Effects of oestradiol on LH, FSH and prolactin in ovariectomized ewes

This study was conducted to test the hypothesis that the rate (dose/time) at which oestradiol-17 beta (oestradiol) is presented to the hypothalamo-pituitary axis influences secretion of LH, FSH and prolactin. A computer-controlled infusion system was used to produce linearly increasing serum concentrations of oestradiol in ovariectomized ewes over a period of 60 h. Serum samples were collected from ewes every 2 h from 8 h before to 92 h after start of infusion, and assayed for oestradiol, LH, FSH and prolactin. Rates of oestradiol increase were categorized into high (0.61-1.78 pg/h), medium (0.13-0.60 pg/h) and low (0.01-0.12 pg/h). Ewes receiving high rates of oestradiol (N = 11) responded with a surge of LH 12.7 +/- 2.0 h after oestradiol began to increase, whereas ewes receiving medium (N = 15) and low (N = 11) rates of oestradiol responded with a surge of LH at 19.4 +/- 1.7 and 30.9 +/- 2.0 h, respectively. None of the surges of LH was accompanied by a surge of FSH. Serum concentrations of FSH decreased and prolactin increased in ewes receiving high and medium rates of oestradiol, when compared to saline-infused ewes (N = 8; P less than 0.05). We conclude that rate of increase in serum concentrations of oestradiol controls the time of the surge of LH and secretion of prolactin and FSH in ovariectomized ewes. We also suggest that the mechanism by which oestradiol induces a surge of LH may be different from the mechanism by which oestradiol induces a surge of FSH.     

Use of a GnRH agonist to prevent the endogenous LH surge and injection of exogenous LH to induce ovulation in heifers superstimulated with FSH: a new model for superovulation

A new protocol for superovulating cattle which allows for control of the timing of ovulation after superstimulation with FSH was developed. The preovulatory LH surge was blocked with the GnRH agonist deslorelin, and ovulation was induced by injection of LH. In Experiment 1, heifers (3-yr-old) were assigned to a control group (Group 1A, n = 4) or a group with deslorelin implants (Group 1B, n = 5). On Day -7, heifers in Group 1A received a progestagen CIDR-B((R))device, while heifers in Group 1B received a CIDR-B((R))device + deslorelin implants. Both groups were superstimulated with twice daily injections of FSH (Folltropin((R))-V): Day 0, 40 mg (80 mg total dose on Day 0); Day 1, 30 mg; Day 2, 20 mg; Day 3, 10 mg. On Day 2, heifers were given PGF (a.m.) and CIDR-B((R)) devices were removed (p.m.). Three heifers in Group 1A had a LH surge and ovulated, whereas neither of these events occurred in Group 1B (with deslorelin implants) heifers. In Experiment 2, heifers (3-yr-old) were assigned to 1 of 4 equal groups (n = 6). On Day -7, heifers in Group 2A received a norgestomet implant, while heifers in Groups 2B, 2C and 2D received norgestomet + deslorelin implants. Heifers were superstimulated with FSH starting on Day 0 as in Experiment 1. On Day 2, heifers were given PGF (a.m.) and norgestomet implants were removed (p.m.). Heifers in Groups 2B to 2D were given 25 mg LH (Lutropin((R))): Group 2B, Day 4 (a.m.); Group 2C, Day 4 (p.m.); Group 2D, Day 5 (a.m.). Heifers in Group 2A were inseminated at estrus and 12 and 24 h later, while heifers in Groups 2B to 2D were inseminated at the time of respective LH injection and 12 and 24 h later. Injection of LH induced ovulation in heifers in Groups 2B to 2D. Heifers in Group 2C had similar total ova and embryos (15.2 +/- 1.4) as heifers in Group 2A (11.0 +/- 2.8) but greater (P < 0.05) numbers than heifers in Group 2B (7.0 +/- 2.3) and Group 2D (6.3 +/- 2.0). The number of transferable embryos was similar for heifers in Group 2A (5.8 +/- 1.8) and Group 2C (7.3 +/- 2.1) but lower (P < 0.05) for heifers in Group 2B (1.2 +/- 0.8) and Group 2D (1.3 +/- 1.0). The new GnRH agonist-LH protocol does not require observation of estrus, and induces ovulation in superstimulated heifers that would not have an endogenous LH surge.     

