Effect of ovine conceptus secretory proteins and purified ovine trophoblast protein-1 on interoestrous interval and plasma concentrations of prostaglandins F-2 alpha and E and of 13,14-dihydro-15-keto prostaglandin F-2 alpha in cyclic ewes

Conceptus secretory proteins (oCSP) were obtained from medium in which sheep conceptuses, collected on Day 16 of pregnancy, were cultured for 30 h. A portion of the culture medium (500 ml) was prepared for intrauterine infusion by concentrating the proteins by Amicon ultrafiltration (Mr 500 cutoff). A second portion (500 ml medium) was used to purify sheep trophoblast protein one (oTP-1). Proteins remaining after oTP-1 purification were concentrated and then passed through an anti-oTP-1 sepharose CL-4B affinity column to remove any remaining oTP-1 (oCSP-oTP-1). Serum proteins (oSP) were collected from a Day-16 pregnant ewe and diluted for infusion. Catheters were placed in the uterus of cyclic (Day 10) ewes. The following combinations of proteins were infused: 0.75 mg oCSP + 0.75 mg oSP (5 ewes), 0.75 mg oCSP - oTP-1 + 0.75 mg oSP (4 ewes), 0.05 mg oTP-1 + 1.45 mg oSP (5 ewes) and 1.5 mg oSP only (5 ewes). Infusions were twice daily on Days 12 and 13 (08:00 and 17:00 h) and once on Day 14 (08:00 h). On Day 14, ewes were injected intravenously at 08:00 h with 0.5 mg oestradiol-17 beta. Blood sampling began 30 min before oestradiol injection and continued every 30 min for 10 h. On Day 15 ewes received 10 i.u. oxytocin intravenously (08:00 h). Blood samples were collected 10 min before oxytocin and every 10 min for 1 h after oxytocin injection. Concentrations of prostaglandin (PG) F, PGE-2/PGE-1 (PGE) and 13,14-dihydro-15-keto-PGF-2 alpha (PGFM) were measured by specific radioimmunoassay. Ewes treated with oTP-1 and oCSP had longer (P less than 0.05) interoestrous intervals (27 and 25 days, respectively) compared to ewes treated with oSP and oCSP--oTP-1 (19 and 19 days, respectively) (s.e.m. = 1.56 days). These results indicate that oTP-1 alone is as potent as total conceptus secretory proteins in extending luteal maintenance. Ewes treated with oTP-1 and oCSP had no increase in PGF after oestradiol injection while production of PGF did increase 6-10 h after oestradiol in ewes treated with oSP and oCSP--oTP-1. PGFM was correlated with PGF concentrations (r = 0.57, P less than 0.01) although presence or absence of increases in production of PGFM for the treatment groups were not the same as those for PGF. No effects of treatment on PGE were detected.(ABSTRACT TRUNCATED AT 400 WORDS)     

Spontaneous electromyographic activity throughout the cycle in the sow and its change by intrauterine oestrogen infusion during oestrus

Two experiments were carried out to monitor influences on the uterine electromyographic activity (EMG) in cyclic gilts with chronic uterine EMG electrodes. In Exp. 1 the EMG was recorded continuously from Day -1 for 24 days and was evaluated for frequency, duration and amplitude. Progesterone and oestradiol in peripheral plasma were measured daily. As high amounts of oestrogens are characteristic for boar semen, in Exp. 2 the influence of seminal oestrogens on uterine contractions at Day 0 (first day of standing reflex) was investigated in gilts with chronic intrauterine catheters. They were infused with 10 ml saline (N = 4) or saline with physiological amounts of oestrogens (5 micrograms oestradiol + 2 micrograms oestrone + 4.5 micrograms oestrone sulphate; N = 4). Sham-treated gilts (infusion catheters, no infusion; N = 5) served as controls. The EMG was recorded for 2 h before and 9 h after infusion. In Exp. 1 the maximal amplitude (2040 +/- 98 microV) and duration (32 +/- 1.7 sec) but the lowest frequency (15.8 +/- 2.1 contractions/h) were found on Day 0. With decreasing oestrogen and increasing progesterone concentrations the frequency increased continuously until Day 5 (63.5 +/- 1.0 contractions/h) while the amplitude (183 +/- 13 microV) and duration (3.3 +/- 0.7 sec) decreased. During Days 6-13 the EMG activity was not detectable. The reverse pattern was found from the onset of luteolysis until the following Day 0. On Day 0 a significant correlation between oestradiol and the duration (r = 0.81; P less than 0.01; n = 10) but not the frequency was observed.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of FSH on ovarian inhibin secretion in anoestrous ewes

