Increased ovulation rate in androstenedione-immune ewes is not due to elevated plasma concentrations of FSH

Two experiments were undertaken to determine the hormonal response of Merino ewes to immunization against androstenedione (Fecundin). In Exp. 1 peripheral concentrations of LH, FSH and progesterone were monitored in spontaneously cycling ewes (20 immunized and 21 controls). In Exp. 2 (10 immunized and 10 controls) the same hormones were measured in ewes before and after prostaglandin (PG)-induced luteolysis and, in addition, the pattern of pulsatile LH secretion was determined during the luteal (PG + 12 days), early follicular (PG + 24 h) and late follicular (PG + 40 h) phase of the oestrous cycle. Ovulation rates were measured in both experiments. The results of these experiments indicate that androstenedione-immune animals have elevated ovulation rates (0.6-0.7 greater than control animals; P less than 0.05) associated with elevated plasma concentrations of LH and progesterone. The magnitude of the increase in plasma progesterone was correlated with androstenedione antibody titre (r = 0.6, P less than 0.001). LH pulse frequency of androstenedione-immune ewes tended to be higher at all stages of the oestrous cycle, but this difference was only significant (P less than 0.05) during the luteal phase. Mean plasma concentrations of FSH did not differ significantly between immunized and control ewes at any stage of the cycle. Analysis of periodic fluctuations in FSH during the luteal phase revealed that androstenedione-immune animals had a similar number of fluctuations of a similar amplitude to those of control animals, but the nadir of these fluctuations was lower (P less than 0.05) in immunized animals. A significant (P less than 0.05) negative correlation existed between androstenedione antibody titre and the interval between FSH peaks (r = -0.49) and androstenedione antibody titre and FSH nadir concentrations (r = -0.46). It is concluded that plasma FSH concentrations are not a determinant of ovulation rate in androstenedione-immune ewes and that increased LH concentrations, or perturbation of normal intraovarian mechanisms, may be responsible for the increase in ovulation rate observed in ewes immunized against androstenedione.     

Steroid secretion rates and plasma binding activity in androstenedione-immune ewes with an autotransplanted ovary

Mature Merino ewes in which the left ovary and its vascular pedicle had been autotransplanted to the neck were divided into control (N = 5) and immunized groups (N = 6). The immunized ewes were treated (2 ml s.c.) with Fecundin 1 and 4 weeks before the start of blood sampling. Ovarian and jugular venous blood was collected every 10 min at two stages of the follicular phase (21-27 h and 38-42 h after i.m. injection of 125 micrograms of a prostaglandin (PG) analogue) and during the mid-luteal phase (8 h at 15-min intervals). The ewes were monitored regularly for luteal function and preovulatory LH surges. Hormone concentrations and anti-androstenedione titres were assayed by RIA and ovarian secretion rates of oestradiol-17 beta, progesterone and androstenedione were determined. After the booster immunization, progesterone increased simultaneously with titre in immunized ewes, reaching 30 ng/ml at the time of PG injection when median titre was 1:10,000. All ewes responded to PG with LH surges 42-72 h later: 2 of the immunized ewes then had a second LH surge within 3-4 days at a time when peripheral progesterone values were 2-3 ng/ml. The frequency of steroid and LH pulses was greater in immunized ewes (P less than 0.05) during the luteal phase but not the follicular phase. The secretion rate of androstenedione was 6-10 times greater (19-37 ng/min; P less than 0.001) in immunized ewes at all sampling stages. Progesterone secretion rates were 3 times greater (16 micrograms/min; P less than 0.001) during the luteal phase in immunized ewes. The amplitude of oestradiol pulses was significantly reduced in immunized ewes (4.8 vs 2.1 ng/min at +24 h and 6.5 vs 2.8 ng/min at +40 h in control and immunized ewes, respectively: P less than 0.05) during the follicular phase. However, the mean secretion rate of oestradiol at each phase of the cycle was not significantly different between treatment groups. Analysis of bound and free steroid using polyethylene glycol showed that greater than 98% of peripheral and ovarian venous androstenedione and 86% of peripheral progesterone was bound in immunized ewes but there was no appreciable binding (less than 0.1%) in control ewes. Similarly, 50% of ovarian venous oestradiol was bound in immunized ewes compared to 15% in control ewes. We conclude that immunization against androstenedione increases the secretion rate of androstenedione and progesterone but not of oestradiol.(ABSTRACT TRUNCATED AT 400 WORDS)     

Effect of oxytocin infusion on secretion of progesterone and luteinizing hormone and the concentration of uterine oxytocin receptors during the periovulatory period in cloprostenol-treated ewes.

