Effects of indomethacin, luteinizing hormone (LH), prostaglandin E(2) (PGE(2)), trilostane, mifepristone, ethamoxytriphetol (MER-25) on secretion of prostaglandin E (PGE), prostaglandin F(2alpha) (PGF(2alpha)) and progesterone by ovine corpora lutea of pregnancy or the estrous cycle

Two experiments were conducted to determine the luteotropin of pregnancy in sheep and to examine autocrine and paracrine roles of progesterone and estradiol-17 beta on progesterone secretion by the ovine corpus luteum (CL). Secretion of progesterone per unit mass by day-8 or day-11 CL of the estrous cycle was similar to day-90 CL of pregnancy (P >/= 0.05). In experiment 1, secretion of progesterone in vitro by slices of CL from ewes on day-8 of the estrous cycle was increased (P /= 0.05) while PGE(2) increased (P /= 0.05) detectable quantities of PGF(2alpha) or PGE while day-90 ovine CL of pregnancy secreted PGE (P /= 0.05). Trilostane, mifepristone, or MER-25 did not affect secretion of progesterone, PGE, or PGF(2alpha) by day-11 CL of the estrous cycle or day-90 CL of pregnancy (P >/= 0.05). It is concluded that PGE(2), not LH, is the luteotropin at day-90 of pregnancy in sheep and that progesterone does not modify the response to luteotropins. Thus, we found no evidence for an autocrine or paracrine role for progesterone or estradiol-17 36 on luteal secretion of progesterone, PGE or PGF(2alpha).     

Effects of prostaglandins E2 and F2alpha (PGE2; PGF2alpha), trilostane,mifepristone, palmitic acid (PA), indomethacin (INDO), ethamoxytriphetol (MER-25), PGE2 + PA,or PGF2alpha + PA on PGE2, PGF2alpha,and progesterone secretion by bovine corpora lutea of mid-pregnancy in vitro

The effects of PGE2, PGF2alpha, trilostane, RU-486, PA, INDO, MER-25, PGE2, or PGF2alpha + PA on secretion of progesterone, PGE2, or PGF2alpha by bovine corpora lutea (CL) of mid-pregnancy in vitro for 4 and 8 hr was examined. Secretion of PGE2 and PGF2alpha increased with time in culture (P < or = 0.05). PGE2 and PGE2 + PA increased (P < or = 0.05) secretion of progesterone at 4 and 8 h, progesterone secretion was increased (P < or = 0.05) at 4 h; but not at 8 h (P > or = 0.05) by trilostane, mifepristone, PGF2alpha and PGF2alpha + PA, and was decreased at 8 h by PGF2alpha and PGF2alpha + PA. Indomethacin decreased (P < or = 0.05) secretion of PGE2, PGF2alpha, and progesterone at 4 and 8 h. Trilostane, PA, PGF2alpha, RU-486 and PGF2alpha + PA increased (P < or = 0.05) PGE2 at 4 h only. Palmitic acid decreased (P < or = 0.05) PGF2alpha at 4 h, while trilostane, RU-486, or MER-25 did not affect (P < or = 0.05) PGE2 of PGF2alpha secretion. It is concluded that PGE2 of luteal tissue origin is the luteotropin at mid-pregnancy in cows. Also, it is suggested that PA may alter progesterone secretion by affecting the inter conversion of PGE2 and PGF2alpha.     

Effect of luteinizing hormone (LH), pregnancy-specific protein B (PSPB), or arachidonic acid (AA) on secretion of progesterone and prostaglandins (PG) E (PGE; PGE(1) and PGE(2)) and F(2alpha) (PGF(2alpha)) by ovine corpora lutea of the estrous cycle or pregnancy in vitro