In Exp. 1, 7 Finn-Merino ewes which had one ovary autotransplanted to a site in the neck had jugular and timed ovarian venous blood samples collected at 10-min intervals for 2 h before and 3 h after injection of 5 micrograms NIAMDD-oFSH-S16. In Exp. 2, 8 Finn-Merino ewes with ovarian autotransplants had jugular and timed ovarian venous blood samples collected at 15-min intervals for 2 h before and 12 h after bolus injection of 40 micrograms NIAMDD-oFSH-S16 and infusion of oFSH-S16 at 6 micrograms/min for 4 h. In Exp. 2 the follicular population of the ovary was assessed by real-time ultrasound at the beginning and end of the experimental period. In both experiments the secretion rates of inhibin (1-3 ng/min) and oestradiol (0.5-8 ng/min) were similar to those observed during the luteal phase of the cycle in the breeding season, indicating significant follicular development in these animals. In Exp. 1 there was no change in the secretion of oestradiol or inhibin after the injection of FSH which resulted in a 25% increase (P less than 0.05) in the concentration of FSH in plasma. Inhibin secretion was pulsatile but there was no difference in inhibin pulse frequency before (1.6 +/- 0.2 pulses/h) or after (1.2 +/- 0.5 pulses/h) injection of FSH. In Exp. 2 injection of FSH resulted in an increase (P less than 0.001) in plasma concentrations of FSH in the sample taken 10 min after injection from a baseline of 1.2 +/- 0.2 ng/ml to a peak of 10.6 +/- 1.0 ng/ml (mean +/- s.e.m.).(ABSTRACT TRUNCATED AT 250 WORDS)     

Extension of oestrous cycles and prolonged secretion of progesterone in non-pregnant cattle infused continuously with oxytocin

The experimental objective was to evaluate how continuous infusion of oxytocin during the anticipated period of luteolysis in cattle would influence secretion of progesterone, oestradiol and 13,14-dihydro-15-keto-prostaglandin F-2 alpha (PGFM). In Exp. I, 6 non-lactating Holstein cows were infused with saline or oxytocin (20 IU/h, i.v.) from Day 13 to Day 20 of an oestrous cycle in a cross-over experimental design (Day 0 = oestrus). During saline cycles, concentrations of progesterone decreased from 11.0 +/- 2.0 ng/ml on Day 14 to 2.0 +/- 1.3 ng/ml on Day 23; however, during oxytocin cycles, luteolysis was delayed and progesterone secretion remained near 11 ng/ml until after Day 22 (P less than 0.05). Interoestrous interval was 1.6 days longer in oxytocin than in saline cycles (P = 0.07). Baseline PGFM and amplitude and frequency of PGFM peaks in blood samples collected hourly on Day 18 did not differ between saline and oxytocin cycles. In Exp. II, 7 non-lactating Holstein cows were infused with saline or oxytocin from Day 13 to Day 25 after oestrus in a cross-over experimental design. Secretion of progesterone decreased from 6.8 +/- 0.7 ng/ml on Day 16 to less than 2 ng/ml on Day 22 of saline cycles; however, during oxytocin cycles, luteolysis did not occur until after Day 25 (P less than 0.05). Interoestrous interval was 5.9 days longer for oxytocin than for saline cycles (P less than 0.05). In blood samples taken every 2 h from Day 17 to Day 23, PGFM peak amplitude was higher (P less than 0.05) in saline (142.1 +/- 25.1 pg/ml) than in oxytocin cycles (109.8 +/- 15.2 pg/ml). Nevertheless, pulsatile secretion of PGFM was detected during 6 of 7 oxytocin cycles. In both experiments, the anticipated rise in serum oestradiol concentrations before oestrus, around Days 18-20, was observed during saline cycles, but during oxytocin cycles, concentrations of oestradiol remained at basal levels until after oxytocin infusion was discontinued. We concluded that continuous infusion of oxytocin caused extended oestrous cycles, prolonged the secretion of progesterone, and reduced the amplitude of PGFM pulses. Moreover, when oxytocin was infused, pulsatile secretion of PGFM was not abolished, but oestrogen secretion did not increase until oxytocin infusion stopped.     

Uterine secretion of prostaglandin F2 alpha in response to oxytocin in ewes: changes during the estrous cycle and early pregnancy.

