G.E. Lamming

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

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

The role of sub-optimal preovulatory oestradiol secretion in the aetiology of premature luteolysis during the short oestrous cycle in the cow.

Premature regression of the corpus luteum, following the first post partum ovulation, is often preceded by sub-optimal preovulatory oestradiol secretion and accompanied by elevated levels of oxytocin receptors early in the luteal phase. We have investigated the role of preovulatory oestradiol in the control of subsequent oxytocin receptor concentration and activity by treating ovariectomised cows, over a simulated 48 h follicular phase, with high (600 microg per day) medium (300 microg per day) or low (150 microg per day) levels of oestradiol. These doses of oestradiol generated mean+/-S.E.M. plasma oestradiol concentrations of 12.1+/-1.0, 4.9+/-0.5 and 2.9+/-0.4 pg ml(-1), respectively. In Study 1 (n=4 per group), we found that by day 4 following oestrus there was a significant (P< 0.05) effect of the level of oestradiol on the inhibition of oxytocin binding activity measured in endometrial biopsy samples. This had fallen to mean+/-S.E.M. concentrations of 25+/-2 fmol per mg protein in the high group, 47+/-8 fmol per mg protein in the medium group and 65+/-12 fmol per mg protein in the low group. In Study 2, cows (n=3 per group) were treated with the same three levels of oestradiol followed by treatment with increasing levels of progesterone from days 3 to 6 following oestrus, generating mean+/-S.E.M. plasma concentrations of 2.17+/-0.18 ng ml(-1) by day 6. On day 6, there was a significant (P< 0.01) effect of the level of oestradiol on PGF(2alpha) release in response to oxytocin challenge. High, medium and low oestradiol groups exhibiting mean+/-S.E.M., increase plasma PGF(2alpha) metabolite concentrations of 10.0+/-2.2, 21.3+/-4.3 and 41.3+/-1.2 pg ml(-1), respectively, during the hour after oxytocin administration. From these results, we postulate that at the first post partum ovulation a low level of preovulatory oestradiol can result in the early generation of a luteolytic mechanism during the subsequent luteal phase due to impaired inhibition of oxytocin receptors allowing increased PGF(2alpha) release.

Effects of a GnRH agonist (Buserelin) on LH secretion in post-partum beef cows

Plasma LH concentrations were monitored in post-partum beef cows treated with the GnRH agonist Buserelin, administered via biodegradable implants. Animals were given either untreated Buserelin implants (Group 1, n = 6) or implants that had been preincubated for 7 (Group 2, n = 6) or 14 (Group 3,n = 6) days. Control cows were sham-implanted (Group 4, n = 3). The implants were inserted subcutaneously at a mean time of 25 ± 2.8 days after caiving, and LH concentrations were monitored over 8 h periods of blood sampling (samples taken at 10 min intervals) undertaken on Days — 2, 1, 2, 3, 7, 14, 21 and 28 (Day 1 was the day of implantation). In all animals treated with Buserelin, plasma LH concentrations increased within 1 h of implantation to reach a maximum concentration (mean 35.4 ± 2.86 ng ml−1) within 4 h. In all three Buserelin-treated groups, mean LH concentrations were higher on Day 1 (P < 0.001) and on Days 2 and 3 (P < 0.05) than on Day — 2, before the start of treatment. On subsequent sampling days (Days 7, 14, 21 and 28), mean LH concentrations did not differ significantly from pretreatment values, but on Days 14, 21 and 28, mean LH concentrations were lower (P < 0.05) in Buserelin-treated than in control cows. There were no significant differences in either the magnitude or the duration of the LH response between Buserelin treatment groups. Before treatment, LH episodes occurred with a mean frequency of 1.9 ± 0.4 episodes per 8 h, and in control cows occurred at a frequency of 2.3 ± 0.2 episodes per 8 h throughout the 28-day sampling period. In contrast, LH episodes were not recorded in any of the Buserelin-treated cows on the first 3 days after implantation, and were recorded in only 1 of 18, 2 of 18, 1 of 18 and 4 of 18 animals on Days 7, 14, 21 and 28, respectively. Following luteal regression, plasma progesterone concentrations remained low for the remainder of the sampling period (mean 18.2 ± 1.2 days) in 17 of 18 Buserelin treated cows. These results demonstrate that Buserelin treatment induces a short period of increased LH secretion, after which there is a prolonged period of suppression of LH release which blocks ovulation.