LH regulates luteal progesterone secretion during the estrous cycle in ewes and cows. However, PGE, not LH, stimulated ovine luteal progesterone secretion in vitro at day 90 of pregnancy and at day 200 in cows. The hypophysis is not obligatory after day 50 nor the ovaries after day 55 to maintain pregnancy in ewes. LH has been reported to regulate ovine placental PGE secretion up to day 50 of pregnancy and by pregnancy-specific protein B (PSPB) after day 50 of pregnancy. The objective of this experiment was to determine if and when a switch from LH to PGE occurred as the luteotropin regulating luteal progesterone secretion during pregnancy in ewes. Ovine luteal tissue slices of the estrous cycle (days 8, 11, 13, and 15) or pregnancy (days 8, 11, 13, 15, 20, 30, 40, 50, 60, and 90) were incubated in vitro with vehicle, LH, AA (precursor to PGE(2) and PGF(2alpha) synthesis), or PSPB in M199 for 4 h and 8 h. Concentrations of progesterone in jugular venous plasma of bred ewes increased (P< or =0.05) after day 50 and continued to increase through day 90. Secretion of progesterone by luteal tissue of non-bred ewes on days 8, 11, 13 and 15 and by bred ewes on days 8, 11, 13, 15, 20, 30, 40, and 50 was increased (P< or =0.05) by LH, but not by luteal tissue from pregnant ewes after day 50 (P> or =0.05). LH-stimulated progesterone secretion by luteal tissue from day 15 bred ewes was greater (P< or =0.05) than day 15 luteal tissue from non-bred ewes. Concentrations of progesterone in media were increased (P< or =0.05) when luteal tissue from pregnant ewes on day 50, 60, or 90 were incubated with AA or PSPB. Concentrations of PGE in media of non-bred ewes on days 8, 11, 13, or 15 and bred ewes on days 8 and 11 did not differ (P> or =0.05). Concentrations of PGE were increased (P< or =0.05) in media by luteal slices from bred ewes on days 13, 15, 20, 30, 40, 50, 60, and 90 of vehicle, LH, AA or PSPB-treated ewes. In addition, PSPB increased (P< or =0.05) PGE in media by luteal slices from pregnant ewes only on days 40, 50, 60, and 90. Concentrations of PGF(2alpha) were increased in media (P<0.05) of vehicle, AA, LH, or PSPB-treated luteal tissue from non-bred ewes and bred ewes on day 15 and by luteal tissue from bred ewes on days 20 and 30 after which concentrations of PGF(2alpha) in media declined (P< or =0.05) and did not differ (P> or =0.05) from non-bred or bred ewes on days 8, 11, or 13. It is concluded that LH regulates luteal progesterone secretion during the estrous cycle of non-bred ewes and up to day 50 of pregnancy, while only PGE regulates luteal progresterone secretion by ovine corpora lutea from days 50 to 90 of pregnancy. In addition, PSPB appears to regulate luteal secretion of progesterone from days 50 to 90 of pregnancy through stimulation of PGE secretion by ovine luteal tissue.     

Effect of luteinizing hormone (LH), pregnancy specific protein B (PSPB), or arachidonic acid (AA) on ovine endometrium of the estrous cycle or placental secretion of prostaglandins E2 (PGE2) and F2alpha (PGF2alpha) and progesterone in vitro

The objective of this experiment was to determine the effect of AA, LH, or PSPB on secretion of PGE2, PGF2alpha, or progesterone by ovine caruncular endometrium of the estrous cycle or placental tissue of pregnancy in vitro. Ovine caruncular endometrium of the estrous cycle (days 8, 11, 13, and 15) or caruncular/placental tissue on days 8, 11, 13, 15, 20, 30, 40, 50, 60, and 90 postbreeding were incubated in vitro with vehicle, AA, LH, or PSPB in M-199 for 4 and 8 h. Secretion of PGF2alpha by caruncular endometrium of non-bred ewes on days 13 and 15 and by caruncular/placental tissue of bred ewes on days 13, 15, 20, 30, and 40 was increased (P < or = 0.05) when incubated with vehicle and declined (P < or = 0.05) after day-40 in bred ewes. Secretion of PGF2alpha by day-15 caruncular endometrium of non-bred ewes and bred ewes was increased (P < or = 0.05) by AA on days 13 and 15 and by LH on day-15. Secretion of PGF2alpha by caruncular/placental tissue from bred ewes was (P < or = 0.05) by AA on days 13, 15, 20, 30, and 40 and by LH on days 15, 20, 30, and 40, after which the response decreased (P < or = 0.05). Secretion of PGF2alpha by caruncular endometrium of non-bred ewes during the estrous cycle or by caruncular/placental tissue of bred ewes during the first trimester was not affected by PSPB (P > or = 0.05). Secretion of PGE2 by caruncular endometrium of non-bred ewes did not change (P > or = 0.05) and was increased (P < or = 0.05) by caruncular/placental tissue on days 13-90 from bred ewes when incubated with vehicle. Secretion of PGE2 by endometrium from non-bred ewes was not affected (P > or = 0.05) by AA, LH, or PSPB, but was increased (P < or = 0.05) by AA or LH on days 13-50 and by PSPB on days 60 and 90 when incubated with caruncular/placental tissue from bred ewes. Secretion of progesterone by placental tissue of bred ewes increased (P < or = 0.05) on day-50 and continued to increase through day-90. In summary, uterine/placental tissue secretion of PGF2alpha is not reduced until the end of the first trimester of pregnancy in ewes. In addition, LH appears to play a role in luteolysis of non-bred ewes by stimulating caruncular endometrial secretion of PGF2alpha and on day-5 postbreeding to prevent luteolysis during early pregnancy by stimulating caruncular/placental secretion of PGE2 throughout the first trimester of pregnancy in sheep. Secretion of PGE2 by caruncular/placental tissue after day-50 of pregnancy appears to be regulated by PSPB, not LH.     

