The control of temporal changes in oxytocin receptor concentrations and oxytocin-induced 13,14-dihydro-15-keto PGF_2α (PGFM) release was examined in ewes. One week after ovariectomy, 36 ewes were administered fluorogesterone acetate for 10 days followed by oestradiol (3 × 16 μg day⁻¹) for 2 days (pretreatment cycle). Day 0 was designated as the time of the final 'oestrous' oestradiol injection. Ewes were then treated for up to 12 days with progesterone (24 mg day⁻¹ maximum) with or without oestradiol (both hormones administered in 1 ml of corn oil i.m. at 8 h intervals) in a pattern known to simulate natural plasma profiles of the oestrous cycle. The three treatments were zero oestradiol, low oestradiol (12 μg day⁻¹ maximum), and high oestradiol (36 μg day⁻¹ maximum). Subgroups of four ewes from each treatment group were given 1 μg of oxytocin (i.v.) on day 10, 11 or 12 of the simulated cycle, and endometrial oxytocin receptor concentrations were determined in samples collected within 3 h of oxytocin administration. On day 10 only one ewe in each group exhibited a PGFM response to oxytocin, and the mean response was unaffected by the concentration of oestradiol administered. On days 11 and 12 there was a significant effect of oestradiol concentration (P < 0.05) on the pattern of PGFM release in response to oxytocin, the high oestradiol concentration causing a rapid increase in the concentration of PGFM following oxytocin administration. On day 12 the oestradiol concentration was positively correlated with the PGFM mean response (P < 0.01). Oxytocin receptor concentrations were positively correlated with the concentration of PGFM released on days 11 and 12. We conclude that the quantitative effect of oestradiol is mediated principally through the oestrogenic stimulation of uterine oxytocin receptors, although additional effects on post-receptor events cannot be excluded. These results demonstrate that the oestradiol concentration affects the timing, the magnitude and the pattern of the PGF_2α response to oxytocin in progesterone-treated ovariectomized ewes. In the natural cycle, a high oestradiol concentration may therefore be associated with an earlier onset of luteolysis.

A steroid-treated ovariectomized ewe model was used to investigate the role of progesterone pretreatment in the control of functional oxytocin receptor concentrations during the early luteal phase. Ovariectomized ewes (n = 28) were injected with oestradiol for 2 days (final injection = day 0) with or without progesterone pretreatment (progestagen sponge for 10 days). Ewes were then given high or low concentrations of progesterone combined with high, low or zero concentrations of oestradiol in a pattern known to simulate the early luteal phase profile (n = 4 per group). Ewes were given 1 μg oxytocin (i.v.) on day 4 and plasma was collected to assay 13,14-dihydro-15-keto PGF_2α. The concentration of progesterone and oestradiol administered had no effect on the concentration of 13,14-dihydro-15-keto PGF_2α following oxytocin administration (P > 0.05). However, the group that was not pretreated exhibited a small but significant 13,14-dihydro-15-keto PGF_2α response in comparison with the equivalent pretreated group (P < 0.05). In a subsequent study, ewes were divided into groups pretreated and not pretreated with progesterone; both groups were given oestrous concentrations of oestradiol and high concentrations of progesterone and oestradiol together. On day 0, 2, 3 or 4, ewes from each group (n = 3, 3, 4 and 4, respectively) were given 1 μg of oxytocin i.v., and the endometrium was collected to measure the binding of oxytocin receptors. Oxytocin caused a significant (P < 0.05) increase in the concentration of 13,14-dihydro-15-keto PGF_2α in all ewes on day 0 but not on days 2, 3 or 4. Oxytocin receptor concentrations were maximal on day 0 and basal by day 4. The decline in receptor concentrations occurred more rapidly in the progesterone-pretreated than in the ewes that were not pretreated. This study has shown that progesterone pretreatment alters the subsequent steroid hormone control of oxytocin receptor concentrations, and has identified the delayed decline in oxytocin receptor concentrations as the potential cause of premature luteolysis in ewes that are not pretreated.