Compensation by pulsatile GnRH infusions for incompetence for oestradiol-induced LH surges in long-term ovariectomized gilts and castrated male pigs.

The aim of this study was to investigate incompetence for oestradiol-induced LH surges in long-term ovariectomized gilts and male pigs. Gilts (250 days old; n = 36), which had been ovariectomized 30 (OVX 30) or 100 days (OVX 100) before the start of treatment, were challenged i.m. with oestradiol benzoate and were either given no further treatment, fed methallibure to inhibit endogenous GnRH release or fed methallibure and given i.v. pulses of 100 or 200 ng GnRH agonist at 1 h intervals during the LH surge (48-96 h after oestradiol benzoate). The same treatments were applied to long-term orchidectomized male pigs (ORC, n = 23). In addition, one ORC group was not injected with oestradiol benzoate but was fed methallibure and given pulses of 200 ng GnRH agonist. Oestradiol benzoate alone induced an LH surge in the OVX 30 group only (5/6 gilts), methallibure suppressed (P < 0.05) oestradiol benzoate-induced LH secretion, while pulses of 100 ng GnRH agonist in animals fed methallibure produced LH surges in four of six OVX 30 and four of six OVX 100 gilts. The induced LH surges were similar to those produced by oestradiol benzoate alone in OVX 30 gilts. Pulses of 200 ng GnRH agonist produced LH surges in OVX 30 (6/6) and OVX 100 (6/6) gilts and increased the magnitude of the induced LH surge in OVX 100 gilts (P < 0.05 compared with 100 ng GnRH agonist or OVX 30 control). Pulses of 200 ng GnRH agonist also induced LH surge release in ORC male pigs (5/6), but were unable to increase LH concentrations in a surge-like manner in ORC animals that had not been given oestradiol benzoate, indicating that oestradiol increases pituitary responsiveness to GnRH. These results support the hypothesis that oestradiol must inhibit secretion of LH before an LH surge can occur. It is concluded that incompetence for oestradiol-induced LH surges in long-term ovarian secretion-deprived gilts and in male pigs is due to the failure of oestradiol to promote a sufficient increase in the release of GnRH.     

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.     

Effect of photoperiod on LH, FSH, prolactin and melatonin patterns in ovariectomized prepubertal heifers

Angus and Angus crossbred prepubertal heifers were ovariectomized and randomly assigned to either increasing light simulating the photoperiod of the vernal equinox to the summer solstice (I) or decreasing light simulating the photoperiod of the autumnal equinox to the winter solstice (D) for 43 degrees N latitude. Three blood samples were taken each week for 14 weeks, the first at 11:00 h and two others 2 days later, 1 h before lights on (dark), 1 h before lights off (light). At the end of 14 weeks 4 heifers from each treatment group were cannulated and samples were taken for 12 h at 15-min intervals, 6 h in the light and 6 h in the dark. All sera were assayed for LH, FSH and prolactin. In addition, the samples taken at 15-min intervals were assayed for melatonin. In samples taken weekly at 11:00 h circulating concentrations of LH and prolactin were higher among animals in Group I, while FSH concentrations were not different between Groups D and I. In samples collected weekly in the light or the dark, LH and prolactin concentrations were higher in Group I animals. However, prolactin concentrations were higher and LH concentrations tended to be higher in samples taken in the dark. FSH concentrations were not different between either D or I or dark and light. In samples taken at 15-min intervals the prolactin baseline was higher and pulse amplitude tended to be higher for Group I animals. Neither LH nor FSH pulse characteristics differed between I and D; however, LH baseline and LH pulse amplitude were higher in the dark. Melatonin pulse amplitude was higher among animals in Group D and higher in serum collected in the dark. These results suggest that photoperiod alters circulating concentrations of LH and prolactin and alters pulsatile release of LH, prolactin and melatonin in the prepubertal heifer.     

