Use of spironolactone to investigate the role of testosterone secretion during luteolysis in the goat

In non-pregnant goats, appreciable amounts of testosterone (2.1 ng/g) and 5 alpha-dihydrotestosterone (DHT, 0.8 ng/g) were present in the corpus luteum on Day 12 of the oestrous cycle. Significant (P less than 0.01, N = 18) veno-arterial concentration differences of testosterone were found across ovaries bearing corpora lutea. No such difference in testosterone concentration occurred across ovaries without corpora lutea (P greater than 0.5, N = 12). Increased peripheral plasma concentrations of testosterone and DHT occurred at the start of luteal regression, as monitored by progesterone concentration, and before the day of oestrus. Subcutaneous injections of spironolactone (10 mg/kg/day) in peanut oil between Days 10 and 20 of the oestrous cycle inhibited the increase in testosterone and DHT concentrations and delayed luteolysis and oestrus. It is suggested that aromatization of testosterone to oestrogens is needed for luteal regression and expression of oestrus in goats.     

Failure of LH and/or prolactin to prevent PGF2α-induced luteolysis of ovine corpora lutea

Infusions of phosphate buffered saline, LH (4 μg/min or 14 μg/min), prolactin (42 μg/min) or LH (4 μg/min) plus prolactin (42 μg/min) for 12 hr did not prevent luteolysis following intramuscular injections of prostaglandin F₂α-tham salt two and six hr after beginning the infusion. Likewise, these treatments did not delay luteolysis since a similar rate of decline in peripheral plasma progesterone occurred in all groups. It was concluded that elevation of serum concentrations of LH and prolactin to high levels had no effect on PGF₂α-induced luteolysis on day 8 following induced ovulation.     

Efficacy of a prostaglandin analogue in reproduction in the anestrous mare

A prostaglandin F analogue was studied in anestrous mares: a dose-response study; a study in mares presumed pregnant; and a field evaluation of effective doses in breeding establishments. A dose of 2.0mg given by single subcutaneous injection to mares with initial plasma progesterone levels greater than 1.0ng/ml, caused luteolysis on the basis of decline in plasma progesterone concentrations. Follicle maturation leading to ovulation, accompanied by estrus, was observed, and fertility at mating either by natural service or artificial insemination was satisfactory. A dose of 1.0mg was generally effective for luteolysis, but pregnancy rates were lower than after 2.0mg. A proportion of mares which had less than 1.0mg of plasma progesterone at the time of injection ovulated and became pregnant.     

Pulsatile output of prostaglandin F(2alpha) does not increase around the time of luteolysis in the pregnant goat.

Prostaglandin (PG) F(2alpha) secreted from the uterus is the luteolysin of the estrous cycle and is also believed to be responsible for luteolysis in the pregnant doe at term. We have reported that basal progesterone concentrations decrease before basal PGF(2alpha) concentrations increase, which is inconsistent with this view. In this study we investigated whether luteolysis is associated with increased frequency or amplitude of pulsatile PGF(2alpha) secretion in does over the last 2 wk of gestation. Progesterone concentrations decreased approximately 1 wk before parturition. There was no accompanying increase in PGF(2alpha) concentrations or pulse frequency, and those pulses that were observed were of lesser amplitude and duration than those that have been associated with luteolysis in cycling ewes. A small increase in PGF(2alpha) pulse frequency was identified during the 3 days before parturition, but this was not associated with any change in progesterone concentrations. The biological significance of these small changes in PGF(2alpha) pulse frequency is obscure, although the high concentration of this eicosanoid at labor may have been related to the final, precipitous decline in plasma progesterone concentrations. These findings do not support the notion that PGF(2alpha) is the principal luteolysin in the pregnant doe at term.     