Experiment 1 was conducted to determine when the ovine uterus develops the ability to secrete prostaglandin F2 alpha (PGF2 alpha) in response to oxytocin and how development is affected by pregnancy. Pregnant and nonpregnant ewes received an injection of oxytocin (10 IU, i.v.) on Day 10, 13, or 16 postestrus. Jugular venous blood samples were collected for 2 h after injection for quantification of 13,14-dihydro-15-keto-PGF2 alpha (PGFM). In nonpregnant ewes, concentrations of PGFM increased following oxytocin on Day 16 but not on Day 10 or 13. Concentrations of PGFM did not increase following treatment on Day 10, 13, or 16 in pregnant ewes. Therefore, the ability of oxytocin to induce uterine secretion of PGF2 alpha develops after Day 13 in nonpregnant but not in pregnant ewes. Experiment 2 was conducted to precisely define when uterine secretory responsiveness to oxytocin develops. Pregnant and nonpregnant ewes received oxytocin on Day 12, 13, 14, or 15. In nonpregnant ewes, concentrations of PGFM increased following treatment on Days 14 and 15, but not earlier. Peripheral concentrations of progesterone showed that uterine secretory responsiveness to oxytocin developed prior to the onset of luteal regression. As in experiment 1, the increase in concentrations of PGFM following administration of oxytocin was much lower in pregnant than in nonpregnant ewes; however, some pregnant ewes did respond to oxytocin with an increase in PGFM. In experiment 3, pregnant ewes received an injection of oxytocin on Day 18, 24, or 30 postmating. Concentrations of PGFM increased following oxytocin on Days 18 and 24. The conceptus appears to delay and attenuate the development of uterine secretory responsiveness to oxytocin.     

Uteroplacental production of eicosanoids in ovine pregnancy

Dramatic cardiovascular alterations occur during normal ovine pregnancy which may be associated with increased prostaglandin production, especially of uteroplacental origin. To study this, we examined (Exp 1) the relationships between cardiovascular alterations, e.g., the rise in uterine blood flow and fall in systemic vascular resistance, and arterial concentrations of prostaglandin metabolites (PGEM, PGFM and 6-keto-PGF1 alpha) in nonpregnant (n = 4) and pregnant (n = 8) ewes. To determine the potential utero-placental contribution of these eicosanoids in pregnancy, we also studied (Exp 2) the relationship between uterine blood flow and the uterine venous-arterial concentration differences of PGE2, PGF2 alpha, PGFM, 6-keto-PGF1 alpha, and TxB2 in twelve additional late pregnant ewes. Pregnancy was associated with a 37-fold increase in uterine blood flow and a proportionate (27-fold) fall in uterine vascular resistance (p less than 0.01). Arterial concentrations of PGEM were similar in nonpregnant and pregnant ewes (316 +/- 19 and 245 +/- 38 pg/ml), while levels of PGFM and PGI2 metabolite 6-keto-PGF1 alpha were elevated 23-fold (31 +/- 14 to 708 +/- 244 pg/ml) and 14-fold (12 +/- 4 to 163 +/- 78 pg/ml), respectively (p less than 0.01). Higher uterine venous versus uterine arterial concentrations were observed for PGE2 (397 +/- 36 and 293 +/- 22 pg/ml) and 6-keto-PGF1 alpha (269 +/- 32 and 204 +/- 32 pg/ml), p less than 0.05, but not PGF2 alpha or TxB2. Although PGFM concentrations appeared to be greater in uterine venous (1197 +/- 225 pg/ml) as compared to uterine arterial (738 +/- 150 pg/ml) plasma, this did not reach significance (0.05 less than p less than 0.1). In normal ovine pregnancy arterial levels of PGI2 are increased, which may in part reflect increased uteroplacental production. Moreover the gravid ovine uterus also appears to produce PGE2 and metabolize PGF2 alpha.     

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)     

Progestagen synchronisation in the absence of a corpus luteum results in the ovulation of a persistent follicle in cyclic ewe lambs

Progestagens are widely used to synchronise oestrous in sheep but the effects on follicular dynamics are not clear. We tested the hypothesis that when luteolysis occurs early during progestagen synchronisation prolonged growth of the ovulatory follicle will occur. Cyclic ewe lambs (40.0+/-0.3 kg) were divided into three groups: eight ewes (Long group) received a progestagen sponge (60 mg medroxyprogesterone acetate) from Days 5 to 19 after oestrous and eight ewes (Short group) received a progestagen sponge on Day 5 which was replaced on Day 10 and again on Day 15, and removed on Day 19 after oestrous. On Days 6 and 7, ewes in both groups received prostaglandin. A third group (n=5, Control) did not receive any treatment. The growth and development of follicles > or =2 mm in diameter were characterised using daily transrectal ultrasonography. On Day 18, blood samples were collected every 12 min for 8 h from five ewes in the Long and Short groups. Data were analysed by ANOVA. The maximum diameter and age (emergence to ovulation) of the ovulatory follicle was greater (P<0.01) in ewes in the Long group (7. 4+/-0.2 mm and 12.1+/-0.6 days) than in ewes in the Short group (6. 3+/-0.2 mm and 5.1+/-0.5 days) and Control group (6.3+/-0.4 mm and 6. 8+/-0.6 days). On Day 18 of the cycle, LH pulse frequency and oestradiol concentrations were greater (P<0.05) in ewes in the Long group (3.2+/-1.1 pulse per 8 h and 1.15+/-0.09 pg ml(-1)) than the Short group (0.8+/-0.4 pulses per 8 h and 0.54+/-0.08 pg ml(-1)).We suggest that the negative feedback efficacy of a long-term progestagen sponge decreased with time and led to an increase in LH pulse frequency and prolonged growth of the ovulatory follicle. We conclude that, in the absence of luteal progesterone, synchronisation with a single progestagen sponge for 14 days resulted in higher LH pulse frequency and ovulation of a persistent follicle with a larger maximum diameter, compared with controls.     

