Effect of RU 486 on luteal function in the early pregnant rat

A dose of 30 mg RU 486/kg, an antiprogesterone, was administered to pregnant rats on Day 2 (Group 1) or Day 4 (Group 2) of pregnancy. RU 486 significantly changed serum progesterone and oestradiol concentrations and luteal 3 beta-HSD and 20 alpha-HSD activities in Group 1, and implantation was significantly inhibited. The luteal 3 beta-HSD activity in Group 2 rats on Day 6 was significantly (P less than 0.01) lower than the control value (7.5 +/- 0.6 and 10.1 +/- 0.6 mU/mg protein respectively). This decline in the 3 beta-HSD activity was followed by a marked decrease in the serum progesterone concentration, resulting in a significant decrease of the progesterone/oestradiol ratio and implantation was completely inhibited. The 20 alpha-HSD activity, which could not be detected on Day 6 in the control rats, was twice as great in Group 2 than in Group 1 rats (17.5 +/- 1.2 and 7.4 +/- 3.1 mU/mg protein respectively). Ultrastructural examination of corpora lutea of Group 2 rats confirmed luteolysis. These results suggest that RU 486 has a luteolytic effect and its anti-implantation effect is concomitant with luteolysis of the corpora lutea of pregnancy.     

In vitro secretion of progestins by rat luteal cells and their 20 alpha-hydroxysteroid dehydrogenase activity

Functionally active or regressing corpora lutea were harvested from pseudopregnant (psp) rats between days 5-8 of psp or day 15 of psp, respectively. They were enzymatically dispersed and cultured for 24 h to assess progestins in the medium and 20 alpha-hydroxysteroid dehydrogenase [20 alpha-HSD, catalyzing the conversion of progesterone to 20 alpha-dihydroprogesterone (20 alpha-OH-P)] activity in the cell. Though the active luteal cells retained low 20 alpha-HSD activity, they secreted 6-7 times more 20 alpha-OH-P than progesterone as the regressing luteal cells did. There was no significant difference between the total amounts of progestins in the 2 groups. When increasing doses of pregnenolone were added to the media, progesterone secretion from the active luteal cells was promoted and the progesterone to 20 alpha-OH-P ratio became comparable to the circulating progestins ratio during the mid-luteal phase. In contrast, from the regressing luteal cells only 20 alpha-OH-P secretion was promoted. These results indicate that an insufficient precursor supply results in the catabolism of a large part of synthesized progesterone before its release from luteal cells and suggest the presence of a high affinity but low capacity 20 alpha-HSD in active corpora lutea.     

Luteolytic action of RU486: Modulation of luteal 3β-hydroxysteroid dehydrogenase and 20α-hydroxysteroid dehydrogenase activities in late pregnant rats

The effect of the synthetic antiprogestin RU486 on luteal function in late pregnant rats was studied by evaluating the activities of the enzymes 3β-hydroxysteroid dehydrogenase (3β-HSD) and 20α-hydroxysteroid dehydrogenase (20α-HSD). RU486 (2 mg/kg) administered to rats on day 18 of pregnancy at 10.00 h induced preterm delivery 26.4 ± 0.35 h (n = 8) after treatment. Luteal 3β-HSD activity increased 24 and 34 h after RU486 injection, but a significant and progressive decrease started at 48 h with the maximal reduction 72 h after RU486 treatment, when compared with controls. Serum progesterone concentration decreased at the time of 3β-HSD activity reduction. Interestingly, 20α-HSD activity started to increase 58 h after RU486 injection. The administration of the cyclooxygenase inhibitor, diclofenac (1.3 mg/kg), on days 17–19 of pregnancy to RU486-treated rats, delayed abortion and the duration of delivery, and prevented the decrease in 3β-HSD and the increase in 20α-HSD activities observed 58 h after antiprogesterone treatment. RU486 administered intrabursally (1 μg per ovary) on day 20 (14.00–15.00 h) increased 3β-HSD and decreased 20α-HSD luteal activities at 18.00 h on day 21 of pregnancy, without modifying serum progesterone concentration, when compared with normal pregnant rats. In conclusion, the luteolytic process after preterm delivery induced by RU486 administration in late pregnant rats is characterized by a decrease in luteal 3β-HSD activity and circulating progesterone, which may trigger the increase in luteal 20α-HSD activity. Prostaglandins seems to be involved in the increase of 20α-HSD activity and therefore, in the demise of corpora lutea.     