Effect of prostaglandin F2alpha (PGF2alpha), indomethacin, tamoxifen or estradiol-17beta on prostaglandin E (PGE), PGF2alpha and estradiol-17beta secretion in 88 to 90-day pregnant sheep

Treatment with PGF2alpha plus estradiol-17beta aborts 90-day pregnant ewes, whereas PGF2alpha or estradiol-17beta alone does not abort ewes. The objective of this experiment was to evaluate whether tamoxifen, an estrogen receptor antagonist, estradiol-17beta, prostaglandin F2alpha (PGF2alpha), indomethacin, or some of their interactions affected ovine uterine/placental secretion of PGF2alpha, estradiol-17beta or prostaglandins E (PGE), because a single treatment with PGF2alpha and estradiol-17beta given every 6 h aborts 90-day pregnant ewes. Concentrations of PGF2alpha in uterine venous blood were increased (P < or = 0.05) by estradiol-17beta, PGF2alpha + estradiol-17beta, and PGF2alpha + tamoxifen, and decreased (P < or = 0.05) by indomethacin or PGF2alpha + indomethacin at 72 h when compared to the 0 h samples. Concentrations of PGE in uterine venous blood were decreased (P < or = 0.05) by indomethacin and PGF2alpha + indomethacin and increased (P < or = 0.05) by PGF2alpha + estradiol-17beta at 72 h when compared to the 0 h samples. Concentrations of PGF2alpha in inferior vena cava blood at 6 h were increased (P < or = 0.05) by PGF2alpha either alone or in combination with indomethacin, tamoxifen, or estradiol-17beta, which is due to the PGF2alpha injected. Concentrations of PGF2alpha in inferior vena cava blood in PGF2alpha + estradiol-17beta-treated 88- to 90-day pregnant ewes increased (P < or = 0.05) linearly over the 72-h sampling period and averaged 4.0 + 0.4 ng/ml. Concentrations of PGF2alpha in inferior vena cava blood of control, PGF2alpha, tamoxifen, PGF2alpha + indomethacin, PGF2alpha + tamoxifen, and estradiol-17beta-treated ewes did not differ (P > or = 0.05) and averaged 0.4 + 0.04 ng/ml. Profiles of PGE in inferior vena cava blood of 88- to 90-day pregnant ewes treated with vehicle, PGF2alpha, estradiol-17beta, tamoxifen, tamoxifen + PGF2alpha, or estradiol-17beta + PGF2alpha did not differ (P > or = 0.05). Concentrations of PGE in inferior vena cava blood of 88- to 90-day pregnant ewes treated with indomethacin or PGF2alpha + indomethacin were lower (P < or = 0.05) than in control ewes. Concentrations of estradiol-17beta in jugular venous plasma of PGF2alpha + estradiol-17beta-treated 88- to 90-day pregnant ewes increased linearly and differed (P < or = 0.05) from controls. Profiles of estradiol-17beta in jugular venous plasma of PGF2alpha, indomethacin, tamoxifen, and PGF2alpha + tamoxifen and PGF2alpha + indomethacin, estradiol-17beta, and controls did not differ (P > or = 0.05). It is concluded that treatment with a single injection of PGF2alpha and estradiol-17beta given every 6 h causes a linear increase in PGF2alpha and estradiol-17beta.     

Effect of mifepristone on pregnancy,pregnancy-specific protein B (PSPB), progesterone,estradiol-17beta,prostaglandin F2alpha (PGF2alpha) and prostaglandin E (PGE) in ovariectomized 90-day pregnant ewes

The objective of this experiment was to determine the effect of mifepristone, a progesterone receptor antagonist, on pregnancy and secretion of steroids, pregnancy-specific protein B (PSPB) and prostaglandins at mid-pregnancy in ewes. Ninety-day pregnant ewes were ovariectomized (OVX) and treatments were initiated 72 h post-OVX. Ewes received (1) vehicle, (2) prostaglandin F2alpha (PGF2alpha, 8 mg/58 kg/bw, i.m.) 84 h post-OVX, (3) mifepristone (50 mg intrajugular at 72, 84, 96, and 108 h post-OVX), (4) mifepristone (50mg) + PGF2alpha, (5) mifepristone (100 mg intrajugular at 72, 84, 96, and 108 h), and (6) mifepristone (100 mg) + PGF2alpha. Ewes treated with vehicle or PGF2alpha alone did not abort (P > or = 0.05). But, 60, 80, 60, and 100% of ewes treated with mifepristone (50 mg), mifepristone (50 mg) + PGF2alpha, mifepristone (100 mg), and mifepristone (100 mg) + PGF2alpha, respectively, aborted (P < or = 0.05). Profiles of progesterone, estradiol-17beta, prostaglandin E (PGE), or PSPB did not differ (P > or = 0.05) among treatment groups. Profiles of PGF2alpha of treatment groups receiving mifepristone with or without PGF2alpha differed (P < 0.05) from vehicle or PGF2alpha alone-treated ewes. It is concluded that progesterone actions are necessary to suppress uterine/placental secretion of PGF2alpha and that maintenance of critical progesterone: estradiol-17beta and PGE:PGF2alpha ratios are necessary for maintenance of pregnancy.     