Photoperiodic modification of negative and positive feedback effects of oestradiol on LH secretion in ovariectomized goats

Ovariectomized Shiba goats carrying an oestradiol implant (4-10 pg/ml) were kept under a short-day light regimen (10L:14D; Group 1, N = 4) or a long-day regimen (16L:8D; Group 2, N = 4). Plasma LH concentrations were lower (P less than 0.05) in Group 2 than in Group 1 between Days 40 and 200, suggesting an enhanced negative feedback effect of oestradiol on LH secretion under a long-day regimen. On Days 30, 60, 100, 149 and 279, an LH surge was induced by i.v. infusion of oestradiol for 48 h; the infusion rate was gradually increased from 0.5 (0 h) to 4.1 (48 h) micrograms/h, thereby mimicking the preovulatory increase of oestradiol secretion. The duration and magnitude of the induced LH surge were indistinguishable between the groups. The latency from the onset of oestradiol infusion to the LH surge was relatively constant in Group 1, 41.1 +/- 0.9 h (mean +/- s.e.m., n = 17) but was shorter in Group 2 (19.7 +/- 3.7 h, P less than 0.05) on Day 149; less oestradiol was therefore required for induction of the LH surge (27.4 vs 89.7 micrograms, P less than 0.01), suggesting an increased sensitivity to the oestradiol positive feedback under a long-day regimen. These results might be interpreted to indicate that the hypothalamic-pituitary axis of the goat becomes hypersensitive to the positive as well as the negative feedback effect of oestradiol under long-day conditions.     

Plasma 13,14-dihydro-15-keto-PGF alpha (PGFM) in pregnant and nonpregnant heifers prior to and during surgery and following intrauterine injection of PGF2 alpha

On day 17 postestrus or postmating, heifers were given intrauterine injections of saline (2 pregnant, 2 non-pregnant) or 200 micrograms PGF2 alpha (7 pregnant, 6 nonpregnant) through cannulae installed surgically into the uterine horn ipsilateral to the corpus luteum bearing ovary. Jugular blood samples were collected prior to the laparotomy at which the cannulae were installed during surgery, and for 90 min following the intrauterine injection. Plasma was assayed for progesterone and 13,14-dihydro-15-keto-PGF2 alpha (PGFM). Laparotomies were reopened to confirm proper cannula placement and to determine if blastocysts were present in mated heifers. Concentrations of PGFM were higher in pregnant compared to nonpregnant heifers during the presurgery (68 +/- 26 vs 24 +/- 26 pg/ml; P less than .025) and surgery (186 +/- 47 vs 65 +/- 17 pg/ml; P less than .05) periods. Pregnancy status did not alter the mean concentrations of PGFM (pregnant, 554 +/- 70 pg/ml; nonpregnant, 422 +/- 81 pg/ml) or the half-life of its decline in concentration (18 min) following intrauterine injection of PGF2 alpha. Pregnancy at 17 days in cattle does not appear to influence PGF2 alpha transport from the uterine lumen or its metabolism in the uterus or elsewhere in response to an acute intrauterine injection.     