Hormonal changes during luteal regression in farmed fallow deer, Dama dama

Concentrations of progesterone, oxytocin and PGFM (pulmonary metabolite of PGF-2 alpha) were measured in plasma from peripheral blood samples collected from 5 fallow does every hour or 2 h for 12-h periods on Days 15-20 inclusive of the oestrous cycle (i.e. luteolysis). For 3 does that exhibited oestrus on Day 21, plasma progesterone concentrations fluctuated between 3 and 10 ng/ml on Days 15-18 inclusive. Thereafter, values declined progressively to attain minimum concentrations of less than 0.05 ng/ml on Day 20. Basal concentrations of plasma oxytocin and PGFM fluctuated between 5 and 20 pg/ml and 10 and 100 pg/ml respectively. Episodic pulses of plasma oxytocin (greater than 300 pg/ml) occurred on Days 15 and 16, whereas pulses of plasma PGFM (greater than 400 pg/ml) occurred on Days 19 and 20. There was little apparent correlation between episodic pulses of the two hormones. For 2 does that exhibited oestrus on Day 22, plasma progesterone concentrations declined to minimum values of 1.0-1.5 ng/ml by Day 20. One of these does showed very high levels of oxytocin secretion throughout the sampling period while the other showed an apparent paucity of oxytocin secretory periods. Two does hysterectomized on Day 13 of their second oestrous cycle failed to exhibit further oestrous cycles. Continual elevation of plasma progesterone concentrations (2-6 ng/ml) for an 8-month period indicated persistence of the corpus luteum after hysterectomy. It is concluded that luteolysis in fallow deer involves episodic secretion of both oxytocin and PGF-2 alpha.     

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.     

Comparison of the rate of decline in plasma progesterone concentrations at a natural and progesterone-synchronized oestrus and its effect on tonic LH secretion in the ewe

The pattern of change in plasma progesterone and LH concentrations was monitored in Clun Forest ewes at a natural oestrus and compared to that observed after removal of progesterone implants. The rate of decline in plasma progesterone concentrations after implant withdrawal (1.8 +/- 0.2 ng/ml h-1) was significantly greater (P less than 0.001) than that observed at natural luteolysis (0.2 +/- 0.1 ng/ml h-1), and this resulted in an abnormal pattern of change in tonic LH secretion up to the time of the preovulatory LH surge. This more rapid rate of progesterone removal was also associated with a shortening of the intervals from the time that progesterone concentrations attained basal values to the onset of oestrus (P less than 0.05) and the onset of the preovulatory LH surge (P less than 0.01). However, there were no significant differences in the duration of the LH peak, preovulatory peak LH concentration, ovulation rate or the pattern of progesterone concentrations in the subsequent cycle. It is suggested that the abnormal patterns of change in progesterone and tonic LH concentrations may be one factor involved in the impairment of sperm transport and abnormal patterns of oestradiol secretion known to occur at a synchronized oestrus.     

Concentrations of 13, 14-dihydro-15-keto prostaglandin F2 alpha after pulsatile progesterone administration at the time of luteolysis of heifers

Progesterone was administered in pulses to 12 dairy heifers from days 17.5 to 22.5 post-estrus in order to determine its ability to modify secretion of PGF2 alpha around the time of luteolysis. Control heifers exhibited pulses of PGFM concomitant with a sharp decline in progesterone concentrations and thus these pulses were temporally associated with luteolysis. Additional pulses of PGFM were observed in heifers receiving exogenous progesterone, but these were not statistically predictable by either dose of progesterone (50 or 100 micrograms) or time of administration (3 or 6 hour intervals). However, all heifers (4/4) treated with progesterone at 3 hour intervals had additional pulses of PGFM as compared to only one heifer (1/4) treated at 6 hour intervals. When pulses of PGFM were induced by exogenous progesterone there was a substantial lag time between the initiation of progesterone treatment and their occurrence. The limited response to progesterone administration and the lack of synchrony is not consistent with an ability of exogenous progesterone to directly stimulate secretion of PGF2 alpha at the time of luteolysis.     