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.     

Effects of progesterone administered to dairy heifers on sensitivity of corpora lutea to PGF(2)alpha and on plasma LH concentration

To determine whether progesterone facilitates PGF(2)alpha-induced luteolysis prior to day 5 of the estrous cycle, 48 Holstein-Friestian heifers were assigned at random to four treatments: 1) 4 ml corn oil/day + 5 ml Tris-HCl buffer (control); 2) 25 mg prostaglandin F(2)alpha (PGF(2)alpha); 3) 100 mg progesterone/day (progesterone); 4) 100 mg progesterone/day + 25 mg PGF(2)alpha (combined treatment). Progesterone was injected subcutaneously daily from estrus (day 0) through day 3. The PGF(2)alpha was injected intramuscularly on day 3. Estrous cycle lengths were decreased by progesterone: 20.2 +/- 0.56, 19.2 +/- 0.31 (control and PGF(2)alpha); 13.2 +/- 1.40, and 11.7 +/- 1.27 (progesterone and combined). The combination of progesterone and PGF(2)alpha did not shorten the cycle any more than did progesterone alone (interaction, P>0.05). PGF(2)alpha treatment reduced progesterone concentrations on day 6 (P<0.05) and both progesterone and PGF(2)alpha reduced plasma progesterone on day 8 (P<0.01 and P<0.05, respectively). LH was measured in blood samples collected at 10- min intervals for 4 hr on day 4 from three heifers selected at random from each of the four treatment groups. Mean LH concentration for control heifers ranged from 0.35 to 0.63 ng/ml (overall mean, 0.49 ng/ml) and for progesterone-treated heifers ranged from 0.12 to 0.30 ng/ml (overall mean, 0.23 ng/ml). LH concentrations were greater in control heifers (P<0.01). The mean LH pulse rate for control heifers was 2.7 pulses/heifers/4 hr, while that for the progesterone-treated heifers was 1.7 pulses/heifer/4 hr. The mean pulse amplitude for control and progesterone treatments was 0.47 ng/ml and 0.36 ng/ml, respectively. Neither pulse amplitude nor frequency were different between treatment groups.     

Prostaglandin E1 (PGE1), but not prostaglandin E2 (PGE2), alters luteal and endometrial luteinizing hormone (LH) occupied and unoccupied LH receptors and mRNA for LH receptors in ovine luteal tissue to prevent luteolysis

Loss of luteal progesterone secretion at the end of the ovine estrous cycle is via uterine PGF₂α secretion. However, uterine PGF₂α secretion is not decreased during early pregnancy in ewes. Instead, the embryo imparts a resistance to PGF₂α. Prostaglandins E (PGE; PGE₁ + PGE₂) are increased in endometrium and uterine venous blood during early pregnancy in ewes to prevent luteolysis. Chronic intrauterine infusion of PGE₁ or PGE₂ prevents spontaneous or IUD, estradiol-17β, or PGF₂α-induced premature luteolysis in nonbred ewes. The objective was to determine whether chronic intrauterine infusion of PGE₁ or PGE₂ affected mRNA for LH receptors, occupied and unoccupied receptors for LH in luteal and caruncular endometrium, and luteal function. Ewes received Vehicle, PGE₁, or PGE₂ every 4 h from days 10 to 16 of the estrous cycle via a cathether installed in the uterine lumen ipsilateral to the luteal-containing ovary.Jugular venous blood was collected daily for analysis of progesterone and uterine venous blood was collected on day-16 for analysis of PGF₂α and PGE. Corpora lutea and caruncular endometrium were collected from day-10 preluteolytic control ewes and day-16 ewes treated with Vehicle, PGE₁ or PGE₂ for analysis of the mRNA for LH receptors and occupied and unoccupied receptors for LH. Luteal weights on day-16 in ewes treated with PGE₁ or PGE₂ and day-10 control ewes were similar (P ≥ 0.05), but were greater (P ≤ 0.05) than in day-16 Vehicle-treated ewes. Progesterone profiles on days 10–16 differed (P ≤ 0.05) among treatment groups: PGE₁ > PGE₂ > Vehicle-treated ewes. Concentrations of PGF₂α and PGE in uterine venous plasma on day-16 were similar (P ≥ 0.05) in the three treatment groups. Luteal mRNA for LH receptors and unoccupied and occupied LH receptors were similar (P ≥ 0.05) in day-10 control ewes and day-16 ewes treated with PGE₂ and were lower (P ≤ 0.05) in day-16 Vehicle-treated ewes. PGE₂ prevented loss (P ≤ 0.05) of day-16 luteal mRNA for LH receptors and occupied and unoccupied LH receptors. Luteal and caruncular tissue mRNA for LH receptors and occupied and unoccupied LH receptors were greater (P ≤ 0.05) on day-16 of PGE₁-treated ewes than any treatment group. mRNA for LH receptors and occupied and unoccupied receptors for LH in caruncules were greater (P ≤ 0.05) in day-16 Vehicle or PGE₂-treated ewes than in day-10 control ewes. It is concluded that PGE₁ and PGE₂ share some common mechanisms to prevent luteolysis; however, only PGE₁ increased luteal and endometrial mRNA for LH receptors and occupied and unoccupied LH receptors. PGE₂ prevents a decrease in luteal mRNA for LH receptors and occupied and unoccupied receptors for LH without altering endometrial mRNA for LH receptors or occupied and unoccupied receptors for LH.     