In-vivo and in-vitro effects of ethanol on mouse preimplantation embryos

In Exp. 1A, hybrid mice (N = 10) were provided with food and 25% (v/v) ethanol as the only source of liquid for 72 h, beginning at the detection of the copulatory plug (08:00 h, Day 1). Control mice received food and tap water. Food consumption (P less than 0.001) but not total caloric intake (P greater than 0.05) was less for the alcohol-treated mice than the controls. Ethanol-derived calories averaged 35% of caloric intake during the 72 h of treatment. Alcohol-treated animals showed a dramatic weight loss until Day 5 while controls gained weight (P less than 0.05). Ethanol consumption did not influence pregnancy rate, litter size or litter weight. In Exp. 1B, animals were treated as in Exp. 1A, but were killed at various times between 24:00 h, Day 1, and 08:00 h, Day 4. Trunk blood was used to determine haematocrit and serum to determine alcohol concentration. Haematocrit was greater (P less than 0.05) for all alcohol-treated mice than for controls at all time periods sampled except one. Dehydration was therefore probably responsible for the weight loss seen in Exps 1A and 1B. Average blood alcohol concentrations fluctuated with time of day and day of treatment. Average maximum concentration was 91.4 mg ethanol/100 ml serum. In Exp. 2, hybrid mouse 2-cell embryos were cultured in vitro in 0 or 0.1% ethanol (Exp. 2A) and 0 or 1.0% ethanol (Exp. 2B) for 8 days.(ABSTRACT TRUNCATED AT 250 WORDS)     

Oxytocin-stimulated inositol phosphate turnover in endometrium of ewes is influenced by stage of the estrous cycle, pregnancy, and intrauterine infusion of ovine conceptus secretory proteins

Three experiments (Exp) assessed the influence of stage of the estrous cycle, pregnancy, and intrauterine infusion of ovine conceptus secretory proteins (oCSP) on turnover of inositol trisphosphate (the putative second-messenger for oxytocin-stimulated secretion of prostaglandin F2 alpha) in ovine endometrium during luteolysis and maternal recognition of pregnancy. In Exp 1, endometrium was collected from 5 cyclic (Cy) and 6 pregnant (P) ewes on Day 16 after onset of estrus. In Exp 2, endometrium was collected from Day 12 Cy (n = 5), Day 12 P (n = 3), Day 16 Cy (n = 4), and Day 16 P (n = 3) ewes. In Exp 3, 12 Cy ewes were allotted randomly, in a 2 x 2 factorial arrangement, to receive serum protein (SP), or oCSP and estradiol-17 beta (E2), or vehicle treatments. Ewes were injected i.v. with 0.5 mg E2 or vehicle on Day 12 and received twice-daily infusions of 1.5 mg SP or oCSP (containing 25 micrograms ovine trophoblast protein-1 by radioimmunoassay [RIA]) + SP (1.5 mg total protein) into each uterine horn on Days 12, 13, and 14. Blood samples for RIA of plasma progesterone were collected on Days 10-15 (before treatment on each day) and endometrium was collected on Day 15. For each Exp, 100 mg endometrium was incubated, in duplicate, for 2 h with 10 microCi [3H] inositol and treated with 0 or 100 nM oxytocin (OT) for 20 min, then [3H]inositol mono-, bis-, and trisphosphates (IP1, IP2, and IP3, respectively) were quantified.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of post-mating GnRH analogue (buserelin) treatment on PGF2alpha release in ewes and ewe lambs

The objectives of this study were to determine the effects of buserelin or saline treatment on ovarian function (Experiment 1), plasma PGFM concentrations and oxytocin stimulated prostaglandin F(2alpha) (PGF(2alpha)) release (Experiment 2) in ewe lambs and ewes. Welsh Halfbred ewes (n=26) and ewe lambs (n=24) were mated to vasectomised rams at synchronised oestrus and on Day 12 post-mating each animal was injected intramuscularly either normal saline or 4 microg buserelin. In Experiment 1, plasma progesterone and oestradiol concentrations were determined in samples collected by jugular venepuncture 1h before and at 0, 2, 4, 6, 8, 24, 48 and 72 h after treatment (n=7 per treatment group). Progesterone concentrations increased (P<0.05) from 2 to 8h after buserelin treatment and returned to basal levels after 72 h, whereas oestradiol concentrations were maximal at 2h post-treatment and returned to basal levels after 24h (P<0.05). Oestradiol concentrations were lower (P<0.05) in buserelin-treated animals than controls at 72 h post-treatment. Basal and post-treatment progesterone concentrations were greater (P<0.05) in ewes than in ewe lambs but oestradiol levels were similar for both age groups. Ovulation rate, determined by laparoscopy on Day 14, was similar for both age groups (ewes 1.1; ewe lambs 1.0). Buserelin treatment induced accessory corpora lutea in ewes (4/7; 57%) but not in ewe lambs (0/7; 0%). In the Experiment 2, plasma PGFM concentrations were determined in samples collected at 20-min intervals for 6h on Day 14 and at 20-min intervals for 1h before and at 10-min intervals for 1h and then at 20-min intervals for a further 3h period after an intravenous injection of oxytocin (1IU/kg body weight) on Day 15 post-oestrus. In this experiment there were five ewe lambs and six ewes per treatment group. There was no effect of buserelin treatment or age on basal PGFM concentrations on either Day 14 or 15. Although peak PGFM concentrations tended to be lower in buserelin-treated animals, the difference was not significant (P>0.05). However, peak duration following oxytocin challenge on Day 15 post-mating was shorter (P<0.05) in control ewes compared with control ewe lambs. In conclusion, buserelin treatment given on Day 12 post-oestrus enhances luteal function more in ewes than ewe lambs and after a transitory increase, reduces oestradiol concentrations in both ewes and ewe lambs. However, buserelin treatment does not significantly attenuate the luteolytic signal.     