Oxytocin infusions were initiated on day 10 of the oestrous cycle in ewes, and luteal regression was induced by injection of 100 micrograms cloprostenol on day 12. Blood samples were collected at frequent intervals via an indwelling jugular vein cannula to measure concentrations of progesterone and luteinizing hormone (LH) during the luteal and follicular phases in saline (n = 6) and oxytocin (n = 5) infused animals. The oxytocin infusion maintained peripheral plasma concentrations of 53 +/- 3.2 pg oxytocin ml-1 (mean +/- SEM) compared with values of about 1 pg ml-1 during oestrus in control ewes. Oxytocin infusion had no effect on luteal phase progesterone concentrations, the timing of luteolysis, basal luteinizing hormone (LH) secretion, LH pulse frequency, or the timing or height of the LH surge. Treated ewes came into oestrus significantly earlier than controls (P < 0.05) but ovulated normally. Uterine samples collected 96 h after cloprostenol injection (approximately day 2 of the cycle) showed that oxytocin receptor concentrations were significantly higher in the endometrium in ewes that had been given a 5 day oxytocin infusion than in control animals (556 and 262 fmol mg-1 protein, respectively: geometric means from ANOVA, P < 0.001), whereas myometrial receptor concentrations were not affected (113 and 162 fmol mg-1 protein, respectively). We conclude that the previously reported delay in luteal development caused by oxytocin infusion (Wathes et al., 1991) is not due to the inhibition or delay of ovulation, but must instead occur via a direct influence on the developing corpus luteum.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of immunization against progesterone on oestrus, cycle length, ovulation rate, luteal regression and LH secretion in the ewe

The effects of active immunization against progesterone on reproductive activity were studied in Merino ewes. Immunization against progesterone caused a shortening (P less than 0.01) of the interval between ovulations from 17-18 days (controls) to between 6 and 10 days (immunized group); this was associated with a corresponding reduction in the interval between LH surges. The immunized ewes also had higher (P less than 0.05) ovulation rates (1.72) than controls (1.25) and exhibited a reduced (P less than 0.01) incidence of oestrus (26% v. 95%). Many immunized ewes continued to ovulate despite the persistence of corpora lutea from earlier ovulations which led to an accumulation on the ovaries of many corpora lutea of different ages. The frequency of LH pulses in ewes immunized against progesterone (1.8 +/- 0.2 pulses/4 h) was significantly (P less than 0.001) higher than that of control ewes (0.3 +/- 0.1 pulses/4 h). This study highlights the importance of progesterone in the control of oestrus, ovulation, ovulation rate, luteal regression and the secretion of LH in the ewe.     

Administration of recombinant bovine interferon-alpha I at the time of maternal recognition of pregnancy inhibits prostaglandin F2 alpha secretion and causes luteal maintenance in cyclic ewes.