Growth hormone, but not luteinizing hormone, acts with luteal peptides on prostaglandin F2alpha and progesterone secretion by bovine corpora lutea in vitro

Prostaglandin F2alpha (PGF2alpha) is a major physiological luteolysin in the cow. However, injection of PGF2alpha before day 5 (day 0 = estrus) of the estrous cycle dose not induce luteolysis. On the other hand, the early corpus luteum (CL) actively produces PGF2alpha. This indicates that luteal PGF2alpha may play a key role in the refractoriness to PGF2alpha injected during the early luteal phase when angiogenesis is active in the CL. Thus, this study aimed to investigate the possible interaction between pituitary hormones and local factors (luteal peptides) on secretion of PGF2alpha and progesterone (P) by the early bovine CL, and to evaluate the effect of growth hormone (GH) as well as its interactions on production of PGF2alpha in the developing CL. A RT-PCR analysis revealed that mRNA for GH receptor in CL was fully expressed from early in the luteal phase throughout the estrous cycle, while luteinizing hormone (LH) receptor mRNA was expressed less by the early and regressing CL than those at mid or late luteal phases (P < 0.05). For the stimulation test, an in vitro microdialysis system (MDS) was used as a model. Each bovine early CL (days 3-4) was implanted with the MDS, and maintained in an organ culture chamber. The infusion of GH, insulin-like growth factor-I (IGF-I) and oxytocin (OT) increased (P < 0.05) PGF2alpha and P release. In contrast, LH had no effect (P > 0.05) on PGF2alpha secretion and little effect on P release. Unexpectedly, there was no distinct interaction between pituitary hormones and luteal peptides on secretion of PGF2alpha and P. These results indicate that GH is a more powerful stimulator of PGF2alpha and P production in the early bovine CL than LH and suggest that GH and luteal peptides, IGF-1 and OT, contribute to maintenance of elevated PGF2alpha production in the developing bovine CL.     

Growth hormone, but not luteinizing hormone, acts with luteal peptides on prostaglandin F2alpha and progesterone secretion by bovine corpora lutea in vitro*

Prostaglandin F2alpha (PGF2alpha) is a major physiological luteolysin in the cow. However, injection of PGF2alpha before day 5 (day 0 = estrus) of the estrous cycle dose not induce luteolysis. On the other hand, the early corpus luteum (CL) actively produces PGF2alpha. This indicates that luteal PGF2alpha may play a key role in the refractoriness to PGF2alpha injected during the early luteal phase when angiogenesis is active in the CL. Thus, this study aimed to investigate the possible interaction between pituitary hormones and local factors (luteal peptides) on secretion of PGF2alpha and progesterone (P) by the early bovine CL, and to evaluate the effect of growth hormone (GH) as well as its interactions on production of PGF2alpha in the developing CL. A RT-PCR analysis revealed that mRNA for GH receptor in CL was fully expressed from early in the luteal phase throughout the estrous cycle, while luteinizing hormone (LH) receptor mRNA was expressed less by the early and regressing CL than those at mid or late luteal phases (P < 0.05). For the stimulation test, an in vitro microdialysis system (MDS) was used as a model. Each bovine early CL (days 3-4) was implanted with the MDS, and maintained in an organ culture chamber. The infusion of GH, insulin-like growth factor-1 (IGF-1) and oxytocin (OT) increased (P < 0.05) PGF2alpha and P release. In contrast, LH had no effect (P > 0.05) on PGF2alpha secretion and little effect on P release. Unexpectedly, there was no distinct interaction between pituitary hormones and luteal peptides on secretion of PGF2alpha and P. These results indicate that GH is a more powerful stimulator of PGF2alpha and P production in the early bovine CL than LH and suggest that GH and luteal peptides, IGF-1 and OT, contribute to maintenance of elevated PGF2alpha production in the developing bovine CL.     