Effect of pregnancy on oxytocin-induced release of prostaglandin F2 alpha in heifers

The effect of pregnancy on the release of prostaglandin F2 alpha (PGF2 alpha) in response to oxytocin (OT) has been examined. Fourteen cyclic heifers received one intravenous injection of 1 IU OT (n = 6) or 100 IU OT (n = 8) 17, 18, or 19 days (Day 17-19) after the onset of estrus (Day 0). Five of these animals also received 100 IU OT at Days 6 and 13 to determine the effect of OT at different times of the cycle. Frequent blood samples were taken for 60 min before and for 90 min after OT injection for the measurement of 15-keto-13,14-dihydro-PGF2 alpha (PGFM) by radioimmunoassay. The experiment was then repeated using the same animals at Day 17-19 of pregnancy (confirmed by the recovery of an embryo the day after OT injection). Following the injection of 1 IU OT, plasma PGFM reached its peak within 30 min with the increase significantly lower (P less than 0.05) in pregnant (1.13 +/- 0.10-fold) than in nonpregnant animals (2.07 +/- 0.27-fold). However, because only 3 of the 6 cyclic animals showed a response to 1 IU OT, the dose was increased to 100 IU in subsequent experiments. The animals that received 100 IU at Days 6 and 13 had no significant increase in PGFM concentrations (1.18 +/- 0.05-fold and 1.01 +/- 0.04-fold, respectively). At Day 17-19 the increase in plasma PGFM reached its peak 5-15 min after 100 IU OT and the increase was significantly greater in nonpregnant (3.23 +/- 0.17-fold) than in pregnant (1.21 +/- 0.02-fold; P = 0.003) heifers. Six of 11 animals injected at Day 17-19 of the cycle showed a decrease in progesterone (P4) the day after OT administration. These data show that the release of PGF2 alpha in response to OT is suppressed in pregnant animals in vivo, suggesting an antiluteolytic role for the embryo in luteostasis.     

Control of birth in rats by RU 486, an antiprogesterone compound

Rats, isolated at mating (Day 1 of pregnancy), were submitted to either 8 h (8L:16D, Exp. I) or 14 h (14L:10D, Exp. II) of light daily with lights on from 12:00 h to 20:00 h and from 06:00 to 20:00 h respectively. In Exp. I, a single dose of RU 486 (10 mg in 0.2 ml ethanol) was given cutaneously at 08:00 h (Group A1), 12:00 h (Group B1), 19:00 h (Group C1) on Day 21 and at 08:00 h (Group D1) and 12:00 h (Group E1) on Day 22. In Exp. II, the same dose of RU 486 was given at 08:00 h (Group A2), 12:00 h (Group B2) and 19:00 h (Group C2) on Day 21. The solvent was given once at each of the preceding times to the control groups (T1 and T2) in both experiments. Groups T1 and T2 gave birth at two periods, the first on Day 22, the second on Day 23; the proportion of births during each of these periods depended on the light regimen (66.3% in 8L:16D; 50% in 14L:10D on Day 22). The distribution of births in Groups D1 and E1 treated on Day 22 were similar to their controls (T1). Rats treated on Day 21 (Groups A1, A2, B1, B2, C1, C2) gave birth over single periods on Day 22 after an interval correlated with the time of RU 486 administration. The earlier the treatment was given, the higher was the number of dead young and the lower the weight of live young 1 day after birth. These effects of prematurity did not impair further survival rates or weight at weaning.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of injected bovine interferon-alpha I1 on estrous cycle length and pregnancy success in sheep