The antiluteolytic protein, ovine trophoblast protein-1, which is secreted by sheep embryos at about the time of the maternal recognition of pregnancy, exhibits significant structural homology with alpha interferons. Experiments were conducted to examine the effects of intra-uterine and systemic administration of a recombinant bovine interferon-alpha I (rboIFN-alpha I) upon the interoestrus interval, endometrial oxytocin receptor concentrations and secretion of prostaglandin (PG) F2 alpha in cyclic ewes. In Expt 1, each ewe had a cannula placed in the tip of a uterine horn ipsilateral to a corpus luteum, 7 days after an induced oestrus. From day 9 after oestrus until day 19, ewes received either 200 (n = 4), 667 (n = 5) or 2000 (n = 9) micrograms/24 h of rboIFN-alpha I, meclofenamic acid (n = 4) or vehicle (n = 11). Other ewes received 2000 micrograms rboIFN-alpha I/24 h (n = 5) between days 12 and 15 only. All ewes were killed on day 19. Mean luteal phase, as determined by daily plasma progesterone measurements, was significantly longer (P less than 0.01) and mean concentrations of 13,14-dihydro-15-keto PGF 2 alpha (PGFM) in plasma were lower (P less than 0.05) in ewes receiving 667 or 2000 micrograms rboIFN-alpha I between days 9 and 19, or 2000 micrograms between days 12 and 15, than in animals from other treatment or control groups. A similar protocol was used in Expt 2, in which further ewes received either 2000 micrograms rboIFN-alpha I/24 h (n = 5) or vehicle (n = 5) by bolus infusions twice a day into one uterine horn. Mean luteal phase was significantly (P less than 0.05) longer in treated than in control animals, but differences in PGFM concentrations were not significant. In Expt 3, after a synchronized oestrus, ewes received either 2.5 mg rboIFN-alpha I by i.m. injection twice a day between days 12 and 15 (n = 10), 2.5 mg rboIFN-alpha I by i.m. injection twice a day between days 9 and 15 (n = 11), i.m. injection of vehicle alone twice a day (n = 20), or continual intra-uterine infusion of 2 mg rboIFN-alpha I/day between days 12 and 15 (n = 7). The mean luteal phase of ewes receiving rboIFN-alpha I by intrauterine infusion or i.m. injection between days 9 and 15 was significantly longer than for animals from the other two groups (P less than 0.05).(ABSTRACT TRUNCATED AT 400 WORDS)     

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

Anoestrous Romney Marsh ewes with and without progesterone treatment (+P, -P) were treated with small-dose (250 ng) multiple injections of GnRH at 2-h intervals for 48 h. Animals were slaughtered on Days 4, 5, 7 and 11 after the end of GnRH treatment and luteal function was assessed by the measurement of daily plasma progesterone concentrations. In all animals which ovulated (29/32, 91%) peripheral progesterone concentrations rose to 0.5-1.0 ng/ml within 3 days of the end of GnRH treatment. In 7/7 (100%) +P animals and 5/22 (23%) -P animals, progesterone concentrations continued to rise and were maintained at levels greater than 1.5 ng/ml until slaughter. In the remaining -P animals, plasma progesterone concentrations declined to reach basal levels by Day 5. Corpora lutea recovered from these animals showed signs of premature regression on Day 5 and were fully regressed by Day 7. Progesterone priming delayed the occurrence of the LH surge which occurred 39.1 +/- 3.6 h after the end of GnRH treatment in the +P animals compared to 20.2 +/- 1.74 h (P less than 0.001) in the -P animals in which luteal function was abnormal and 22.4 +/- 4.35 h in the -P animals in which luteal function was normal. These results show that abnormal luteal function occurs in the majority of GnRH-treated ewes in the absence of progesterone pretreatment.(ABSTRACT TRUNCATED AT 250 WORDS)     

Influence of oxytocin infusion during oestrus and the early luteal phase on progesterone secretion and the establishment of pregnancy in ewes

In Experiment 1, an osmotic minipump containing oxytocin was implanted s.c. in ewes for 12 days beginning on Day 10 of the oestrous cycle, producing approximately 100 pg oxytocin/ml in the plasma. Two days after the start of infusion, all ewes were injected with 100 micrograms cloprostenol and placed with a fertile ram. At slaughter 22 days later, 9 (75%) of the 12 control (saline-infused) ewes were pregnant compared with 1 (11%) of the 9 ewes infused with oxytocin. In the control group, midcycle plasma concentrations of oxytocin were significantly higher in nonpregnant than in pregnant ewes. In Experiment 2, an infertile ram was used throughout to avoid any possible effects of pregnancy and oxytocin infusions were given at different stages of the oestrous cycle. Otherwise the protocol was similar to that in Exp. 1. Oxytocin infusion during luteolysis and the early follicular phase had no effect on the subsequent progesterone secretion pattern, but infusions beginning the day before cloprostenol-induced luteolysis and lasting for 7 or 12 days and infusions beginning on the day of oestrus for 4 days all delayed the subsequent rise in plasma progesterone by approximately 3-4 days. In these animals, the cycle tended to be longer. It was concluded that an appropriate oxytocin secretion pattern may be necessary for the establishment of pregnancy in ewes and that a high circulating oxytocin concentration during the early luteal phase delays the development of the young corpus luteum.     