Effect of trilostane on PGE, PGF2alpha, estradiol-17beta, and progesterone secretion and pregnancy of 90-day ovariectomized pregnant ewes

Ninety-day pregnant sheep were ovariectomized and received vehicle or trilostane every 12 h through 132 h, starting at 72 h postovariectomy. All trilostane-treated ewes aborted (P < or = 0.05) between 36 and 50 h after initiation of treatment. Profiles of progesterone in jugular venous blood differed (P < or = 0.05) and was lower (P < or = 0.05) in trilostane-treated ewes. Profiles of estradiol-17beta in jugular venous plasma of trilostane-treated ewes differed (P < or = 0.05) from controls. Estradiol-17beta increased after the first two treatments, followed by a return 2 h later to pretreatment levels (P > or = 0.05), which was followed by a sustained increase (P < or = 0.05) in estradiol-17beta. Profiles of PGF2alpha in inferior vena cava plasma of trilostane-treated ewes differed and were greater (P < or = 0.05) and occurred with the sustained increase in estradiol-17beta and the onset of most of the abortions. Profiles of PGE in inferior vena cava plasma between control and trilostane-treated 90-day pregnant ewes did not differ (P > or = 0.05). It is concluded that abortions occur at midpregnancy in sheep when the estradiol-17beta : progesterone ratio changes sufficiently to cause a sustained increase in estradiol-17beta and PGF2alpha but without changing placental secretion of PGE.     

Effect of local interaction of reactive oxygen species with prostaglandin F(2alpha) on the release of progesterone in ovine corpora lutea in vivo

Prostaglandin (PG) F(2alpha) is implicated in the process of luteal regression in many species, and has been shown to increase the generation of reactive oxygen species. In this study, the role of reactive oxygen species in the local regulatory mechanisms of functional luteolysis in the ewe was examined. In Experiment 1, we studied local effects of hydrogen peroxide (H(2)O(2)) and its interaction with PGF(2alpha) on P secretion in ovine corpus luteum (CL) in vivo. For this purpose, a microdialysis system (MDS) was used, where only the cells surrounding the capillary membrane in the microenvironment of the CL are exposed to these factors, and the P secretory ability of the CL is maintained as if intact. The study used a multiple CL model to implant the MDS, enabling us to examine in parallel several experimental infusions into the MDS implanted in different CLs (one MDS line per CL) developed after superovulation in one ewe. On Day 8 after GnRH treatment, the MDS were implanted into multiple CL in both ovaries of six ewes. A 4-h infusion with PGF(2alpha) (10(-6)M) at 8-12 h slightly increased P release during infusion, while a 4-h infusion with H(2)O(2) (10(-3)M) at 20-24 h decreased P release at 27-38 h. A pre-infusion with PGF(2alpha) for 4h at 8-12h, followed by infusion of H(2)O(2) at 20-24 h rapidly decreased the P release at 20-40 h (P<0.05); this decrease occurred 7h earlier than in the CL treated with H(2)O(2) alone. In Experiment 2, by utilizing the MDS we also applied free radical scavengers to examine their possible weakening effect on the inhibition of P secretion in the microenvironment within the regressing CL induced by PGF(2alpha) treatment. On Day 8 after estrus, the MDS were implanted into the CL (single CL model, two MDS lines per CL). Infusion of free radical scavengers, superoxide dismutase (SOD;50mg/ml)+catalase (CAT; 10mg/ml), at 0-28 h first increased P release until 12 h (P<0.05), and consequently delayed the decrease in P release until 30 h after administration of PGF(2alpha) i.m. (P<0.05). The present results support the concept that the leading pathway from PGF(2alpha) induces an increase of reactive oxygen species in luteolysis in the ewe.     