In Exp. 1 twice daily i.m. injections of 2 mg recombinant bovine IFN-alpha I1 (rboIFN-alpha I1) (N = 24) or placebo (N = 25) were administered to ewes from Day 12 to Day 16 during a normal oestrous cycle. Treatment did not increase (P greater than 0.10) oestrous cycle length (20.7 +/- 1.2 versus 18.5 +/- 1.4 days). In Exp. 2, ewes were injected twice daily with 2 mg IFN (N = 34) or placebo (N = 36) from Days 11 to 18 after natural mating. The rboIFN-alpha I1 significantly (P = 0.05) improved pregnancy rate (79% versus 58%) as determined by a failure of ewes to return to oestrus within 50 days. The number of ewes that lambed was greatest in the rboIFN-alpha I1-treatment group (71% versus 50%; P = 0.07), and no teratogenic effects were observed in the young born to IFN-treated ewes. The study was repeated a second year with a more fecund group of ewes (Exp. 3). More (P = 0.08) ewes injected with rboIFN-alpha I1 (58/65) than placebo-treated ewes (48/61) were judged pregnant by ultrasound. Again more ewes lambed (55 versus 45) and more lambs were born (98 versus 80) from the rboIFN-alpha I1-treated group. Combining the data from both studies revealed a significant (P = 0.01) effect of treatment. The amount of antiviral activity in jugular vein blood of ewes injected with rboIFN-alpha I1 (2 mg) was determined over time in Exp. 4. Activity rose to a maximum (approximately 450 IRU/ml) within 1-2 h and declined by over 75% in 24 h. Single injections of 1, 2 and 5 mg in buffer or 2 mg emulsified in sesame oil all gave similar profiles of antiviral activity in jugular blood over a 48-h period. In Exp. 5, antiviral activity was measured in uterine vein, ovarian artery and jugular vein serum of untreated pregnant (N = 7) and non-pregnant (N = 11) ewes at Day 15 after mating. Activity was detected in the uterine vein (58 +/- 19 IRU/ml) of all pregnant ewes. The observations in Exps 1-5 are consistent with a role for conceptus-derived IFN-alpha in maternal recognition of pregnancy and suggest that supplemental IFN-alpha might be useful in improving pregnancy success in sheep.     

Receptors for prostaglandins F2 alpha and E2 in ovine corpora lutea during maternal recognition of pregnancy.

Plasma membrane receptors for prostaglandins (PG) F2 alpha and E2 were quantified in ovine corpora lutea obtained from nonpregnant and pregnant ewes on Days 10, 13, and 15 post-estrus, and from additional ewes on Days 25 and 40 of pregnancy. Regardless of reproductive status or day post-estrus, concentrations of luteal receptors for PGF2 alpha were 7- to 10-fold greater than those for PGE2. In pregnant ewes the concentration of receptors for PGF2 alpha was highest on Day 10 (35.4 +/- 2.8 fmol/mg) and lowest on Day 25 (22.3 +/- 2.5 fmol/mg). A difference in the concentration of luteal receptors for PGF2 alpha between pregnant and nonpregnant ewes was apparent only on Day 15 post-estrus, at which time the concentration of receptors for PGF2 alpha was higher in pregnant ewes than in nonpregnant ewes (27.1 +/- 2.7 vs. 17.7 +/- 2.7 fmol/mg). Concentrations of receptors for PGE2 in pregnant ewes were similar (p > 0.05; 2.8 +/- 0.3 to 3.7 +/- 0.2 fmol/mg) between Days 13 and 40 but were higher (p < 0.05) than in corpora lutea obtained from nonpregnant ewes on Days 10 (5.0 +/- 0.4 vs. 4.1 +/- 0.2 fmol/mg) and 15 (3.7 +/- 0.2 vs. 2.0 +/- 0.4 fmol/mg) post-estrus. Although concentrations of receptors for both PGF2 alpha and PGE2 were lowest in corpora lutea obtained from nonpregnant ewes on Day 15, this was not due to luteal regression since the weights and concentrations of progesterone in corpora lutea on Day 15 were not lower than those for corpora lutea obtained on Days 10 and 13.(ABSTRACT TRUNCATED AT 250 WORDS)     

Independence of progesterone blockade of the luteinizing hormone surge in ewes from opioid activity at naloxone-sensitive receptors.

Fifteen ovariectomized ewes were treated with implants (s.c.) creating circulating luteal progesterone concentrations of 1.6 +/- 0.1 ng ml-1 serum. Ten days later, progesterone implants were removed from five ewes which were then infused with saline for 64 h (0.154 mol NaCl l-1, 20 ml h-1, i.v.). Ewes with progesterone implants remaining were infused with saline (n = 5) or naloxone (0.5 mg kg-1 h-1, n = 5) in saline for 64 h. At 36 h of infusion, all ewes were injected with oestradiol (20 micrograms in 1 ml groundnut oil, i.m.). During the first 36 h of infusion, serum luteinizing hormone (LH) concentrations were similar in ewes infused with saline after progesterone withdrawal and ewes infused with naloxone, but with progesterone implants remaining (1.23 +/- 0.11 and 1.28 +/- 0.23 ng ml-1 serum, respectively, mean +/- SEM, P greater than 0.05). These values exceeded circulating LH concentrations during the first 36 h of saline infusion of ewes with progesterone implants remaining (0.59 +/- 0.09 ng ml-1 serum, P less than 0.05). The data suggested that progesterone suppression of tonic LH secretion, before oestradiol injection, was completely antagonized by naloxone. After oestradiol injection, circulating LH concentrations decreased for about 10 h in ewes of all groups. A surge in circulating LH concentrations peaked 24 h after oestradiol injection in ewes infused with saline after progesterone withdrawal (8.16 +/- 3.18 ng LH ml-1 serum).(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of continuous infusion of oxytocin on length of the oestrous cycle and luteolysis in cattle