Temporal relationships between peripheral plasma concentrations of oxytocin, progesterone and 13, 14-dihydro-15-keto-prostaglandin F2α during the oestrous cycle and early pregnancy in the ewe

Peripheral plasma concentrations of oxytocin, 13,14-dihydro-15-keto-prostaglandin F_2α(PGFM), progesterone and LH were determined at 3 hourly intervals during the oesterous cycle (n = 3) and in early pregnancy (n = 4) in sheep. The progesterone and LH concentrations showed that the cycling ewes were samples during the periods of luteal regression (decreasing progesterone concentrations), the preovulatory gonadotrophin surge and the beginning of the next luteal phase (increasing progesterone concentrations). The pregnant ewes had basal LH concentrations and luteal phase concentrations of progesterone (>lng/ml afte day 5 following mating) throughout the whole of the sampling period. Oxytocin concentrations in the non-pregnant ewes decreased around the time of luteal regression to reach low concentrations (mean concentrations of approximately 18pg/ml) during the preovulatory period and then increased after the preovulatory surge. PGFM concentrations exhibited a pulsatile pattern with increasing concentrations as progesterone levels fell. In the pregnant ewes oxytocin concentrations gradually fell until approximately 16 days post-mating (approximately 7–8pg/ml). The magnitude of the pulses in PGFM concentrations were also lower than in the cycling ewes. These results demonstrate that the increased concentrations of PGFM which are found during the period of luteal regression are not caused by increased peripheral concentrations of oxytocin.     

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.     

Pulsatile release of oxytocin into the circulation of the ewe during oestrus, mating and the early luteal phase

Two experiments were designed to investigate release patterns of oxytocin into plasma during oestrus and the early luteal phase. In Exp. 1, blood samples were collected from 5 ewes every 30 min for 10 h during 6 days around oestrus and the early luteal phase. During oestrus concentrations of oxytocin were generally low (1.27 +/- 0.54 pg/ml; mean +/- s.d.) but with occasional pulses up to 6 pg/ml. By Day 5 mean basal concentrations had risen to 4.5 +/- 2.1 pg/ml with a fluctuating release pattern. In Exp. 2, a method was developed for continuous blood sampling from conscious, unrestrained ewes. On the predicted day of oestrus following an untreated oestrous cycle, 8-ml blood samples were collected every minute for two 35-min periods (8 ewes: 16 sampling periods). For 6 ewes a ram was introduced to the pen for part of this time, and resulting behaviour was recorded. Additional blood samples were assayed for LH and progesterone to determine the stage of the cycle. Overall mean oxytocin concentrations ranged from 1.5 +/- 0.53 to 6.8 +/- 5.25 pg/ml in different animals. Ewes which were both in oestrus and exposed to the ram showed a pulsatile oxytocin release pattern consisting of low baseline concentrations with short-duration pulses superimposed (duration 1-4 min; amplitude 2.5-31.7 pg/ml; frequency 3.18/h). Coitus was not temporally associated with pulsatile release. However, the importance of the presence of the ram was indicated by total separation of 2 oestrous ewes from the ram until after experimentation. In these animals only 1 pulse of oxytocin was detected in 2.7 h of sampling. It is concluded that, although mean oxytocin concentrations at oestrus were low, short duration pulses were released into the plasma at this time. This effect may be dependent on the presence of a ram.     