Levels of progesterone and changes in prostaglandin F(2alpha) release during luteolysis and early pregnancy in llamas and the effect of treatment with flunixin meglumine

The secretory patterns of progesterone in relation to concentrations of 15-ketodihydro-PGF(2alpha) (PGFM) during the period of luteolysis or of maternal recognition of pregnancy were determined in the blood of llamas mated either with an intact or a vasectomized male. The ability of flunixin meglumine (FM) to postpone luteolysis in non-pregnant llamas was investigated by injecting the drug intravenously every 6 h at a dose of 2.2 mg/kg from days 6 to 12 post-copulation into a group of non-pregnant llamas. A pulsatile pattern of prostaglandin release was recorded during luteolysis in non-pregnant llamas, giving further support to the hypothesis that PGF(2alpha) is the luteolytic agent in llamas. The mean number of peaks per animal rose from 0.3 on day 7 to 3.8 on day 10 and then declined to 1.1 on day 12 with corresponding mean peak amplitude changing from 465 to 1234 and 566 pmol l(-1), respectively. In pregnant llamas, prostaglandin pulsatile release also occurred. The mean number of peaks per animal rose from 0.4 on day 7 to 0.8 on day 10 and then declined to 0.2 on day 11 and 0.6 on day 12, with corresponding mean peak amplitude changing from 494 to 676, 388 and 547 pmol l(-1), respectively. The transient decrease and subsequent recovery in progesterone concentrations was observed to occur in connection with prostaglandin release during early pregnancy. Oestradiol-17beta plasma peak concentrations attained after luteolysis were significantly higher than those recorded in early pregnant animals (around 30 pmol l(-1) and ll pmol l(-1)). Concentrations of PGFM decreased rapidly after the first administration of FM and remained low throughout the first 2 days of treatment. Thereafter, pulsatile release of prostaglandins started, and luteolysis proceeded; but a delay of 1-1.5 days in the progesterone decline was observed. Thus, it might be suggested that a higher dose and/or a more intensive injection schedule is required in llamas than in other ruminants to prevent luteolysis.     

Effects of dihydrotestosterone on progesterone secretion in pseudopregnant rats

To determine if the administration of dihydrotestosterone (DHT) suppresses serum progesterone concentrations in pseudopregnant rats and if so what role the decidua play in mediating this effect, two pellets (8 mg) of DHT were inserted under each ovarian bursa on Day 9 of pseudopregnancy in hysterectomized or intact rats with or without the decidua. The treatment induced within 24 hr a 50% decline in serum progesterone concentrations in decidua-bearing rats only; a further reduction was observed on Days 11 and 12. To further determine if the effect of DHT on serum progesterone concentrations is due to its effect on luteal luteinizing hormone (LH)-receptor content, rats were similarly treated on Day 9 and the capacity of corpora lutea to bind 125I-labeled human chorionic gonadotropin (hCG) on Day 12 in all three groups of rats was examined. DHT treatment had no effect on luteal LH-receptor content in any group. These results suggest that DHT is luteolytic in pseudopregnant rats only in the presence of the decidua and it is speculated that this effect is mediated by suppressing the decidual luteotropin.     

Luteolytic action of two antiprogestational agents (RU-38486 and ZK-98734) in the rat

RU-38486 or ZK-98734 treatment (3 mg/day, s.c.) to intact or hysterectomized adult female rats on Days 5-7 post coitum induced changes characteristic of luteolysis. Ultrastructurally, the luteal cells exhibited an extensive vacuolization of the cytoplasm and perinuclear areas, degeneration of mitochondrial cristae, massive accumulation of lipid droplets, increase in number of lysosome like granules and heterochromatinization of the nucleus. In general, RU-38486 induced more marked degeneration of the luteal cells than did ZK-98734. There was also a significant decrease in peripheral plasma progesterone concentrations in treated rats. We suggest that these antiprogestagens act via inhibition of luteal function in addition to their antagonism at the uterine progesterone receptor level.     