Surges of FSH during the follicular and early luteal phases of the estrous cycle in heifers

Surges of FSH were characterized in each of 12 Holstein heifers using a computerized cycle detector program, and as mean changes averaged over all heifers. Blood samples were collected 6 times a day at 4-h intervals beginning at late diestrus. Concentrations of FSH were adjusted relative to the preovulatory LH peak (Hour 0) and profiled beginning 48 h before and ending 120 h after the LH peak. Peak concentrations of FSH and LH occurred synchronously in 11 of 12 (92%) heifers, and only a 4-h interval separated peak concentrations in the remaining heifer. The FSH surge that was synchronous with the LH surge was designated FSH Surge 1 and was used as a reference to designate other FSH surges. Surge -1 of FSH was detected in 58% of the heifers at mean Hour -21.2, and Surges 2, 3 and 4 were detected in 92%, 92% and 75% of the heifers, respectively, at mean Hours 25.1, 57.8 and 78.7. Mean peak levels and duration of FSH Surges-1, 2, 3 and 4 were significantly lower than for FSH Surge 1. Mean concentrations of FSH significantly increased and decreased before and after the LH peak, resulting from the synchrony between FSH Surge 1 and the LH surge in individual heifers. Additionally, there was a tendency (P < 0.08) for a second and third increase in mean FSH concentrations at Hours 24 and 60, which was attributed to FSH Surges 2 and 3 that occurred in individuals. Peak FSH concentrations of Surge 2 occurred (mean, Hour 25.1) within 8 h of maximal mean concentrations at Hour 24 in 91% of the heifers. Correspondingly, peak FSH concentrations of Surge 3 occurred (mean, Hour 57.8) within 8 h of maximal mean concentrations at Hour 60 in 64% of the heifers. Surges -1 and 4 of FSH occurred less frequently and at various times within and among heifers compared with Surges 1 to 3; therefore, they were not detected as mean increases in FSH concentrations but were masked as a result of concentrations being averaged over all heifers. In summary, FSH surges were detected in individual heifers before and after the combined FSH/LH surge. The interpeak intervals for FSH Surges 1 to 2 (25 h), 2 to 3 (33 h) and 3 to 4 (21 h) suggests a rhythmic nature to the surges.     

Plasma LH and FSH responses and ovarian activity in prepubertal heifers treated with repeated injections of low doses of GnRH for 72 h

Twelve 5-month-old Hereford X Friesian heifers were injected i.v. with 2.0 micrograms GnRH at 2-h intervals for 72 h. Blood samples were collected at 15-min intervals from 24 h before the start until 8 h after the end of the GnRH treatment period. Over the 24-h pretreatment period, mean LH concentrations ranged from 0.4 to 2.2 ng/ml and FSH concentrations from 14.1 to 157.4 ng/ml; LH episodes (2-6 episodes/24 h) were evident in all animals. Each injection of GnRH resulted in a distinct episode-like response in LH, but not FSH. Mean LH, but not FSH, concentrations were significantly increased by GnRH treatment. The GnRH-induced LH episodes were of greater magnitude than naturally-occurring episodes (mean maximum concentration 6.7 +/- 0.5 and 4.9 +/- 0.6 ng/ml respectively). Preovulatory LH surges occurred between 17.0 and 58.8 h after the start of treatment in 9/12 heifers, with a coincident FSH surge in 8 of these animals. This was not followed by normal luteal function. There were no apparent correlations between pretreatment hormone concentrations, and either the pituitary response to GnRH or the occurrence of preovulatory gonadotrophin release.     

Estradiol induces and progesterone inhibits the preovulatory surges of luteinizing hormone and follicle-stimulating hormone in heifers