Oxytocin is luteolytic in the rhesus monkey (Macaca mulatta)

Oxytocin (10 mi.u./microliter/h) or vehicle (0.5% chlorobutanol in saline, 1 microliter/h) was chronically infused directly into the corpus luteum of normally cyclic rhesus monkeys, by means of an Alzet pump-ovarian cannula system. Infusion of oxytocin (N = 6) or vehicle (N = 5) began 6 days after the preovulatory oestradiol surge, and daily peripheral blood samples were taken. Oxytocin caused a significant (P less than 0.05) decrease in progesterone, beginning 1 day after treatment, and oestradiol after 4 days; progesterone and oestradiol remained significantly depressed until menstruation. However, peripheral LH concentrations remained unchanged. The duration of the luteal phase, menstrual cycle and the onset of menses from the initiation of oxytocin infusion were significantly (P less than 0.01) shorter when compared to those of vehicle-treated controls. These results show that oxytocin can induce functional luteolysis in the primate and supports the hypothesis that oxytocin of luteal origin may play a role in spontaneous luteolysis.     

The active immunization of the cyclic ewe against an estrone protein conjugate

Four mature, cyclic ewes were given injections (I.M.) of a conjugate of 1,3,5 (10)-estratrien-3-ol-6,17-dione, 6 carboxyoxime bovine serum albumin (immunized ewes) on day 3 after estrus, and at days 10, 20, 40, 58, 91 and 134 after this initial treatment. Six control ewes treated with carrier emulsion alone continued to cycle normally. Three of the immunized ewes failed to exhibit estrus, an associated preovulatory surge of LH and ovulation. One ewe showed 1 abnormally short estrous period and then became anestrus. Injection of an estrone-protein-conjugate at days 3 and 13 after estrus did not appear to interfere with the rate of structural luteolysis of the corpus luteum present, but plasma concentrations of progesterone reached abnormally high luteal phase levels and in 2 ewes failed, subsequently to decline to normal follicular phase levels. Estrone binding capacity rose as early as day 9 after first treatment, and concentrations of LH rose as early as day 14. Subsequently, plasma levels of LH, estrone and progesterone and antisera titer rose; the only significant cross reaction of the antisera was with estradiol 17beta (11.32 +/- 2.80%).     

Continuous infusion of oxytocin prevents induction of uterine oxytocin receptor and blocks luteal regression in cyclic ewes

Continuous intravenous infusion of oxytocin (3 micrograms/h) between Days 13 and 21 after oestrus delayed return to oestrus by 7 days (length of cycle 23.3 +/- 0.6 days compared to 16.6 +/- 0.2 days in control ewes). At a lower infusion rate (0.3 micrograms/h) oxytocin delayed luteolysis in only 2 of 5 ewes. Treatment from Day 14, when luteolysis had already begun, was ineffective. Delay of luteal regression by oxytocin had no effect on the length of subsequent cycles. Measurement of circulating progesterone concentrations and luteal weight showed that prolongation of the oestrous cycle was due to prevention of luteal regression. Luteal regression and behavioural oestrus were induced during continuous oxytocin administration begun on Day 13 when cloprostenol was given on Day 15 (mean cycle length, 17.3 +/- 0.21 days). Continuous oxytocin infusion from Day 13 blocked the rise in uterine oxytocin receptor concentrations which normally precedes oestrus. Mean receptor concentrations in caruncular and intercaruncular endometrium and in myometrium were 76, 36 and 9 fmol/mg protein on Day 17 in ewes receiving continuous oxytocin (3 micrograms/h); in control ewes these values were 675, 638 and 130 fmol/mg protein respectively at oestrus. Receptor concentrations on the day of oestrus in ewes receiving oxytocin and cloprostenol were not significantly different from those in control ewes (649, 852, and 109 fmol/mg protein respectively). Since cloprostenol, a PGF-2 alpha analogue, overcame the antiluteolytic action of oxytocin, it is suggested that continuous oxytocin treatment may inhibit uterine production of PGF-2 alpha, possibly by down regulating the uterine oxytocin receptor.(ABSTRACT TRUNCATED AT 250 WORDS)     

Growth differentiation factor 9 and bone morphogenetic protein 15 are essential for ovarian follicular development in sheep

