diestrus. Five experimental Beagle bitches had been mated during the estrus preceding treatment, and copulation had been confirmed in 2/7 client-owned bitches presented for termination of unwanted pregnancy. Serum progesterone concentration (mean +/- SD) declined from 26.1 +/- 66 ng/ml before treatment to 0.3 +/- 0.4 ng/ml on the fourth day of treatment One of the 7 client-owned bitches maintained her pregnancy even though serum progesterone concentrations were less than 0.5 ng/ml on the third and fourth day of treatment. Mean (+/- SEM) inter-estrous intervals before and following prostaglandin-induced luteolysis were 207.3 +/- 12.4 (n = 11 cycles in 6 bitches) and 95.5 +/- 20.0 d (n = 6 cycles in the same 6 bitches; P < 0.0001), respectively These results suggest that effective prostaglandin-induced luteolysis can be achieved with administration of 180 microg/kg during the third week of diestrus in pregnant and nonpregnant bitches.     

Receptors for prostaglandins F2 alpha and E2 in ovine corpora lutea during maternal recognition of pregnancy.

Plasma membrane receptors for prostaglandins (PG) F2 alpha and E2 were quantified in ovine corpora lutea obtained from nonpregnant and pregnant ewes on Days 10, 13, and 15 post-estrus, and from additional ewes on Days 25 and 40 of pregnancy. Regardless of reproductive status or day post-estrus, concentrations of luteal receptors for PGF2 alpha were 7- to 10-fold greater than those for PGE2. In pregnant ewes the concentration of receptors for PGF2 alpha was highest on Day 10 (35.4 +/- 2.8 fmol/mg) and lowest on Day 25 (22.3 +/- 2.5 fmol/mg). A difference in the concentration of luteal receptors for PGF2 alpha between pregnant and nonpregnant ewes was apparent only on Day 15 post-estrus, at which time the concentration of receptors for PGF2 alpha was higher in pregnant ewes than in nonpregnant ewes (27.1 +/- 2.7 vs. 17.7 +/- 2.7 fmol/mg). Concentrations of receptors for PGE2 in pregnant ewes were similar (p > 0.05; 2.8 +/- 0.3 to 3.7 +/- 0.2 fmol/mg) between Days 13 and 40 but were higher (p < 0.05) than in corpora lutea obtained from nonpregnant ewes on Days 10 (5.0 +/- 0.4 vs. 4.1 +/- 0.2 fmol/mg) and 15 (3.7 +/- 0.2 vs. 2.0 +/- 0.4 fmol/mg) post-estrus. Although concentrations of receptors for both PGF2 alpha and PGE2 were lowest in corpora lutea obtained from nonpregnant ewes on Day 15, this was not due to luteal regression since the weights and concentrations of progesterone in corpora lutea on Day 15 were not lower than those for corpora lutea obtained on Days 10 and 13.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of prostaglandin E2 alpha on the hCG-stimulated progesterone production by human corpora lutea

The effect of prostaglandin PGF2 alpha on the hCG stimulated and basal progesterone production by human corpora lutea was examined in vitro. hCG (40 i.u./ml) stimulated progesterone formation in corpora lutea of early (days 16-19 of a normal 28 day cycle), mid (days 20-22) and late (days 23-27) luteal phases. This stimulation was inhibited by PGF2 alpha (10 micrograms/ml) in corpora lutea of mid and late luteal phases. PGF2 alpha alone did not show a consistent effect on basal progesterone production. The inhibition of hCG stimulated progesterone production by PGF2 alpha at times corresponding to luteolysis indicates a role for that prostaglandin in the process of luteolysis in the human corpus luteum.