The role of oestradiol in the control of premature luteolysis (previously shown to occur by the normal luteolytic mechanism involving PGF_2α and oxytocin) was investigated in anoestrous ewes induced to ovulate using GnRH (250 ng every 2 h for 24 h followed by 125 μg on day 0) without progesterone pretreatment. Seven ewes were administered charcoal stripped bovine follicular fluid (bFF) on days 1–5 (2 ml by s.c. injection every 8 h) together with oestradiol on days 2–4 (4 μg in 1 ml of corn oil by i.m. injection every 8 h). Ten ewes were treated with bFF and corn oil (as above), and ten ewes received saline and corn oil (control group). All ewes were treated with 1 μg oxytocin (i.v.) on day 4 and plasma was collected for measurement of 13,14-dihydro-15-keto PGF_2α (PGFM). Blood samples were collected for measurement of progesterone and oestradiol (day−2 to 15). The ewes in the control group that responded to GnRH formed either normal (50% of ewes) or short-lived (50% of ewes) corpora lutea identified by progesterone profiles. The proportion of ewes that displayed premature luteolysis was reduced (P < 0.05) by bFF treatment alone (to 11% of ewes), and increased (P < 0.001) by bFF plus oestradiol treatment (to 100%). bFF treatment suppressed oestradiol concentrations (P < 0.01), whereas bFF plus oestradiol treatment increased oestradiol concentrations (P < 0.001) on days 1–5. The high oestradiol concentrations appeared to stimulate the premature luteolytic mechanism as the mean PGFM response to oxytocin was higher in the ewes treated with bFF plus oestradiol than in the other two groups (P < 0.001). In addition, the control ewes that formed short-lived corpora lutea had higher oestradiol concentrations (days 1–5) than did ewes with normal corpora lutea (P = 0.05). This study suggests that short-lifespan corpora lutea are the result of increased oestrogenic stimulation of the luteolytic mechanism during the early luteal phase (following a lack of prior exposure to progesterone) which can be overcome by suppressing oestradiol secretion. This finding demonstrates that oestradiol plays a key role in the initiation of premature luteolysis, probably through stimulation of the prostaglandin response to oxytocin.

The mammalian reproductive system is sensitive to exposure to endocrine disrupting chemicals, particularly during sexual maturation. The purpose of this study was to examine reproductive function in second and third generation male and female mink exposed to pesticides from conception to maturity. The mink were fed untreated feed or feed treated with Lindane (1 mg kg⁻¹ day⁻¹), Carbofuran (0.05 mg kg⁻¹ day⁻¹ or Pentachlorophenol (1 mg kg⁻¹ day⁻¹) from the time they were weaned. The second generation mink had also been exposed to the pesticides in utero and from their mother's milk as their mothers were similarly fed pesticides, from 3 weeks before breeding. The third generation mink were the offspring of mink (second generation females) who had themselves undergone long-term exposure to pesticides from conception onwards. Blood samples and endocrine tissues were obtained at necropsy from both generations of mink. No overt signs of toxicity were seen. The pesticides did not affect the percentage of mink mated. Lindane treatment reduced the proportion of mated mink that subsequently whelped (P < 0.1) and the litter size of mink that whelped (P <0.05). Testis size was reduced in the Lindane-treated, third generation males (P < 0.05). Serum concentrations of cortisol, testosterone and oestradiol were not affected by any pesticide treatment; however, thyroxine concentration was reduced by Pentachlorophenol (P < 0.05). In conclusion, exposure of mink to Lindane from conception resulted in a decrease in reproductive efficiency when they were subsequently mated, leading to a 60% reduction in the number of kits born.

Two experiments investigate the effects of oxytocin and progesterone on premature luteolysis in ewes. In Expt 1, 20 anoestrous ewes were induced to ovulate by multiple injections of GnRH (250 ng i.v. every 2 h for 24 h) followed by a bolus injection of GnRH (125 μg, i.v.). Ten ewes received a continuous infusion of oxytocin from the day after the GnRH bolus injection and the other ten ewes were infused with saline. Oxytocin infusion had no significant effect on the proportion of ewes with short luteal phases (P > 0.05). All ewes that had luteal phases of normal duration from either group (n = 9) exhibited a transient increase in plasma concentrations of progesterone 2 h after insertion of the pump. In Expt 2, 25 anoestrous ewes were treated with GnRH as in Expt 1. Five ewes were pretreated with progestagen for 11 days and ten ewes received progesterone (12 mg, i.m.) 24 h after the bolus injection of GnRH. All animals received an oxytocin injection (1 μg, i.v.) on day 4 after the GnRH bolus. All five ewes that were pretreated with progestagen had normal luteal function and none exhibited a 13,14-dihydro-15-keto PGF_2α (PGFM) response to oxytocin. None of the ten ewes injected with progesterone had a normal luteal phase and six ewes exhibited a PGFM response to oxytocin. Four ewes in the control group had normal luteal function and three had short luteal phases. It is concluded that (1) administration of oxytocin from about the time of ovulation does not prevent premature luteal regression; (2) a transient increase in progesterone at about the time of ovulation is associated with luteal phases of normal duration; (3) a more extended exposure to progesterone at about the time of ovulation prevents normal luteal function and may inhibit luteinization and (4) pretreatment with progesterone prevents luteolysis by reducing the uterine response to oxytocin early in the luteal phase.