Luteal expression of cytochrome P450 side-chain cleavage, steroidogenic acute regulatory protein, 3beta-hydroxysteroid dehydrogenase, and 20alpha-hydroxysteroid dehydrogenase genes in late pregnant rats: effect of luteinizing hormone and RU486

A decrease in serum progesterone at the end of pregnancy is essential for the induction of parturition in rats. We have previously demonstrated that LH participates in this process through: 1) inhibiting 3beta-hydroxysteroid dehydrogenase (3beta-HSD) activity and 2) stimulating progesterone catabolism by inducing 20alpha-hydroxysteroid dehydrogenase (20alpha-HSD) activity. The objective of this investigation was to determine the effect of LH and progesterone on the luteal expression of the steroidogenic acute regulatory protein (StAR), cytochrome P450 side-chain cleavage (P450(scc)), 3beta-HSD, and 20alpha-HSD genes. Gene expression was analyzed by Northern blot analysis 24 and 48 h after administration of LH or vehicle on Day 19 of pregnancy. StAR and 3beta-HSD mRNA levels were lower in LH-treated rats than in rats administered with vehicle at both time points studied. P450(scc) mRNA levels were unaffected by LH. The 20alpha-HSD mRNA levels were not different between LH and control rats 24 h after treatment; however, greater expression of 20alpha-HSD, with respect to controls, was observed in LH-treated rats 48 h after treatment. Luteal progesterone content dropped in LH-treated rats at both time points studied, whereas serum progesterone decreased after 48 h only. In a second set of experiments, the anti-progesterone RU486 was injected intrabursally on Day 20 of pregnancy. RU486 had no effect on 3beta-HSD or P450(scc) expression but increased 20alpha-HSD mRNA levels after 8 h treatment. In conclusion, the luteolytic effect of LH is mediated by a drop in StAR and 3beta-HSD expression without effect on P450(scc) expression. We also provide the first in vivo evidence indicating that a decrease in luteal progesterone content may be an essential step toward the induction of 20alpha-HSD expression at the end of pregnancy in rats.     

Contrasting effects of oestradiol-17 beta and human chorionic gonadotrophin on steroidogenesis in the rabbit corpus luteum

On Day 10 of pseudopregnancy, rabbits were given an i.v. injection of hCG (10-20 i.u.) that was sufficient to cause new ovulations and the loss of follicular oestradiol secretion. There was an immediate 3-4-fold rise in serum progesterone which returned to near prestimulation values (approximately 27 ng/ml) within 12 h in the presence of an implant containing oestradiol-17 beta. In the absence of oestradiol, serum progesterone continued to decline to reach low values (approximately 4 ng/ml) within 24 h and the original corpora lutea subsequently regressed. The administration of oestradiol 24 h after injection of hCG, when progesterone secretion was low, arrested any further decline in progesterone and then restored serum progesterone to normal values. This steroidogenic effect of oestradiol in vivo was a function of enhanced luteal steroidogenesis; corpora lutea removed and incubated for 12 h produced progesterone at high, linear rates, whereas the corpora lutea from animals that did not receive oestradiol produced low or insignificant quantities of progesterone in vitro. We conclude that hCG at these doses is compatible with continued responsiveness of the corpora lutea to oestrogen and that hCG produces its luteolytic effect primarily by ovulating follicles, thus stopping the secretion of the luteotrophic hormone, oestradiol.     

Effect of prostaglandin F2 alpha on release of progesterone and leukotriene B-4 by cells from corpora lutea of mares.