Objectives were to determine: 1) whether estradiol, given via implants in amounts to stimulate a proestrus increase, induces preovulatory-like luteinizing hormone (LH) and follicle-stimulating hormone (FSH) surges; and 2) whether progesterone, given via infusion in amounts to simulate concentrations found in blood during the luteal phase of the estrous cycle, inhibits gonadotropin surges. All heifers were in the luteal phase of an estrous cycle when ovariectomized. Replacement therapy with estradiol and progesterone was started immediately after ovariectomy to mimic luteal phase concentrations of these steroids. Average estradiol (pg/ml) and progesterone (ng/ml) resulting from this replacement were 2.5 and 6.2 respectively; these values were similar (P greater than 0.05) to those on the day before ovariectomy (2.3 and 7.2, respectively). Nevertheless, basal concentrations of LH and FSH increased from 0.7 and 43 ng/ml before ovariectomy to 2.6 and 96 ng/ml, respectively, 24 h after ovariectomy. This may indicate that other ovarian factors are required to maintain low baselines of LH and FSH. Beginning 24 h after ovariectomy, replacement of steroids were adjusted as follows: 1) progesterone infusion was terminated and 2 additional estradiol implants were given every 12 h for 36 h (n = 5); 2) progesterone infusion was maintained and 2 additional estradiol implants were given every 12 h for 36 h (n = 3); or 3) progesterone infusion was terminated and 2 additional empty implants were given every 12 h for 36 h (n = 6). When estradiol implants were given every 12 h for 36 h, estradiol levels increased in plasma to 5 to 7 pg/ml, which resembles the increase in estradiol that occurs at proestrus. After ending progesterone infusion, levels of progesterone in plasma decreased to less than 1 ng/ml by 8 h. Preovulatory-like LH and FSH surges were induced only when progesterone infusion was stopped and additional estradiol implants were given. These surges were synchronous, occurring 61.8 +/- 0.4 h (mean +/- SE) after ending infusion of progesterone. We conclude that estradiol, at concentrations which simulate those found during proestrus, induces preovulatory-like LH and FSH surges in heifers and that progesterone, at concentrations found during the luteal phase of the estrous cycle, inhibits estradiol-induced gonadotropin surges. Furthermore, ovarian factors other than estradiol and progesterone may be required to maintain basal concentrations of LH and FSH in heifers.     

Oestradiol and the preovulatory surges of luteinising hormone and follicle stimulating hormone in ewes during the breeding season and transition into anoestrus

This study was designed to see if giving exogenous oestradiol, during the follicular phase of the oestrous cycle of intact ewes, during the breeding season or transition into anoestrus, would alter the occurrence, timing or magnitude of the preovulatory surge of secretion of luteinising hormone (LH) or follicle stimulating hormone (FSH). During the breeding season and the time of transition, separate groups of ewes were infused (intravenously) with either saline (30 ml h⁻¹; n = 6) or oestradiol in saline (n = 6) for 30 h. Infusion started 12 h after removal of progestin-containing intravaginal sponges that had been in place for 12 days. The initial dose of oestradiol was 0.02 μg h⁻¹; this was doubled every 4 h for 20 h, followed by every 5 h up to 30 h, to reach a maximum of 1.5 μg h⁻¹. Following progestin removal during the breeding season, peak serum concentrations of oestradiol in control ewes were 10.31 ± 1.04 pg ml⁻¹, at 49.60 ± 3.40 h after progestin removal. There was no obvious peak during transition, but at a time after progestin removal equivalent to the time of the oestradiol peak in ewes at mid breeding season, oestradiol concentrations were 6.70 ± 1.14 pg ml⁻¹ in ewes in transition (P < 0.05). In oestradiol treated ewes, peak serum oestradiol concentrations (24.8 ± 2.1 pg ml⁻¹) and time to peak (41.00 ± 0.05 h) did not differ between seasons (P > 0.05). During the breeding season, all six control ewes and four of six ewes given oestradiol showed oestrus with LH and FSH surges. The two ewes not showing oestrus did not respond to oestrus synchronisation and had persistently high serum concentrations of progesterone. During transition, three of six control ewes showed oestrus but only two had LH and FSH surges; all oestradiol treated ewes showed oestrus and gonadotrophin surges (P < 0.05). The timing and magnitude of LH and FSH surges did not vary with treatment or season. In blood samples collected every 12 min for 6 h, from 12 h after the start of oestradiol infusion, mean serum concentrations of LH and LH pulse frequency were lower in control ewes during transition than during mid breeding season (P < 0.05). Oestradiol treatment resulted in lower mean serum concentrations of LH in season and lower LH pulse frequency in transition (P < 0.05). We concluded that enhancing the height of the preovulatory peak in serum concentrations of oestradiol during the breeding season did not alter the timing or the magnitude of the preovulatory surge of LH and FSH secretion and that at transition into anoestrus, oestradiol can induce oestrus and the surge release of LH and FSH as effectively as during the breeding season.     