Influence of luteinizing hormone and prostaglandin F(2alpha) on progesterone secretion in superfused minced bovine luteal tissue in the early stage of the estrous cycle

Minced luteal tissue of bovine corpora lutea from Day 4, 5, and 6 of the estrous cycle (n = 4 corpora lutea each) was superfused for 9 h, and the progesterone secretion under the influence of 100 ng luteinizing hormone (LH)/ml and/or 1,000 ng prostaglandin F(2alpha) (PGF(2alpha))/ml was determined. In vivo, this period of the estrous cycle is characterized by a transition from PGF(2alpha) refractoriness to PGF(2alpha) sensitivity. The investigations were carried out in order to examine whether this transition is reflected by a change in the hormone secretion pattern in vitro. The basal secretion was higher on Day 6 than on Day 4 and 5 (P < 0.01). PGF(2alpha) slightly increased the progesterone secretion, but there was no statistically significant difference (P > 0.05). LH, however, stimulated the progesterone secretion by about 30% in luteal tissue collected from Day 4 and 5 (P < 0.01). In luteal tissue collected from Day 6, the LH-induced increase in hormone secretion was not statistically significant due to two corpora lutea that showed no response at all to LH. The progesterone secretion of the two other corpora lutea, however, was increased by 30% (P < 0.01). When PGF(2alpha) and LH were simultaneously added, the LH-induced progesterone secretion was not inhibited; PGF(2alpha) even seemed to intensify the action of LH. The difference between the hormone secretion under the influence of LH alone and that under the influence of a combination of LH and PGF(2alpha), however, was not statistically significant. It is concluded that in cattle the end of the refractoriness to PGF(2alpha) in vivo is not reflected by a corresponding change of the hormone secretion pattern in vitro.     

In vivo intra-luteal implants of prostaglandin (PG) E(1) or E(2) (PGE(1), PGE(2)) prevent luteolysis in cows. I. Luteal weight, circulating progesterone, mRNA for luteal luteinizing hormone (LH) receptor, and occupied and unoccupied luteal receptors for LH

Previously, it was reported that chronic intra-uterine infusion of PGE(1) or PGE(2) every four hours inhibited luteolysis in ewes. However, estradiol-17β or PGE(2) given intra-uterine every 8h did not inhibit luteolysis in heifers, but infusion of estradiol+PGE(2) inhibited luteolysis in heifers. The objective of this experiment was to determine whether and how intra-luteal implants containing PGE(1) or PGE(2) prevent luteolysis in Angus or Brahman cows. On day-13 post-estrus, Angus cows received no intra-luteal implant and corpora lutea were retrieved or Angus and Brahman cows received intra-luteal silastic implants containing Vehicle, PGE(1), or PGE(2) and corpora lutea were retrieved on day-19. Coccygeal blood was collected daily for analysis for progesterone. Breed did not influence the effect of PGE(1) or PGE(2) on luteal mRNA for LH receptors or unoccupied or occupied luteal LH receptors did not differ (P>0.05) so the data were pooled. Luteal weights of Vehicle-treated Angus or Brahman cows from days-13-19 were lower (P<0.05) than those treated with intra-luteal implants containing PGE(1) or PGE(2). Day-13 Angus luteal weights were heavier (P<0.05) than Vehicle-treated Angus cows on day-19 and luteal weights of day-13 corpora lutea were similar (P>0.05) to Angus cows on day-19 treated with intra-luteal implants containing PGE(1) or PGE(2). Profiles of circulating progesterone in Angus or Brahman cows treated with intra-luteal implants containing PGE(1) or PGE(2) differed (P<0.05) from controls, but profiles of progesterone did not differ (P>0.05) between breeds or between cows treated with intra-luteal implants containing PGE(1) or PGE(2). Intra-luteal implants containing PGE(1) or PGE(2) prevented (P<0.05) loss of luteal mRNA for LH receptors and unoccupied or occupied receptors for LH compared to controls. It is concluded that PGE(1) or PGE(2) alone delays luteolysis regardless of breed. We also conclude that either PGE(1) or PGE(2) prevented luteolysis in cows by up-regulating expression of mRNA for LH receptors and by preventing loss of unoccupied and occupied LH receptors in luteal tissue.     

Role of progesterone in regulating uteroovarian venous concentrations of PGF2 alpha and PGE2 during the estrous cycle and early pregnancy in ewes

The role of progesterone in regulation of uteroovarian venous concentrations of prostaglandins F2 alpha(PGF2 alpha) and E2 (PGE2) during days 13 to 16 of the ovine estrous cycle or early pregnancy was examined. At estrus, ewes were either mated to a fertile ram or unmated. On day 12 postestrus, ewes were laparotomized and a catheter was inserted into a uteroovarian vein. Six mated and 7 unmated ewes received no further treatment. Fifteen mated and 13 unmated ewes were ovariectomized on day 12 and of these, 7 mated and 5 unmated ewes were given 10 mg progesterone sc and an intravaginal pessary containing 30 mg of progesterone. Uteroovarian venous samples were collected every 15 min for 3 h on days 13 to 16 postestrus. Mating resulted in higher mean daily concentrations of PGE2 in the uteroovarian vein than in unmated ewes. Ovariectomy prevented the rise in PGE2 with day in mated ewes but had no effect in unmated ewes. Progesterone treatment restored PGE2 in ovariectomized, mated ewes with intact embryos. Mating had no effect on mean daily concentrations of PGE2 alpha or the patterns of the natural logarithm (1n) of the variance of PGF2 alpha. Ovariectomy resulted in higher mean concentrations and 1n variances of PGF2 alpha on day 13 and lower mean concentrations and 1n variances of PGF2 alpha on days 15 and 16. Replacement with progesterone prevented these changes in patterns of mean concentrations and 1n variances of PGF2 alpha following ovariectomy. It is concluded that progesterone regulates the release of PGF2 alpha from the uterus, maintaining high concentrations while also preventing the occurrence of the final peaks of PGF2 alpha which are seen with falling concentrations of progesterone. This occurs in both pregnant and non-pregnant ewes. Progesterone is also needed to maintain increasing concentrations of PGE2 in mated ewes.     