Corpora lutea were recovered from mares either 4 to 5 days or 12 to 13 days after ovulation. Mixed populations of luteal cells were prepared by collagenase digestion and were incubated for 24 h in the presence or absence of prostaglandin (PG) F-2 alpha (250 ng/ml). PGF-2 alpha significantly (P = 0.03) reduced progesterone secretion by cells from late diestrous corpora lutea and tended (P = 0.06) to reduce secretion by early diestrous cells. PGF-2 alpha had no significant effect on leukotriene B-4 (LTB-4) production by cells from early diestrous corpora lutea, but significantly (P = 0.03) increased LTB-4 production by late diestrous luteal cells. It seems possible that LTB-4 could play a role as an intermediary in the action of PGF-2 alpha in luteolysis in the mare.     

Inhibitory effects of the antiprogestin, RU 486, on progesterone actions and luteinizing hormone secretion in pituitary gonadotrophs

The effects of RU 486 on the modulation of LH release by progesterone were investigated in cultured anterior pituitary cells from ovariectomized adult female rats. The inhibitory effect of progesterone on LH secretion was demonstrable in estrogen-treated pituitary cells, in which addition of 10(-6) M progesterone to cells cultured in the presence of 10(-9) M estradiol for 52 h reduced the LH response to GnRH (10(-11) to 10(-7) M). When RU 486 was superimposed upon such combined treatment with estradiol and progesterone, the suppressive effect of progesterone on GnRH-induced LH release was completely abolished. The converse (facilitatory) effect of progesterone on LH secretion was observed in pituitary cells pretreated with 10(-9) M estradiol for 48 h and then with 10(-6) M progesterone for 4 h. When RU 486 was added together with progesterone during the 4 h treatment period, the facilitatory effect of progesterone was blocked and LH release fell to below the corresponding control value. The direct effect of RU 486 on LH secretion in the absence of exogenous progesterone was evaluated in cells cultured in the absence or presence of 10(-9) M estradiol and then treated for 4 to 24 h with increasing concentrations of RU 486 (10(-12) to 10(-5) M) and stimulated with GnRH (10(-9) M) during the last 3 h of incubation. In estrogen-deficient cultures, 4 h exposure to RU 486 concentrations of 10(-6) M and above decreased the LH response to GnRH by up to 50%. In cultures pretreated with 10(-9) M estradiol, GnRH-stimulated LH responses was inhibited by much lower RU 486 concentrations, of 10(-9) M and above. After 24 h of incubation the effects of RU 486 were similar in control and estradiol-pretreated pituitary cell cultures. Thus, RU 486 alone has a significant inhibitory effect on LH secretion that is enhanced in the presence of estrogen. The antiprogestin is also a potent antagonist of both the inhibitory and the facilitatory actions of progesterone upon pituitary gonadotropin release in vitro.     

Progesterone receptor is not required for progesterone action in the rat corpus luteum of pregnancy

In this study, we investigated whether progesterone exerts a local action regulating the function of the corpus luteum of pregnancy in rats. The luteal activities of the enzymes 3beta-hydroxysteroid dehydrogenase (3beta-HSD), involved in progesterone biosynthesis, and 20alpha-hydroxysteroid dehydrogenase (20alpha-HSD), that catabolizes progesterone and reduces progesterone secretion by the corpus luteum, were evaluated after intrabursal ovarian administration of progesterone in pregnant rats that had received a luteolytic dose of prostaglandin F2alpha (PGF2alpha). Luteal 3beta-HSD activity decreased and 20alpha-HSD activity increased after PGF2alpha treatment (100 microg x 2 intraperitoneally on Day 19 of pregnancy at 12:00 p.m. and 4:00 p.m.) when compared with controls sacrificed at 8:00 p.m. on Day 20 of pregnancy. This effect of PGF2alpha on the luteal 3beta-HSD and 20alpha-HSD activities was abolished in animals that also received an intraovarian dose of progesterone (3 microg/ovary on Day 19 of pregnancy at 8:00-9:00 a.m.). In a second functional study, luteal cells obtained from 19-day pregnant rats responded to the synthetic progestin promegestone (R5020) in a dose-dependent manner, with an increase in the progesterone output. In addition, the glucocorticoid agent hydrocortisone did not affect progesterone accumulation in the same luteal cell culture. We also examined by immunocytochemistry the expression of progesterone receptors (PR) in the corpora lutea during pregnancy and demonstrated the absence of PR in this endocrine gland in all the days of pregnancy studied. In the same pregnant rats, positive staining for PR was observed in cells within the uteroplacental unit, such as cells of the decidua basalis and trophoblast giant cells of the junctional zone. In addition, positive PR staining was observed in the ovarian granulosa and theca cells of growing follicles, but not in corpora lutea of ovaries obtained from cycling rats at proestrus. In summary, this report provides further evidence of a local action of progesterone regulating luteal function in the rat despite the absence of a classic PR.     