Effect of the introduction of rams during the anoestrous season on the pulsatile secretion of LH in ovariectomized ewes

Introduction of rams to ovariectomized ewes treated with oestradiol implants (N = 10) increased the frequency of LH pulses from 4 X 8 to 10 X 6 pulses per 12 h. This effect was reflected by increases in mean levels of LH and the basal levels upon which the pulses were superimposed. In ewes that had not been treated with oestradiol (N = 5), there was no significant increase in pulse frequency but mean and basal levels of LH increased slightly after the introduction of rams. In a second experiment, similar effects of the introduction of rams were seen in ovariectomized ewes treated with oestradiol or oestradiol + androstenedione (N = 16), but no significant effects of the rams were observed in untreated ewes (N = 8) or ewes treated only with androstenedione (N = 7). No preovulatory surges of LH were observed in the 30-h period after the introduction of rams. It was concluded that the ram stimulus probably evokes the increase in pulse frequency by inhibiting the negative feedback action of oestradiol, and that the surge normally observed in entire ewes is dependent on the ovarian response to these pulses. However, the observation of responses in some ewes not treated with oestradiol also raises the possibility that the ram stimulus can act directly on the hypothalamic neurones that control the secretion of LH, and that this effect is enhanced in the presence of oestrogen.     

The secretion of bioactive and immunoreactive follicle-stimulating hormone (FSH) and luteinizing hormone (LH) throughout the menstrual cycle of the rhesus monkey (Macaca mulatta)

The present study provides the first evaluation of related changes in serum levels of bioactive FSH (Bio FSH) and immunoreactive FSH (iFSH), and concurrent dynamics of LH and FSH bioactivity throughout the menstrual cycle of the rhesus monkey. Mean concentrations of Bio FSH were elevated on days 0 and 1 (n = 7; P < 0.05; day 0 = preovulatory LH surge). Data from individual animals revealed that an average (± SEM) of 1.43 ± 0.29 and 2.71 ± 0.61 discrete surges of Bio FSH occurred in each monkey's follicular and luteal phase, respectively. Analysis of the collective data indicated that periods of increased Bio FSH secretory activity spanned days −1 to 1 and 8 to 10 (P < 0.025). Increases in serum Bio FSH and iFSH concentrations were not precisely correlated on a daily basis (38.9%), although 72.2% of the peaks of Bio FSH and iFSH surges occurred within a day of one another. Similarly, only 36.1% of the Bio FSH surges were accompanied by elevations in bioactive LH (Bio LH). A significant rise in Bio LH, but not Bio FSH, occurred on day −1 (P < 0.01). Concentrations of Bio LH, but not Bio FSH, were elevated in the early luteal phase (P < 0.01). The bioactivity/immunoactivity ratios (Bio/I) of LH and FSH were maximal on the day of the preovulatory surge (P < 0.01). On day −1, LH Bio/I significantly increased (P < 0.05), but no change in FSH Bio/I was detected. The Bio/I of LH, but not FSH, remained elevated in the early luteal phase. In summary: the relative increase in Bio FSH exceeds iFSH during the preovulatory surge. Surges of Bio FSH occur during the follicular and luteal phases which potentially could support follicle selection/maturation. Divergencies between circulating LH and FSH biopotency may reflect a differential regulation of secretion and/or biosynthesis of these hormones. The prolonged early luteal elevation of LH Bio/I is consistent with the idea of a functional role of elevated LH biopotency in the maintenance of the corpus luteum.     

Differential requirement for pulsatile LH during the follicular phase and exposure to the preovulatory LH surge for oocyte fertilization and embryo development in cattle