Activity of phospholipase C in ovine luteal tissue in response to PGF2 alpha, PGE2 and luteinizing hormone.

Four ewes were utilized to determine the effects of prostaglandin (PG) F2 alpha, PGE2 and luteinizing hormone (LH) on activity of phospholipase C (PLC) in ovine luteal tissue. Corpora lutea were collected on d 10 post-estrus and six slices from one corpus luteum from each ewe were pre-incubated with [3H]-inositol prior to incubation with one of 6 treatments. Treatments were 1) control, 2) PGF2 alpha (100 ng/ml), 3) PGE2 (10 ng/ml), 4) LH (10 ng/ml), 5) PGF2 alpha + PGE2 and 6) PGF2 alpha + LH. Phospholipase C was determined indirectly by measuring the accumulation of [3H]-inositol mono-, bis- and tris-phosphates (IP, IP2, IP3). Effects of PGF2 alpha (0 vs. PGF2 alpha) and luteotropic treatment (0 vs. PGE2 vs. LH) and their interactions were determined by analysis of variance. There was a significant main effect of PGF2 alpha (P less than 0.01) as concentrations of IP, IP2, IP3 and total [3H]-inositol phosphates were greater in tissue slices treated with PGF2 alpha, regardless of luteotropic treatment. Within groups receiving no PGF2 alpha (1,3,4), no effect of luteotropic treatment was observed. Within groups receiving PGF2 alpha (2,5,6), LH caused a significant (P less than .05) increase in the accumulation of total [3H]-inositol phosphates. Thus, PGF2 alpha can stimulate the activity of PLC in ovine luteal tissue and LH can potentiate this effect.     

Sensitivity of bovine corpora lutea to prostaglandin F2alpha is dependent on progesterone, oxytocin, and prostaglandins.

Prostaglandin (PG) F2alpha that is released from the uterus is essential for spontaneous luteolysis in cattle. Although PGF2alpha and its analogues are extensively used to synchronize the estrous cycle by inducing luteolysis, corpora lutea (CL) at the early stage of the estrous cycle are resistant to the luteolytic effect of PGF2alpha. We examined the sensitivity of bovine CL to PGF2alpha treatment in vitro and determined whether the changes in the response of CL to PGF2alpha are dependent on progesterone (P4), oxytocin (OT), and PGs produced locally. Bovine luteal cells from early (Days 4-5 of the estrous cycle) and mid-cycle CL (Days 8-12 of the estrous cycle) were preexposed for 12 h to a P4 antagonist (onapristone: OP; 10(-4) M), an OT antagonist (atosiban: AT; 10(-6) M), or indomethacin (INDO; 10(-4) M) before stimulation with PGF2alpha. Although OP reduced P4 secretion (p < 0.001) only in early CL, it reduced OT secretion in the cells of both phases examined (p < 0.001). OP also reduced PGF2alpha and PGE2 secretion (p < 0.01) from early CL. However, it stimulated PGF2alpha secretion in mid-cycle luteal cells (p < 0.001). AT reduced P4 secretion in early and mid-cycle CL (p < 0.05). Moreover, PGF2alpha secretion was inhibited (p < 0.05) by AT in early CL. The OT secretion and the intracellular level of free Ca2+ ([Ca2+]i) were measured as indicators of CL sensitivity to PGF2alpha. PGF2alpha had no influence on OT secretion, although [Ca2+]i increased (p < 0.05) in the early CL. However, the effect of PGF2alpha was augmented (p < 0.01) in cells after pretreatment with OP, AT, and INDO in comparison with the controls. In mid-cycle luteal cells, PGF2alpha induced 2-fold increases in OT secretion and [Ca2+]i. However, in contrast to results in early CL, these increases were magnified only by preexposure of the cells to AT (p < 0.05). These results indicate that luteal P4, OT, and PGs are components of an autocrine/paracrine positive feedback cascade in bovine early to mid-cycle CL and may be responsible for the resistance of the early bovine CL to the exogenous PGF2alpha action.