Progesterone receptor-mediated inhibition of apoptosis in granulosa cells isolated from rats treated with human chorionic gonadotropin

Almost all ovarian follicles undergo atresia during follicular development. However, the number of corpora lutea roughly equals the number of preovulatory follicles in the ovary. Because apoptosis is the cellular mechanism behind follicle and luteal cell demise, this suggests a change in apoptosis susceptibility during the periovulatory period. Sex steroids are important regulators of follicular cell survival and apoptosis. The aim of the present work was to study the role of progesterone receptor-mediated effects in the regulation of granulosa cell apoptosis. The levels of internucleosomal DNA fragmentation were evaluated in rat granulosa cells before and after induction of the nuclear progesterone receptor, using hCG treatment to eCG-primed rats to mimic the naturally occurring LH surge. Granulosa cells isolated from hCG-treated rats showed a several-fold increase in the expression of progesterone receptor mRNA and a 47% decrease (P < 0.01) in DNA fragmentation after 24 h incubation in serum-free medium compared to granulosa cells isolated from rats treated with eCG only. The effect of hCG treatment in vivo was dose-dependently reversed in vitro by addition of antiprogestins (Org 31710 or RU 486) to the culture medium, demonstrated by increased DNA fragmentation as well as increased caspase-3 activity. Addition of antiprogestins to granulosa cells isolated from immature or eCG-treated rats did not result in increased DNA fragmentation. The results suggest that progesterone receptor-mediated effects are involved in regulating the susceptibility to apoptosis in LH receptor-stimulated preovulatory rat granulosa cells.     

Inhibition of 20 alpha-hydroxysteroid dehydrogenase activity by follicle-stimulating hormone and androgens in cultured rat granulosa cells: a search for the mechanism of action

Alterations of progesterone metabolism and especially of 20 alpha-hydroxysteroid dehydrogenase (20 alpha-HSD) activity were studied in cultured rat granulosa cells following various treatments. The cells were incubated for up to 48 h with or without follicle-stimulating hormone (FSH), androgens, hydroxyflutamide, estrogens, chlorea toxin, and dibutyryl cAMP [Bu2 cAMP]. Subsequently, the cells were incubated for 3 h with [4-14 C] progesterone (0.5 microM). The progesterone utilization and accumulation of 20 alpha-reduced and 5 alpha-reduced metabolites were assessed following thin-layer chromatography separation of radiolabeled steroids. Both FSH (1 microgram/ml) and testosterone (0.5 microM) decreased the 20 alpha-HSD activity by decreasing the maximal velocity (by 52% and 37%, respectively) without changing significantly the Km value. The inhibition of 20 alpha-HSD was demonstrable following 12 and 24 h exposure to FSH and following 24 and 48 h exposure to testosterone. Effects comparable to that induced by testosterone were elicited by other androgens (androstenedione and 5 alpha-dihydrotestosterone), but not by estrogens (estradiol-17 beta and estrone). Hydroxyflutamide reversed testosterone-induced effects: the increase of endogenous progesterone accumulation and the decrease of 20 alpha-HSD activity. Both cholera toxin (0.001-10 micrograms/ml) and Bu2 cAMP (62.5-1000 micrograms/ml) caused a dose-dependent inhibition of 20 alpha-HSD activity. Present results indicate that: the inhibition of 20 alpha-HSD by both FSH and androgens may be of a noncompetitive nature; androgen action on 20 alpha-HSD may be a true androgenic, receptor-mediated effect; and cAMP may mediate the FSH action on 20 alpha-HSD activity.     