Alterations in the androgen metabolism of the female guinea-pig during pregnancy

The peripheral conversion of testosterone (T) into androstenedione (delta 4) and dihydrotestosterone (DHT) were studied in pregnant (days 32, 48 and 67 post coitum) and non-pregnant anesthetized guinea-pigs by constant intravenous infusion of tritium labeled T. Endogenous levels were measured by radioimmunoassay, and the three labeled androgens (T*, DHT* and delta 4*) counted after celite chromatography. The endogenous T, DHT and delta 4 plasma concentrations were respectively 5, 7 and 20 times higher in pregnant females than they were in cyclic ones. In the non-pregnant guinea-pig, no part of the delta 4 or DHT pools was derived from the peripheral conversion of testosterone. On days 32, 48 and 67 of gestation the fractions of the delta 4 pool originating from T were 9.1, 13.1 and 15.1%, and the fractions of DHT pool coming from T were 5.1, 22.1 and 21.3% respectively.     

Plasma oestradiol and progesterone during early pregnancy in the cow and the effects of treatment with buserelin

We have monitored plasma concentrations of oestradiol and progesterone after insemination in dairy cows, and investigated the effects of injection with 10 μg of the gonadotrophin-releasing hormone (GnRH) analogue buserelin, a treatment known to result in an improvement in conception rate. Animals were injected intramuscularly on Day 12 after insemination with 2.5 ml of either control saline (n = 29) or buserelin (n = 26). Blood samples were collected from Day 8 to Day 17 and milk samples from the day of insemination until Day 30. On the basis of milk progesterone profiles, in the control group 15 cows remained pregnant (control pregnant) and 11 underwent luteolysis (control not pregnant), whereas in the treated group 13 cows remained pregnant (treated pregnant) and 13 underwent luteolysis (treated not pregnant). Three cows were excluded from the control group owing to high progesterone at insemination or failed ovulation. In both the control and treated groups mean plasma progesterone concentration was significantly (P < 0.05) lower in non-pregnant cows than in pregnant ones from Day 12 post-insemination. However, no significant effect of buserelin treatment on plasma progesterone concentration was detected. In the control group the plasma oestradiol concentration was similar in the pregnant and non-pregnant cows. In the treated group, plasma oestradiol concentration in the non-pregnant cows was similar to that in the control group, whereas in the treated pregnant cows the plasma oestradiol concentration showed a significant (P < 0.05) decline after treatment. As oestradiol is known to stimulate the development of the luteolytic mechanism at this time, we suggest that buserelin is acting to reduce the strength of the luteolytic drive in some cows, thus improving the chance of the embryo being able to prevent luteolysis.     

Uterine oxytocin receptors in cyclic and pregnant cows.

Binding of [3H]oxytocin to uterine subcellular preparations ('oxytocin receptor concentrations') was measured in uterine tissue of heifers and multiparous dairy cows at various stages of the oestrous cycle and during early pregnancy. A method for the assay of ovine uterine oxytocin receptors was optimized for use on bovine tissue. Oxytocin receptor concentrations were increased in cyclic animals around the period of luteolysis and oestrus, rising on Day 15 in endometrium and on Day 17 in myometrium while pregnant animals showed no comparable rise. Receptor concentrations then declined on Day 3 after oestrus in myometrium and on Day 5 in endometrium. Some cyclic animals did not show the expected rise in receptors in the late luteal phase; these animals had abnormally high progesterone concentrations for this stage of the cycle. In animals slaughtered on Day 18 after oestrus and/or insemination which had low oxytocin receptor levels, plasma progesterone concentrations were consistently high; while all animals showing the late luteal phase elevation in receptor values had low progesterone concentrations. Oxytocin receptor and progesterone concentrations were negatively correlated (P less than 0.05). These data support the hypothesis that oxytocin receptor level is a key factor in the process of luteolysis in cattle and that in pregnancy there is suppression of uterine oxytocin receptor at the expected time of luteolysis. We suggest that uterine oxytocin receptor levels are partly controlled by circulating steroid hormones and are suppressed during early pregnancy.     