The requirement for pulsatile LH and the LH surge for the acquisition of oocyte fertilizing potential and embryo developmental competency was examined in Zebu heifers. Follicular growth was superstimulated using the GnRH agonist-LH protocol in which pulsatile LH and the preovulatory LH surge are blocked. In experiment 1, heifers were assigned on Day 7 of the estrous cycle to receive: group 1A (n = 5), 1.5 mg norgestomet (NOR) implant; group 1B (n = 5), GnRH agonist implant. Follicular growth was superstimulated with 2x daily injections of FSH from Day 10 (a.m.) to Day 13 (p.m.), with PGF2alpha injection on Day 12 (a.m.). Heifers were ovariectomized on Day 15 (a.m.) and oocytes were placed immediately into fertilization, without 24 h maturation. Respective cleavage and blastocyst development rates were: group 1A, 0/64 oocytes (0%) and 0/64 (0%); group 1B, 34/70 oocytes (48.6%) and 2/70 (2.9%). In experiment 2, heifers were assigned on Day 7 of the estrous cycle to receive: group 2A (n = 10), 1.5 mg NOR implant; group 2B (n = 10), GnRH agonist implant; group 2C (n = 10), GnRH agonist implant. Follicular growth was superstimulated as in experiment 1 above. Heifers in groups 2A and 2B received an injection of 25 mg LH on Day 14 (p.m.) and all heifers were ovariectomized on Day 15 (a.m.); oocytes were placed immediately into fertilization without 24 h maturation. Cleavage rates were similar for heifers in group 2A (84/175 oocytes, 48.0%), group 2B (61/112 oocytes, 54.5%) and group 2C (69/163, 42.3%). Blastocyst development rates were similar for heifers in group 2A (22/175 oocytes, 12.6%) and group 2B (25/112 oocytes, 22.3%) and lower (P < 0.05) for heifers in group 2C (9/163 oocytes, 5.5%). Oocytes obtained from heifers treated with GnRH agonist, without injection of exogenous LH, underwent cleavage indicating that neither pulsatile LH nor the preovulatory LH surge are obligatory for nuclear maturation in cattle oocytes. Exposure to a surge-like increase in plasma LH increased embryo developmental competency indicating that the preovulatory LH surge promotes cytoplasmic maturation. The findings have important implications for controlling the in vivo maturation of oocytes before in vitro procedures including nuclear transfer.     

Control of gonadotrophin release in Scottish Blackface and Finnish Landrace ewes during seasonal anoestrus

The patterns of LH and FSH secretion were measured in 4 experimental groups of Finnish Landrace and Scottish Blackface ewes: long-term (18 months) ovariectomized ewes (Group 1), long-term ovariectomized ewes with an oestradiol implant, which has been shown to produce peripheral levels of approximately 5 pg/ml (Group 2), long-term ovariectomized ewes with an oestradiol implant for 18 months which was subsequently removed (surgery on Day 0) (Group 3) and short-term ovariectomized ewes (surgery on Day 0) (Group 4). LH and FSH concentrations were monitored in all groups at approximately weekly intervals, before and after Day 0. Finnish Landrace ewes in Groups 1, 2 and 3 had significantly higher mean FSH concentrations than did Scottish Blackface ewes (P less than 0.01). FSH and LH concentrations increased significantly in Groups 3 and 4, but values in Group 4 were significantly lower (P less than 0.01) than those in Group 1 ewes even up to 30 days after ovariectomy. In Group 3, LH concentrations increased to levels similar to those in Group 1. The pattern of LH release was, however, significantly different, with a lower LH pulse frequency (P less than 0.05), but higher pulse amplitude (P less than 0.05). This difference was maintained at least until 28 days after implant removal. We suggest that removal of negative feedback by ovariectomy demonstrates an underlying breed difference in the pattern of FSH secretion and that ovarian factors other than oestradiol are also involved in the negative-feedback control of hypothalamic/pituitary gland function. Furthermore, negative-feedback effects can be maintained for long periods, at least 28 days, after ovariectomy or oestradiol implant removal.     

Peripheral plasma concentrations of oestradiol, progesterone, cortisol, LH and prolactin during the oestrous cycle in the cow, with emphasis on the peri-oestrous period

Interrelationships of circulating hormone levels and their implications for follicular development were studied throughout the oestrous cycle with emphasis on the perioestrous period in heifers and cows. The oestradiol level showed a major peak (45 pmol/1) before and coinciding with oestrus, and a second peak (27 pmol/1) around day 5–6 (day 0: day of first standing oestrus); it was low during the luteal phase of the cycle when progesterone was higher than 14 nmol/1 from day −12 to day −2. Large antral follicles, which had developed during the luteal phase, did not secrete significant amounts of oestradiol, degenerated after luteolysis, and were replaced by a newly developing follicle which became preovulatory. Parallel with this development the oestradiol level increased from the onset of luteolysis to reach a plateau about 26 h before the onset of oestrus. The interval between the onset of luteolysis and the onset of oestrus was 58 h; luteolysis proceeded at a slower rate in heifers than in cows. At 4.6 h after the onset of oestrus the maximum of the LH surge was recorded; the LH surge appeared to be postponed in the period October–December in comparison to the period August–September. The maximum of the LH surge was higher in heifers (45 μg/l) than in cows (30 μg/l), but its duration was similar (8.0 h). The oestradiol level decreased significantly from 6 h after the maximum of the LH surge, and standing oestrus (duration 18 h) was terminated almost at the same time as the return to basal values of oestradiol. Cortisol and prolactin levels did not show a peak during the peri-oestrus period. Cortisol fluctuated irrespective of the stage of the oestrus cycle and prolactin was significantly higher during the luteal phase.