Mink are carnivores of agroforestry fringe habitats and are exposed to pesticides that biomagnify within the food chain. Some pesticides are thought to disrupt reproductive and endocrine functions. In Expt 1, four groups of mink (n = 10) were fed either a control diet, or diets treated with lindane (1 mg kg⁻¹ day⁻¹), carbofuran (0.05 mg kg⁻¹ day⁻¹) or pentachlorophenol (1 mg kg⁻¹ day⁻¹) from before breeding until weaning. Mink were mated twice, at 7–8 day intervals. The treatments had no effect on the proportion of mink accepting the first mating; however, lindane and pentachlorophenol caused a decrease in the percentage of females accepting the second mating. Lindane and pentachlorophenol caused a decrease in whelping rate, although litter size was not affected. Carbofuran had no effect on fertility. Mink that mated only once had a lower whelping rate than mink that mated twice; therefore, it could not be determined whether the decreased whelping rates were due to the lack of a second mating or to increased embryo loss. In Expt 2, two groups of mink (n = 15) were fed a control diet or a diet treated with lindane (1 mg kg⁻¹ day⁻¹) from before mating until weaning. Mink were mated twice on two consecutive days. Lindane did not affect mating response at either mating. Whelping rate, but not implantation rate, was decreased by the lindane treatment. The proportion of embryos lost after implantation (implantation scars not represented by kits at whelping) was increased by the lindane treatment. In conclusion, both lindane and pentachlorophenol decreased fertility in mink, and the lindane effect was primarily a result of embryo mortality after implantation.

The effect of oestradiol and progesterone concentrations on the uterine PGF_2α response to oxytocin was investigated by measuring 13,14-dihydro-15-keto PGF_2α (PGFM) secretion. One week after ovariectomy, 27 ewes were administered progestagen for 10 days followed by oestradiol for 2 days. Day 0 was designated as the time of the last 'oestrous' oestradiol injection. Six groups of ewes (n = 4) were then treated for 12 days with a high or low dose of progesterone (36 or 12 mg day⁻¹) either alone or with a high or low dose of oestradiol (36 or 12 μg day⁻¹) administered (in 1 ml of corn oil by i.m. injection, at intervals of 8 h) in a pattern designed to simulate a natural oestrous cycle profile. A control group (n = 3) was given corn oil alone. Ewes were treated with 1 μg oxytocin (i.v.) on days 4, 8 and 12 of the simulated cycle and plasma was collected for assay of PGFM. An oxytocin-induced PGFM response occurred only on day 12, when the response was suppressed by high doses of progesterone and stimulated by high oestradiol doses. There was a significant effect of progesterone (P < 0.05) and a highly significant effect of oestradiol (P < 0.01) on the pattern of PGFM release in response to oxytocin. Low progesterone/high oestradiol stimulated the largest and most sustained increase in PGFM following oxytocin. There was a significant relationship between the oestradiol:progesterone ratio and the mean PGFM response on day 12 (P < 0.05). This is the first demonstration of a quantitative effect of steroid hormone concentrations on the PGFM response to oxytocin in ewes, and indicates that in early pregnancy, ewes with a high oestradiol:progesterone ratio may generate larger PGF_2α episodes thus increasing the risk of a failure of the maternal recognition of pregnancy.

Between 6 and 20 weeks of age an early increase in LH secretion has been reported in Hereford bull calves. Delaying this early increase in LH secretion delays testicular development. This study was designed to determine whether a premature increase in LH secretion during the early postnatal period enhances testicular development. Ten age- and body weight-matched Hereford bull calves were divided into two groups. One group (n = 5) received 200 ng LH releasing hormone (LHRH) i.v. every 2 h for 14 days, between 4 and 6 weeks of age. On the basis of blood samples taken every 15 min for 10 h, mean serum LH and testosterone concentrations and LH pulse frequency were increased by LHRH treatment (P < 0.05). Serum concentrations of FSH were not significantly influenced by treatment (P >0.05). In treated animals at 24 weeks of age, mean serum testosterone concentrations and LH pulse amplitude were increased (P <0.05). The concentrations of spermatozoa in electroejeculates collected at 52 weeks of age were greater in LHRH-treated compared with control calves. Testicular growth was enhanced by LHRH treatment and histological evaluation of the testis at 54 weeks of age showed increased spermatogenesis and also larger numbers of Sertoli cells per tubule cross-section as a result of LHRH treatment. We conclude that treatment with LHRH before the early increase in LH secretion altered testicular development and suggest that the early increase in LH secretion in bull calves may be critical for initiating and regulating the progression of reproductive maturation.