The aim of this study was to test the hypothesis that both growth differential factor 9 (GDF9) and bone morphogenetic protein (BMP15; also known as GDF9B) are essential for normal ovarian follicular development in mammals with a low ovulation rate phenotype. Sheep (9-10 per group) were immunized with keyhole limpet hemocyanin (KLH; control), a GDF9-specific peptide conjugated to KLH (GDF9 peptide), a BMP15-specific peptide conjugated to KLH (BMP15 peptide), or the mature region of oBMP15 conjugated to KLH (oBMP15 mature protein) for a period of 7 mo and the effects of these treatments on various ovarian parameters such as ovarian follicular development, ovulation rate, and plasma progesterone concentrations evaluated. Also in the present study, we examined, by immunohistochemistry, the cellular localizations of GDF9 and BMP15 proteins in the ovaries of lambs. Both GDF9 and BMP15 proteins were localized specifically within ovarian follicles to the oocyte, thereby establishing for the sheep that the oocyte is the only intraovarian source of these growth factors. Immunization with either GDF9 peptide or BMP15 peptide caused anovulation in 7 of 10 and 9 of 10 ewes, respectively, when assessed at ovarian collection. Most ewes (7 of 10) immunized with oBMP15 mature protein had a least one observable estrus during the experimental period, and ovulation rate at this estrus was higher in these ewes compared with those immunized with KLH alone. In both the KLH-GDF9 peptide- and KLH-BMP15 peptide-treated ewes, histological examination of the ovaries at recovery (i.e., approximately 7 mo after the primary immunization) showed that most animals had few, if any, normal follicles beyond the primary (i.e., type 2) stage of development. In addition, abnormalities such as enlarged oocytes surrounded by a single layer of flattened and/or cuboidal granulosa cells or oocyte-free nodules of granulosa cells were often observed, especially in the anovulatory ewes. Passive immunization of ewes, each given 100 ml of a pool of plasma from the GDF9 peptide- or BMP15 peptide-immunized ewes at 4 days before induction of luteal regression also disrupted ovarian function. The ewes given the plasma against the GDF9 peptide formed 1-2 corpora lutea but 3 of 5 animals did not display normal luteal phase patterns of progesterone concentrations. The effect of plasma against the BMP15 peptide was more dramatic, with 4 of 5 animals failing to ovulate and 3 of 5 ewes lacking surface-visible antral follicles at laparoscopy. By contrast, administration of plasma against KLH did not affect ovulation rate or luteal function in any animal. In conclusion, these findings support the hypothesis that, in mammals with a low ovulation rate phenotype, both oocyte-derived GDF9 and BMP15 proteins are essential for normal follicular development, including both the early and later stages of growth.     

Oxytocin infusion from day 10 after oestrus extends the luteal phase in non-pregnant cattle

Oestrus was synchronized in 8 cyclic heifers by progesterone treatment (PRID), after which the animals were monitored for one control cycle to measure the inter-oestrous interval. Osmotic minipumps containing saline (controls, N = 3) or oxytocin (N = 5) were implanted subcutaneously on Day 10 of the second cycle, and removed 12 days later. Jugular venous blood samples were collected daily for measurement of progesterone, and every 2 days for oxytocin. In addition, blood samples were taken every 10 min from 1 h before to 3 h after minipump insertion for measurement of plasma 15-keto-13,14-dihydroprostaglandin-F-2 alpha (PGFM) and every 30 min over the same period for measurement of progesterone and oxytocin. The lengths of the first untreated cycle in both groups of heifers were 20.2 +/- 0.56 (mean +/- s.e.m.) days compared with 25.4 +/- 0.81 days after oxytocin treatment (P less than 0.001). Oxytocin plasma concentrations in treated animals rose from less than 10 pg/ml to 70-500 pg/ml by 2 h after the start of oxytocin infusion and remained elevated until treatment was withdrawn. There was no increase in PGFM concentrations immediately after minipump insertion. Plasma progesterone concentrations were similar in treated and control animals but remained at mid-luteal levels for an average of 5 days longer in treated heifers. It is concluded that continuous administration of oxytocin can extend the luteal life-span in cattle.     

Pattern of gonadotropin-releasing hormone (GnRH)-like stimuli sufficient to induce follicular growth and ovulation in ewes passively immunized against GnRH.