The development of subcutaneous-delivery preparations of GnRH for the induction of ovulation in acyclic sheep and cattle

Abstract A series of oil-based preparations and biodegradable implants were tested for their efficacy to release low levels of GnRH for a prolonged period when administered subcutaneously, and thus to stimulate tonic LH secretion, promote preovulatory follicle growth and induce ovulation in acyclic sheep and cattle. A total of 15 oil-based formulations incorporating nine different carrier solutions were assessed. Wide between-animal variation in both the magnitude and the duration of the LH response to these preparations overshadowed differences between the doses of GnRH given, or the formulations used. Only six out of 55 seasonally anoestrous ewes, and none of the 57 heifers treated with these preparations produced an endogenous preovulatory LH surge. The first generation of biodegradable GnRH implants tested (ChB2) increased plasma LH concentrations from mean pretreatment values of 0.36 ± 0.08 ng/ml (mean ± s.e.m.) to a mean maximum concentration of 6.11 ± 0.89 ng/ml, and a preovulatory-surge and ovulation occurred in 61% of treated seasonally anoestrous ewes. The second GnRH implant-type (ChB3) resulted in mean maximum LH concentrations of 13.12 ± 1.72 ng/ml, and preovulatory surges and ovulation in 89% of treated ewes. The administration of two implants per animal induced ovulation in 88% and 94% of treated anoestrous ewes, for implant types ChB2 and ChB3, respectively. These results demonstrate the potential for using biodegradable implants, but not oil-based preparations, for inducing ovulation with GnRH in acyclic animals.

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.

Inter-breed variation in the postpartum ewe response to continuous low dose infusion of GnRH

During the nonbreeding season the pituitary and ovarian responses to a subcutaneous GnRH infusion were investigated in acyclic, lactating Mule ewes which exhibit a deep seasonal anestrus and in Finn x Dorset ewes in which seasonal anestrus is ill-defined. Each breed received 10 d of progestagen priming before being subdivided into 3 groups. In Group L + G, 5 lactating ewes received GnRH (250 ng/h sc) for 96 h; in Group D + G, 5 dry ewes received GnRH (250 ng/h sc) for 96 h; in Group L, 5 lactating ewes received saline vehicle for 96 h. The infusions began when lactating and dry ewes were approximately 28 d and 120 d post partum, respectively. Blood samples were collected for LH, progesterone and estradiol analysis. Estrous behavior was monitored between Day -4 and Day +7. On Day +7 the reproductive tract was also examined. In the Mule ewes the mean plasma LH concentration increased (P < 0.05) following minipump insertion in each treatment group, although mean LH levels were greater (P < 0.05) in Group D + G, than in either Group L + G or Group L. Following the GnRH infusion, mean plasma estradiol levels increased (P < 0.05) in Group D + G but not in Group L + G. A preovulatory LH surge and subsequent ovulation occurred in 5 5 , 2 5 and 0 5 ewes from Group D + G, L + G and L, respectively, and estrus was recorded in 5 5 , 1 5 and 0 5 of these ewes, respectively. The LH surges began earlier (P < 0.05) (43.2 +/- 6.8 h vs 77.0 +/- 1.0 h) and the ovulation rate was greater (2.2 +/- 0.37 vs 1.00 +/- 0.00) in Group D + G than Group L + G. In the Finn x Dorset ewes mean LH concentrations increased (P < 0.05), to a similar level following minipump insertion in Groups D + G and L + G, but not Group L. The elevated LH levels were accompanied by increased (P < 0.05) plasma estradiol levels in Group D + G, but not in Group L + G. The GnRH infusion culminated in an LH surge and estrous behavior in 5 5 , 1 5 and 0 5 ewes from Groups D + G, L + D and L, respectively. The interval to the LH surge was similar between Group D + G (48.4 +/- 6.6 h) and Group L + G (46.0 h). Ovulation was evident in those ewes which exhibited an LH surge plus one additional ewe from Group L + G. The mean ovulation rate was greater in Group D + G (4.00 +/- 1.05) than in Group L + G (1.5 +/- 0.50). These data show that continuous GnRH infusion can consistently induce out of season breeding in the nonlactating Mule and Finn x Dorset ewe but can not break combined seasonal and lactational anestrous in these breeds. Further, between-breed differences are evident in the site along the hypothalamic-pituitary-ovarian axis at which reproduction is compromised in ewes at the same chronological stage post partum.

Ovarian control of gonadotrophin secretion in the prepubertal heifer

Abstract In order to investigate the development of ovarian feedback control of gonadotrophin secretion in the immature animal, three groups of eight Hereford X Friesian heifers were bilaterally ovariectomized at either 2, 6 or 12 weeks of age. Blood samples were collected for gonadotrophin analysis (15-min intervals for 6 h) on days −1, 4, 7, 14, 28, 55 and 110 relative to ovariectomy. The ovaries from these animals showed that follicular development was established soon after birth and the follicles were producing oestradiol. In all animals, ovariectomy was followed by a rise in plasma gonadotrophin concentrations although the response was delayed in heifers ovariectomized at 2 weeks compared to those ovariectomized at 6 or 12 weeks of age. Mean LH concentrations were significantly elevated 14, 7 or 4 days after ovariectomy and mean FSH concentrations were significantly higher than pre-treatment values 28, 7, or 4 days post-ovariectomy in heifers ovariectomized at 2, 6 or 12 weeks of age, respectively. The results suggest that an inhibitory mechanism, of ovarian origin, that controls gonadotrophin secretion develops between 2 and 12 weeks of age in the heifer.