Plasma steroids in benign prostatic hypertrophy and carcinoma of the prostate

Cortisol, 17-hydroxyprogesterone (17-OH-P), progesterone, testosterone, 5 alpha-dihydrotestosterone (DHT), estrone (E1) and estradiol (E2) were determined by RIA after chromatographic separation of steroids on Sephadex LH-20 columns, in 54 hospitalized patients with benign prostatic hyperplasia (BPH) and in 32 hospitalized patients with prostatic carcinoma (PCA) (T34, N01, M01). The patients' values were compared with those of 63 age-matched controls. Increased cortisol and DHT levels, subnormal estrogen and 17-OH-P values and normal progesterone level were found in both benign and malignant groups. Higher mean values for testosterone, and T/DHT ratio and lower mean values for E2/T ratio were found in PCA, as compared with those in BPH. An age invariance of cortisol, testosterone, T/DHT ratio and estradiol was found in both BPH and PCA, instead of the age dependence found in normal subjects. The normal relations between testosterone and its precursor (17-OH-P) and its peripheral metabolite (E2), respectively, and the normal relation between estrone and 17-OH-P were not evident in either BPH or PCA group. The normal direct relation between testosterone and DHT has been found in both patient groups.     

Peripartal changes in plasma progesterone and 15-keto-13,14-dihydro-prostaglandin F2α concentrations in Holstein cows with or without retained foetal membranes

Peripheral blood plasma concentrations of progesterone and the main metabolite of prostaglandin F_2α, (15-keto-13,14-dihydro-PGF_2α) PGFM, were determined in 10 Holstein cows with retained foetal membranes (RFM) and 12 Holstein cows without RFM (NRFM) during the peripartal period. The rate of uterine involution in the postpartum cows was monitored.There was no difference in the rate of uterine involution between cows with or without RFM. Cyclical ovarian activity was resumed within a month after parturition in both group. Increases in the mean peripheral plasma PGFM concentrations were evident in the RFM cows 6 days before parturition, compared to 48 h before parturition in the NRFM cows. A gradual decline in PGFM to prepartum concentrations occurred in both groups by Day 12 after parturition, although in the RFM cows, PGFM concentrations remained high until the placenta was shed.In both groups, the mean peripheral plasma concentrations of progesterone showed a marked decline beginning 48 h before partusition. The mean plasma progesterone concentrations were less than 1 ng/ml during the immediate postpartum period.     

Effects of prostaglandin and prolactin on luteolysis and parturient behaviour in the non-pregnant tammar, Macropus eugenii

In Exp. 1 non-pregnant female tammars were injected, on Day 26 (the day parturition would normally occur) after removal of pouch young, with saline, 200 micrograms ovine prolactin or 5 mg PG and changes in plasma concentrations of progesterone, prolactin, PGF-2 alpha metabolite (PGFM), oestradiol-17 beta and LH were determined. Luteolysis occurred in females treated with prolactin alone, while treatment with PG first induced a rapid rise in prolactin and subsequently a significant decrease in plasma progesterone. After prolactin treatment the oestradiol peak, oestrus and the LH surge were advanced significantly compared to the saline-treated females. In Exp. 2 the effects of the same treatments as used in Exp. 1 were determined on Day 23 and again on Day 26 after removal of pouch young in non-pregnant females. On Day 23 both prolactin and PG induced significant elevations in plasma progesterone, but luteolysis did not occur. On Day 26 the treatments initially induced significant elevations in plasma progesterone but these were followed by luteolysis within 8-12 h after treatment. PG treatment induced parturient behaviour in the non-pregnant females within 3-21 min and this persisted during the period that plasma concentrations of PGFM were elevated. The results show that PG induces birth behaviour and the release of prolactin, while prolactin first induces an elevation of plasma progesterone concentrations and, in the mature CL on Day 26, subsequently induces luteolysis.     