The pattern of GnRH-like stimuli capable of inducing follicular growth, ovulation, and luteal function was evaluated in ewes passively immunized against GnRH. The estrous cycles of 30 regularly cyclic sheep were synchronized using vaginal pessaries impregnated with a synthetic progestogen. Animals were passively immunized against GnRH (groups 2-5, n = 6) or the carrier protein, keyhole limpet hemocyanin (KLH; group 1, n = 6), at the time of pessary removal (PR). Circhoral delivery of saline (groups 1, 2, and 5) or low amplitude GnRH agonist (des-Gly10 GnRH ethylamide [100 ng/hourly pulse]; groups 3 and 4) was initiated at PR and continued for 3 (groups 4 and 5) or 12 days (groups 1-3). In groups 4 and 5, the amplitude of the GnRH-like stimulus was increased to 800 ng/hourly pulse (stimulus-shift) during the 24-h period beginning 72 h after PR. The amplitude of the hourly stimulus was adjusted to 100 ng/pulse 96 h after PR and continued at that level to Day 12. The endocrine changes associated with follicle growth and maturation (serum concentrations of estradiol [E2] above 10 pg/ml), ovulation (surge-like secretion of LH and FSH), and normal luteal function (serum concentrations of progesterone [P] above 2 ng/ml) were evident in ewes passively immunized against KLH (group 1). In this group, the preovulatory surge of gonadotropins was noted 48.7 +/- 1.2 h after PR. These endocrine events were blocked by passive immunization against GnRH (group 2).(ABSTRACT TRUNCATED AT 250 WORDS)     

Does the first LH surge of the breeding season initiate the first full-length cycle in the ewe?

To determine whether the first LH surge of the breeding season initiates a transient rise in progesterone in most ewes, serum progesterone (daily) and LH (every 4 h) concentrations were measured in samples collected from 7 ewes between 19 July and first oestrus or 8 September, whichever came first. In 6 of the 7 ewes, the first LH surge of the breeding season was followed within 5 days by a transient, 2-day rise in progesterone. Within less than 5 (N = 4), or 9 (N = 1) or 10 (N = 1) days later, a second LH surge occurred, which was similar in maximum amplitude and duration to the first surge, and which initiated the first full-length luteal phase of the breeding season. In the remaining ewe, the first LH surge of the breeding season induced an abbreviated (9 days) and insufficient (maximum progesterone, 0.94 ng/ml) luteal phase. These results demonstrate that most ewes have more than one LH surge before the first full-length luteal phase, the first surge inducing a transient rise in progesterone. Therefore, although the seasonal decrease in response to oestradiol negative feedback is sufficient for initiation of the first LH surge of the breeding season, additional endocrine mechanisms may be necessary to induce the first full-length luteal phase.     

Does inadequate luteal function limit the establishment of pregnancy in the early post-partum ewe?

In Exp. 1 the effect of lactation versus early weaning on luteal function was examined in seasonally anoestrous Finn Dorset ewes that were induced to ovulate at 21 (N = 14) or 35 (N = 14) days post partum by using a CIDR device and PMSG. Prolactin concentrations were significantly higher (P less than 0.001) in lactating compared with early weaned ewes throughout the study. The proportion of lactating ewes with inadequate luteal function (as assessed by daily progesterone concentrations) in the 21-day group was 0.43 (3 or 7) compared with 0.67 (4 of 6) for those weaned within 2 days after parturition. Corresponding values for the 35-day group were 0 (0 of 4) and 0.14 (1 of 7) respectively. There was no evidence of abnormal luteal function in standard ewes (N = 8) for which the interval from parturition was greater than 150 days. In Exp. 2 we examined whether pregnancy can be successfully established during the breeding season following transfer of embryos into lactating or early weaned ewes in the early post-partum period. Embryos were donated from Border Leicester x Scottish Blackface ewes for which the interval from previous parturition was greater than 150 days. These embryos were transferred synchronously on Day 5 after behavioural oestrus to recipient ewes with the same breeding history as the donors (standard ewes, N = 15) or to lactating or early weaned recipients that had been induced to ovulate on Day 21 (N = 16) or 35 (N = 24) post partum. In the 21-day group inadequate luteal function was observed in 2 of 7 (0.28) lactating and 4 of 9 (0.44) early weaned ewes compared with corresponding values of 1 of 13 (0.08) and 2 of 11 (0.18) in the 35-day post-partum group. Luteal function was normal in all standard ewes. The proportion of successful pregnancies in the standard ewes was 0.80 (12 of 15) compared with 0 in lactating and early weaned ewes in the 21-day group and 0.08 (1 of 13) and 0.36 (4 of 11) respectively in the 35-day group. The incidence of inadequate luteal function is therefore independent of the suckling stimulus and is higher in ewes induced to ovulate on Day 21 than Day 35 post partum during breeding and non-breeding seasons.(ABSTRACT TRUNCATED AT 250 WORDS)     