Invitro responsiveness of hamster corpora lutea undergoing luteolysis to luteinizing hormone

Corpora lutea removed from pregnant hamster deprived of endogenous luteinizing hormone for varying periods were compared for their responsiveness to externally added luteinizing hormone. The corpora lutea removed on the 8th day of pregnancy exhibited a dose-dependent increase in progesterone production in response to added luteinizing hormone upto a concentration of 2.5 Μg/ml. The total progesterone synthesised by the corpora lutea decreased with increase in the duration ofin vivo luteinizing hormone deprivation. However, the hormone deprivation had to be for a minimum period of 24 h before a marked reduction in thein vitro responsiveness could be seen. Neutralisation of endogenous luteinizing hormone increased the luteal cholesterol ester concentration, whilein vitro incubation of such tissue with luteinizing hormone resulted in a marked reduction in cholesterol ester levels. Corpora lutea removed from hamsters on day 8, 15 and 16 of pregnancy when compared for their responsivenessin vitro to added luteinizing hormone showed that the luteal tissue of day 8 produced more progesterone relative to those of day 15/16. In contrast, depletion of free and esterified cholesterol increased with the increase in age of corpora lutea (from 15% on day 8 to 67% on day 16).     

Variations in oxytocin, vasopressin and neurophysin concentrations in the bovine ovary during the oestrous cycle and pregnancy

Bovine ovaries were obtained from the abattoir and corpora lutea were classified as: (1) early luteal phase (approximately Days 1-4); (2) mid-luteal phase (Days 5-10); (3) late luteal phase (Days 11-17); (4) regressing (Days 18-20) and (5) pregnant (Days 90-230). In addition, preovulatory follicles and whole ovaries without luteal tissue were collected. Concentrations of oxytocin, vasopressin, bovine neurophysin I and progesterone were measured in each corpus luteum by radioimmunoassay. Progesterone and neurophysin I levels increased from Stage 1 to Stage 2, plateaued during Stage 3 and declined by Stage 4. Oxytocin and vasopressin concentrations increased from Stage 1 to Stage 2 but declined during Stage 3 and were low (oxytocin) or undetectable (vasopressin) in follicles, whole ovaries and pregnancy corpora lutea. Therefore the concentrations of both peptide hormones were maximal during the first half of the cycle and declined before those of progesterone. The high concentration of oxytocin within the corpus luteum coupled with the presence of bovine neurophysin I suggests that oxytocin is synthesized locally.     

Modulatory actions of the new antiprogestins ZK 98.299 and ZK 98.734 and of RU 486 on luteinizing hormone secretion and progesterone effects in pituitary gonadotrophs

The effects of the antiprogestins (APs) ZK 98.299, ZK 98.734 and RU 486 on GnRH-stimulated LH secretion and their antagonistic activity on progesterone (P) actions were investigated in cultured pituitary cells from adult female Wistar rats. P (100 nM) was able to exert a facilitatory effect on GnRH (1 nM)-induced LH secretion after short-term (4 h) treatment of estradiol-primed (1 nM, 48 h) rat pituitary cells. When the APs (10 pM-10 microM) were introduced during the 4 h incubation period with P the facilitatory effect of P was totally abolished at concentrations greater than 10 nM (ZK 98.299, ZK 98.734) and greater than 1 nM (RU 486). Also the APs were shown to block the inhibitory action of P which occurs after long-term incubation of pituitary cells with this steroid. However at concentrations greater than 10 nM (ZK 98.734, RU 486) and greater than 100 nM (ZK 98.299) this antagonistic action of the APs was lost. To evaluate whether the APs have direct effects on GnRH-induced LH secretion in the absence of exogenous P pituitary cells cultivated for 48 h with or without 1 nM estradiol were incubated for 4 or 24 h with increasing concentrations of the APs (10 pM-10 microM). Four hour treatment of non-estradiol-primed cells with ZK 98.299 or ZK 98.734 was without any effect on the LH response to a 1 nM GnRH-stimulus. Only the highest concentration of RU 486 (10 microM) reduced the LH response. Twenty-four hour treatment of the cultures with the APs led to enhancement of GnRH-stimulated LH secretion by up to 113, 37 and 33% for ZK 98.734, ZK 98.299 and RU 486, respectively. When estradiol-primed cells were used for the same experiments we observed exclusively inhibitory effects on GnRH-induced LH secretion after 4 and 24 h treatment periods. It is concluded that these new APs are potent inhibitors of P-actions, but also per se they induce diverse effects on GnRH-stimulated LH secretion in cultured rat pituitary cells which have to be taken into account.     