An intra-corpus luteum site for the luteolytic action of prostaglandin F2 alpha in the rhesus monkey

It has not been possible to demonstrate prostaglandin F2 alpha (PGF2 alpha) participation in primate luteolysis under conditions of systemic administration or of acute intraluteal injection. These study designs were hampered by the short biological half-life in the first instance and brevity of administration in the latter. In this study, luteolysis has resulted from chronic, intraluteal delivery of PGF2 alpha. Using the Alzet osmotic pump-cannula system, normally cycling rhesus monkeys were continuously infused, until menses occurred, with PGF2 alpha (10 ng/1/hr) directly into the corpus luteum (CL, n = 6), into the stroma of the ovary not bearing the corpus luteum (NCL, n = 3), or subcutaneously (SC, n = 5). An additional 5 monkeys received vehicle (V) into the corpus luteum. All experiments commenced 5-7 days after the preovulatory estradiol surge. Luteal function was assessed by the daily measurements of plasma progesterone, estradiol, and LH. Intraluteal PGF2 alpha caused premature functional luteolysis in all monkeys, as reflected by a highly significant decline in circulating progesterone and estradiol and the early onset of menstruation, when compared to the other groups. V, NCL, and SC infusions had no effect on either circulating steroid levels or luteal phase lengths. None of the experimental groups showed any change in plasma LH concentrations. These are the first data to indicate that PGF2 alpha can induce functional luteolysis in the primate, and the site of action appears to be the corpus luteum.     

Prostaglandins and post-abortion luteolysis in early pregnancy.

This study was undertaken to determine if post-abortion luteolysis in early pregnancy could be accelerated by the administration of 15(S)15-methyl-PGF2alpha(15-me-PGF2alpha) or delayed following pretreatment with indomethacin. Thirty-nine women were divided into four groups: 7 women were given 400mug 15-me-PGF2alpha extra-amniotically one hour prior to vacuum aspiration; 14 were pretreated with oral indomethacin (50 mg X4) over 24 hours; 7 were given indomethacin (50mg X 6) over 36 hours and 11 served as controls. Plasma progesterone and estradiol were measured at fixed intervals before and after abortion. There was a rapid drop in the plasma progesterone within the first hour after abortion followed by an exponential decline over the next 23 hours. The plasma estradiol fell rapidly duriing the same period. Under the experimental conditions of this study neither 15-me-PGF2alpha nor indomethacin exerted a significant effect on the decline in luteal function. These results are interpreted as suggesting that factors other than prostaglandins have a more significant role in post-abortion luteolysis.     

Trypanosome-induced increase in prostaglandin F(2alpha) and its relationship with corpus luteum function in the goat

Plasma progesterone and 13,14-dihydro-15-keto Prostaglandin F(2alpha) (PGFM) were measured in normal (uninfected) and Trypanosoma congolense -infected adult goats for a period of 121 d, from May to August, during the breeding season in Kenya. Chronic trypanosomiasis rapidly increased the baseline plasma PGFM levels and the occurrence of irregular PGFM peaks in several infected goats. Progesterone luteal levels declined rapidly from the second and subsequent cycles post patency. Estrous cycles also became irregular but predominately shorter (8 to 19 d) before cessation from the second to fourth cycle following infection. The PGFM levels were still high during the acyclic period in all goats when progesterone levels were very low (1.4 to 2.4 nmol/l). The reciprocal increase in peripheral PGFM and decline in progesterone in these goats would suggest, in part, a trypanosome-induced PGF(2alpha) mediated luteolysis, and the possible involvement of prostaglandins in trypanosome-induced infertility in female goats.