Gonadotrophin concentrations and ovulation rates in Suffolk ewes actively or passively immunized against inhibin alpha

Mature Suffolk ewes were either actively or passively immunized against the synthetic fragment of porcine inhibin alpha, pI alpha(1-30), to determine the effects on gonadotrophin secretion and ovulation rate. Thirteen control ewes were immunized against human serum albumin, 12 ewes were actively immunized against pI alpha(1-30) and 36 ewes were passively immunized with pI alpha(1-30) antiserum. Blood samples were collected at 4-h intervals for 72 h from oestrus-synchronized ewes following the withdrawal of the progestagen pessaries. Mean gonadotrophin concentrations measured during the oestrous cycle of control ewes, ewes actively immunized against pI alpha(1-30) and ewes passively immunized against pI alpha(1-30) were similar, but their secretory profiles differed. Serum concentrations of follicle-stimulating hormone (FSH) were highest in ewes which had received antiserum at the time of pessary withdrawal; FSH concentrations did not decrease during the follicular phase of the oestrous cycle in ewes given antiserum 24 h after pessary withdrawal. Subtle but significant increments in serum FSH concentrations were observed in all passively immunized ewes in which sampling commenced at the time of treatment. The amplitude of the preovulatory luteinizing hormone (LH) peak, but not of the FSH peak, and the postovulatory secondary rise in FSH were lower (P less than 0.05) in actively immunized ewes than in control ewes. The mean (+/- s.e.) ovulation rate for actively immunized ewes (6.6 +/- 1.0) was 3 times higher (P less than 0.05) than that for control ewes (2.0 +/- 0.2), but was unaffected by passive immunization (range, 1.8-2.3).(ABSTRACT TRUNCATED AT 250 WORDS)     

Induction of luteal regression in the marmoset monkey (Callithrix jacchus) by a gonadotrophin-releasing hormone antagonist and the effects on subsequent follicular development

Doses of 100 or 200 micrograms of a novel GnRH antagonist ([N-acetyl-D beta Na11-D-pCl-Phe2-D-Phe3-D-Arg6-Phe7-Arg8-D-Ala10]NH2 GnRH) (4 animals/dose) were administered on Days 10/11 of the luteal phase and induced a marked suppression of circulating bioactive LH and progesterone concentrations within 1 day of treatment (P less than 0.01). Thereafter, progesterone concentrations remained low or undetectable until after the next ovulation. Similar results were obtained when 200 micrograms antagonist were given on Days 5/6 of the luteal phase (N = 4). The interval from injection of antagonist (200 micrograms but not 100 micrograms) to ovulation (based on a rise in progesterone above 10 ng/ml) was significantly longer than that from prostaglandin-induced luteal regression to ovulation in control cycles (N = 4/treatment) (range, 13-15 days after antagonist vs 8-10 days after prostaglandin, P less than 0.01). This delay of 4-5 days was equivalent to the duration for which LH concentrations were significantly suppressed by 200 micrograms antagonist when administered to ovariectomized animals (N = 3). Corpus luteum function during the cycle after GnRH antagonist treatment appeared normal according to the pattern of circulating progesterone. These results show that corpus luteum function and preovulatory follicular development in the marmoset monkey are dependent on pituitary gonadotrophin secretion.