In Exp. I oxytocin (60 micrograms/100 kg/day) was infused into the jugular vein of 3 heifers on Days 14-22, 15-18 and 16-19 of the oestrous cycle respectively. In Exp. II 5 heifers were infused with 12 micrograms oxytocin/100 kg/day from Day 15 of the oestrous cycle until clear signs of oestrus. Blood samples were taken from the contralateral jugular vein at 2-h intervals from the start of the infusion. The oestrous cycle before and after treatment served as the controls for each animal. Blood samples were taken less frequently during the control cycles. In Exp. III 3 heifers were infused with 12 micrograms oxytocin/100 kg/day for 50 h before expected oestrus and slaughtered 30-40 min after the end of infusion for determination of oxytocin receptor amounts in the endometrium. Three other heifers slaughtered at the same days of the cycle served as controls. Peripheral concentrations of oxytocin during infusion ranged between 155 and 641 pg/ml in Exp. I and 18 and 25 pg/ml in Exp. II. In 4 our of 8 heifers of Exps I and II, one high pulse of 15-keto-13,14-dihydro-prostaglandin F-2 alpha (PGFM) appeared soon after the start of oxytocin infusion followed by some irregular pulses. The first PGFM pulse was accompanied by a transient (10-14 h) decrease of blood progesterone concentration. High regular pulses of PGFM in all heifers examined were measured between Days 17 and 19 during spontaneous luteolysis. No change in length of the oestrous cycle or secretion patterns of progesterone, PGFM and LH was observed.(ABSTRACT TRUNCATED AT 250 WORDS)     

Induction of ovulation in the acyclic postpartum ewe following continuous, low-dose subcutaneous infusion of GnRH

Pituitary and ovarian responses to subcutaneous infusion of GnRH were investigated in acyclic, lactating Mule ewes during the breeding season. Thirty postpartum ewes were split into 3 equal groups; Group G received GnRH (250 ng/h) for 96 h; Group P + G was primed with progestagen for 10 d then received GnRH (250 ng/h) for 96 h; and Group P received progestagen priming and saline vehicle only. The infusions were delivered via osmotic minipumps inserted 26.6 +/- 0.45 d post partum (Day 0 of the study). Blood samples were collected for LH analysis every 15 min from 12 h before until 8 h after minipump insertion, then every 2 h for a further 112 h. Daily blood samples were collected for progesterone analysis on Days 1 to 10 following minipump insertion, then every third day for a further 25 d. In addition, the reproductive tract was examined by laparoscopy on Day -5 and Day +7 and estrous behavior was monitored between Day -4 and Day +7. Progestagen priming suppressed (P < 0.05) plasma LH levels (0.27 +/- 0.03 vs 0.46 +/- 0.06 ng/ml) during the preinfusion period, but the GnRH-induced LH release was similar for Group G and Group P + G. The LH surge began significantly (P < 0.05) earlier (32.0 +/- 3.0 vs 56.3 +/- 4.1 h) and was of greater magnitude (32.15 +/- 3.56 vs 18.84 +/- 4.13 ng/ml) in the unprimed than the primed ewes. None of the ewes infused with saline produced a preovulatory LH surge. The GnRH infusion induced ovulation in 10/10 unprimed and 7/9 progestagen-primed ewes, with no significant difference in ovulation rate (1.78 +/- 0.15 and 1.33 +/- 0.21, respectively). Ovulation was followed by normal luteal function in 4/10 Group-G ewes, while the remaining 6 ewes had short luteal phases. In contrast, each of the 7 Group-P + G ewes that ovulated secreted progesterone for at least 10 d, although elevated plasma progesterone levels were maintained in 3/7 unmated ewes for >35 d. Throughout the study only 2 ewes (both from Group P + G) displayed estrus. These data demonstrate that although a low dose, continuous infusion of GnRH can increase tonic LH concentrations sufficient to promote a preovulatory LH surge and induce ovulation, behavioral estrus and normal luteal function do not consistently follow ovulation in the progestagen-primed, postpartum ewe.