There is controversy over the potential endocrine modulating influence of pesticides, particularly during sensitive phases of development. In this study, ram lambs were exposed to lindane and pentachlorophenol from conception to necropsy at 28 weeks of age. The rams (and their mothers) were given untreated feed (n = 7) or feed treated with 1 mg kg⁻¹ body weight per day of lindane (n = 12) or pentachlorophenol (n = 5). Semen was collected from 19 weeks onwards and reproductive behaviour was tested at 26 weeks. Serum was collected every 2 weeks and at 27 weeks every 15 min for 6 h during both day and night, and for 1 h before and 5 h after stimulation with GnRH, adrenocorticotrophic hormone and thyroid-stimulating hormone. The pesticides did not affect body weight and ejaculate characteristics, or cause overt toxicity. In pentachlorophenol-treated rams, scrotal circumference was increased. However, seminiferous tubule atrophy was more severe and epididymal sperm density was reduced in comparison with untreated rams at necropsy (P < 0.05). Thyroxine concentrations were lower in pentachlorophenol-treated rams than in untreated rams (P < 0.05). However, after thyroid-stimulating hormone treatment, the thyroxine response was unaltered. Reproductive behaviour was reduced in lindane-treated rams compared with control rams (P < 0.05). Serum LH and oestradiol concentrations during reproductive development, LH pulse frequency at 27 weeks and testosterone secretion after GnRH treatment were lower in lindane-treated rams than in untreated rams (P < 0.05). In summary, the effects of pentachlorophenol on the testis may be linked to a decrease in thyroxine concentrations, and reduced reproductive behaviour in lindane-treated rams may be related to decreased LH, oestradiol and testosterone concentrations.

The aim of the present study was to document ovarian antral follicle dynamics throughout seasonal anoestrus in sheep. Daily transrectal ultrasonography was performed during four 17 day scanning periods from March to July in Western White-faced crossbred ewes. Blood samples were collected each day with ultrasonographic scanning for measurement of serum concentrations of FSH, oestradiol and progesterone. Blood samples were also taken every 15 min for 6 h, mid-way through each period of ultrasonographic examination, to determine the patterns of secretion of gonadotrophic hormones. Hormonal data were then related to observed changes in follicular populations and the patterns of antral ovarian follicle turnover. Ultrasonography showed that the ovaries of anoestrous ewes remained active and that the largest ovarian antral follicles grew to a periovulatory size (≥ 5 mm in diameter) at all stages of anoestrus. The total number of all ovarian follicles ≥ 3 mm in diameter was lower during early anoestrus compared with at mid-anoestrus because of a significantly smaller number of small (3 mm) and medium (4 mm) ovarian follicles. The largest ovarian follicles (attaining ≥5 mm in diameter before regression) exhibited a wave-like pattern of growth; an average of three waves of follicular development were recorded in sheep during each of the four 17 day scanning periods in anoestrus, with follicular waves emerging approximately every 5 days. This rhythmic pattern of follicular emergence was found to be associated with the occurrence of fluctuations in serum FSH concentrations. The growth rate of the largest follicles of the wave increased significantly from early to late anoestrus in sheep. In addition, ovarian follicles not growing beyond 3 mm in diameter showed organized patterns of growth and regression; their numbers tended to be lower (P = 0.09) at 3 days before and on the day of follicular wave emergence. Some ewes were seen to maintain synthesis of progesterone throughout anoestrus. This submaximal progesterone secretion tended to occur at irregular intervals and was not coupled with changes in concentrations or patterns of gonadotrophin release, ovulations or detectable morphological luteinization of ovarian antral follicles. It was concluded that the growth of ovarian antral follicles to an ovulatory size was maintained throughout anoestrus in ewes, with a transient shift in the number of small and medium-sized follicles during mid-anoestrus, and that the periodic emergence of waves of large follicles (≥ 5 mm in diameter) occurred in synchrony with an endogenous rhythm of FSH secretion.