Role of arachidonic acid and its metabolites in the regulation of progesterone and oxytocin release from the bovine corpus luteum.

We have examined the effects of arachidonic acid (AA) and some of its metabolites on progesterone (P4) and oxytocin (OT) release by corpora lutea obtained from Holstein heifers at day 8 of the estrous cycle (Day 0 = estrus). The luteal cells were dispersed with collagenase and small and large cells were separated by unit gravity sedimentation and flow cytometry. After an 18-hr preincubation period, the cells were incubated in the presence of various treatments for 1 hr, followed by a 23-hr incubation period with no treatment. OT was secreted by the large, but not by the small, luteal cells into the incubation medium. AA elicited a significant (P less than 0.05) release of OT from the large cells and P4 from both the large and small cells within 1 hr of incubation, having a specific effect at a concentration of 10 microM. Larger doses (25 and 100 microM) of AA adversely affected the cell viability. Phospholipases A2 (0.5 unit/ml) and C (0.05 unit/ml) and calcium ionophore A23187 (0.1 microM) stimulated OT release from the large cells to the same extent as AA (10 microM). Inhibition of the AA cyclooxygenase metabolic pathway by indomethacin did not affect AA-induced release of OT and P4, although exogenous prostaglandins F2 alpha and I2 (5-25 ng/ml) stimulated the release of OT. Lipoxygenase products of AA (hydroxyeicosatetraenoic acid and leukotrienes; 25 ng/ml) also stimulated OT release. Inhibition of the lipoxygenase metabolic pathway by nordihydroguaiaretic acid abolished AA-induced release of both OT and P4. These results suggest that intracellular accumulation of free AA may modulate secretory functions in the bovine corpora lutea, including OT and P4 release.     

Decreased luteinizing hormone-stimulated progesterone secretion by preovulatory follicles isolated from cyclic rats treated with the progesterone antagonist RU486.

Since administration of the antiprogesterone RU486 to cyclic female rats at metestrus and diestrus results in increased serum levels of LH, estradiol, and testosterone at proestrus, we investigated whether RU486 affects follicular steroidogenesis. Female rats with a 4-day estrous cycle, induced experimentally by a single injection of bromocriptine on the morning of estrus, were given RU486 (2 mg) twice daily (0900 and 1700 h) on metestrus and diestrus. At proestrus the preovulatory follicles were isolated and incubated for 4 h in the absence and presence of LH. In the absence of LH, accumulation of estradiol, testosterone, and progesterone in the medium was not different for RU486-treated rats and oil-treated controls. In contrast, LH-stimulated estradiol, testosterone, and progesterone secretions were significantly lower in RU486-treated rats compared with controls. Addition of pregnenolone to the incubation medium resulted in a significantly lower increase of progesterone in follicles from RU486-treated rats compared with those from oil-treated controls. This suggests that 3 beta-hydroxysteroid dehydrogenase (3 beta-HSD) activity is decreased by administration of RU486 in vivo. Aromatase and 17 alpha-hydroxylase/C17-20 lyase activities were not affected: addition of substrate (androstenedione and progesterone respectively) did not affect differently the amount of product formed (estradiol and testosterone) in RU486- and oil-treated rats. However, LH-stimulated pregnenolone secretion was lower in follicles from RU486-treated rats compared with follicles from oil-treated controls, suggesting that either cholesterol side-chain cleavage activity or LH responsiveness is decreased. At proestrus the preovulatory follicles from RU486- and oil-treated rats were not morphologically different.(ABSTRACT TRUNCATED AT 250 WORDS)     

Synergistic effects of insulin-like growth factor I and gonadotrophins on relaxin and progesterone secretion by ageing corpora lutea of pigs.