The effect of exogenous progesterone on plasma LH and milk progesterone concentrations in multiple-suckling post-partum cows

Abstract Fourteen Friesian cows each suckling four calves were treated for a 7-day period (a) between days 20–40 post partum with progesterone-releasing intravaginal devices (PRID) containing 2% progesterone (Group 1; n = 5), (b) between days 51–264 post partum with PRIDS containing 2% progesterone (Group 2; n = 6) and (c) between days 29–214 days post partum with PRIDS containing 0% progesterone (Group 3; n = 3). Mean plasma LH concentrations decreased during PRID treatment in Group 2 cows only and pre-ovulatory LH surges were observed in 5 6 of these cows between 38 and 84 h after coil removal. All Group 2 cows underwent at least one ovarian cycle following PRID removal. No pre-ovulatory LH surges were observed in either Group 1 or Group 3 cows and only one cow (Group 3) underwent an ovarian cycle after treatment. It is suggested that there is an increase in pituitary responsiveness to the feedback effects of progesterone during the post-partum period.

Ovulatory responses to continuous administration of GnRH in nine-month-old prepubertal beef heifers.

Abstract Continuous infusion of gonadotrophin-releasing hormone (GnRH) will consistently induce ovulation in several species, but has had limited success in post-partum acyclic cattle. We suggest that this treatment is only effective when anovulation is due solely to a lack of gonadotrophic stimulation of the ovary, rather than to a hypothalamic-pituitary block. Therefore, we have attempted to induce ovulation in 9-month-old prepubertal heifers, an experimental model in which lack of ovarian stimulation is likely to be the only limitation to oestrous cycles. GnRH was administered either at a controlled rate (5 μg/h) via osmotic minipumps (inserted s.c.), or at a less predictable rate by s.c. implantation of a biodegradable implant (40 μg GnRH/implant) previously shown to be effective in sheep. The mean magnitude of the maximum luteinizing hormone (LH) concentration in response to GnRH treatment was significantly higher in heifers treated with 24 GnRH implants than in those given 8 implants or osmotic minipumps (18.02 ± 1.23 ng/ml, 11.55 ± 2.37 ng/ml and 8.6 ± 2.29 ng/ml, respectively). In addition, plasma LH concentrations reached preovulatory surge magnitude within 4 h of implantation in 5 5 , 1 6 and 0 5 animals in these treatments groups. Conversely, an endogenous preovulatory LH surge was recorded in 0 5 , 3 6 and 5 5 heifers treated with 24 implants, 8 implants or osmotic minipumps. The consistent ovulatory response recorded in those heifers treated with osmotic minipumps demonstrates the importance of administering GnRH at a constant and controlled rate.

Pituitary and ovarian responses to prolonged administration of low doses of GnRH in prepubertal heifers

Abstract The responses of prepubertal heifers to continuous infusion of GnRH were examined in 4 experiments. Eighteen 4 month old beef heifers received 0.25, 0.5 or 2.5 μg GnRH/h for 48 h by continuous intravenous infusion. No significant changes in plasma LH concentrations were observed during infusion of the two lower GnRH doses. However, in all six heifers receiving 2.5 μg GnRH/h, jugular plasma LH concentrations were significantly increased within 2 h of the start of infusion. Preovulatory-type gonadotrophin surges occurred in three of the six animals receiving 2.5 μg/h GnRH during the period of infusion and elevated plasma progesterone concentrations were detected in these animals 4 and 6 days after the end of treatment. Heifers that exhibited preovulatory gonadotrophin surges had higher plasma oestradiol-17β concentrations before treatment than those that did not. In a second experiment, five heifers received 1.25 μg/h GnRH for 120 h. Mean LH and oestradiol-17β concentrations increased in all animals over the first 60 h and then returned to baseline. Two heifers exhibited preovulatory-type LH surges and some luteal function. In Experiment 3, osmotic infusion pumps implanted s.c. were used to deliver 5.0, 7.5 or 10.0 μG GnRH for 8 h and the LH responses compared with the responses to i.v. infusion of 2.5 μg GnRH for 8 h. The latin square design used four heifers and 4 treatment days. Significant effects of both GnRH-dose and day of GnRH treatment were detected, with subcutaneous infusion of 10.0 μg GnRH/h producing a significantly greater LH response than i.v. infusion of 2.5 μg GnRH/h, and the mean LH response to all treatments being greater during the first treatment period than on subsequent treatment days. In Experiment 4, 12 heifers received progesterone pretreatment for 14 days followed by 5.0 μg GnRH/h or saline, infusion s.c. for 120 h. A third group of five heifers received the GnRH infusion only. Mean LH concentrations were significantly elevated in response to GnRH over the first 8 h of the infusion period in progesterone pretreated heifers and over the first 16 h of infusion in animals receiving GnRH alone. Preovulatory LH surges were detected in four GnRH-treated heifers, two animals that were progesterone pretreated and two that received GnRH alone. Laparoscopy revealed corpora lutea in three of these four heifers, and plasma progesterone concentrations were elevated in two of these. It is concluded that GnRH infusion does not consistently induce ovulation in prepubertal heifers.