Insulin-like growth factor I (IGF-I) is involved in paracrine/autocrine regulation of gonadal steroidogenesis and peptide hormone biosynthesis. This study was designed to determine whether IGF-I alone, or an interaction of IGF-I, is involved in augmenting the actions of luteinizing hormone (LH) and prolactin in controlling relaxin and progesterone secretion from ageing corpora lutea of hysterectomized gilts at days 110, 113 and 116 after oestrus. Luteal tissue slices were incubated for 8 h with IGF-I (0, 50, 300 ng ml-1), LH (0, 100, 1000 ng ml-1), and prolactin (0, 100, 1000 ng ml-1) alone or in combination. Progesterone and relaxin concentrations were determined by radioimmunoassay of spent medium and of homogenates from luteal tissue slices before and after incubation. Porcine luteal tissue from day 110 had a net output of 25 ng progesterone and 26 ng relaxin in the control and of 65 ng progesterone and 2125 ng relaxin in the combined IGF-I, LH and prolactin treatment mg-1 of luteal tissue, respectively. IGF-I, LH and prolactin alone or in combination significantly increased (P < 0.01) progesterone production by luteal tissue from day 110, but they were partially effective at day 113 and ineffective at day 116. By contrast, the same hormone treatments increased relaxin production by luteal tissue from days 110 and 113. Even at day 116, prolactin alone or with LH or IGF-I continued to stimulate relaxin production. In conclusion, IGF-I augments the ability of prolactin and LH to increase relaxin production by ageing corpora lutea; however, a decrease in progesterone secretion and an increase in relaxin secretion at day 113 indicate that different mechanisms control progesterone and relaxin secretion in pigs.     

Function of abnormal corpora lutea in vitro after GnRH-induced ovulation in the anoestrous ewe

Normal and abnormal corpora lutea were recovered from anoestrous Romney Marsh ewes on Days 3, 4, 5 and 6 after treatment with small-dose (250 ng) multiple injections of GnRH followed by a bolus injection (125 micrograms) with (+P) and without (-P) progesterone pretreatment and a study made of their characteristics in vitro. Plasma progesterone concentrations initially rose concurrently in all animals but abnormal luteal function occurred in 70% of the -P ewes and was defined on Day 5 when plasma progesterone concentrations declined relative to those in the +P ewes. All corpora lutea recovered on Days 3 and 4 appeared macroscopically similar and there were no significant differences between the +P and -P groups in terms of luteal weight, progesterone content and binding of 125I-labelled hCG on these days. However, corpora lutea from the -P animals only exhibited a decline in progesterone production in vitro on Day 4 (P less than 0.01), and morphological differences became apparent on Days 5 and 6 when the abnormal corpora lutea from the -P animals also decreased in weight (P less than 0.01) and progesterone content (P less than 0.001). Binding of 125I-labelled hCG increased on Day 5 in the normal corpora lutea only. These results show that, although abnormal luteal function induced by GnRH treatment of anoestrous ewes could not be distinguished from normal corpora lutea before Day 5 by measurement of progesterone in peripheral plasma, a significant decline in progesterone production in vitro occurred on Day 4 in the abnormal corpora lutea. This was followed by significant decreases in weight and progesterone content and a failure to increase 125I-labelled hCG binding. Abnormal corpora lutea are therefore capable of some initial growth and progesterone production, before undergoing a rapid and premature regression from Day 4, which has similar characteristics to natural luteolysis.