The results of this study indicate that development of the preovulatory follicle starts in the cow at the onset of luteolysis, about 2.5 days before the preovulatory LH surge, and that oestradiol secretion by this follicle is possibly inhibited by the LH surge.     

Effect of photoperiod on LH, FSH and prolactin patterns in ovariectomized oestradiol-treated heifers

Angus and Angus crossbred heifers were ovariectomized, treated with oestradiol implants and randomly assigned to the natural photoperiod of fall to spring for 43 degrees N latitude or extra light simulating the photoperiod of spring to fall. Weekly blood samples were taken for 6 months (fall to spring equinox). All heifers were cannulated every 4 weeks and blood samples were taken for 4 h at 15-min intervals. Sera were assayed for LH, FSH, prolactin and oestradiol. In samples taken weekly, serum LH and FSH concentrations were higher while serum prolactin was lower in heifers exposed to natural photoperiod. There was a photoperiod X time interaction for both FSH and prolactin with concentrations diverging as photoperiod diverged. Circulating concentrations of oestradiol were not different between groups. In samples taken every 4 weeks at 15-min intervals, baseline concentrations of LH and FSH and LH pulse amplitude were higher while prolactin pulse frequency was lower in heifers exposed to natural photoperiod. There was a photoperiod X time interaction for each of these pulsatile characteristics. The correlation between LH and prolactin concentrations estimated from the 15-min samples differed between the two photoperiod treatment groups. The pooled correlation coefficient (r) was -0.12 under natural photoperiod and +0.50 under extra light. There was also a photoperiod X time interaction with negative correlations occurring when photoperiod was decreasing and positive correlations occurring when photoperiod was increasing. These results support the hypothesis that photoperiod alters serum concentrations of LH, FSH and prolactin in cattle.     

Influence of pregnancy on the onset of oestrus and luteal function after prostaglandin-induced luteolysis in cattle

Luteolysis was induced by an injection of 500 micrograms cloprostenol (a prostaglandin (PG) analogue) in pregnant (P) Holstein heifers on Days 17 or 24 of gestation and in non-pregnant (NP) Holstein heifers on Day 17 of the oestrous cycle (oestrus = Day 0). Heifers in Groups P-17 (N = 8) and P-24 (N = 8) were inseminated twice whereas those in Group NP-17 (N = 8) were not inseminated. Immediately after PG injection, embryos were recovered by uterine flushing (400 ml) to confirm pregnancy in Groups P-17 and P-24. Uterine flushing with an equivalent volume of physiological saline was also done in Group NP-17. The interval from PG injection to oestrus and to the peak of luteinizing hormone (LH) as well as profile of increase in plasma oestradiol concentrations during that period did not differ (P greater than 0.1) among the groups. However, the proportion of heifers exhibiting abnormal luteal phases (primarily of short duration) during the oestrous cycle after PG injection was greater (P less than 0.01) in Group P-24 than in Groups NP-17 + P-17 pooled (6/8 vs 3/16). These results suggest that the previous presence of a conceptus did not have any effect on the onset of oestrus, or on plasma concentrations of oestradiol and LH after PG-induced luteolysis on Days 17 or 24 of gestation. However, luteal function during the subsequent oestrous cycle was impaired if heifers were 24 days pregnant when luteolysis was induced.     

Role of increased estradiol on altering the follicle diameters and gonadotropin concentrations that have been reported for double-ovulating heifers

During the preovulatory period in heifers that ovulate from two compared to one follicle, circulating concentrations of estradiol-17β (E2) are greater, diameter of follicles and concentration of FSH are reduced, and the LH surge occurs sooner. The effect of increased E2 on the reported characteristics of double ovulation was studied by treating heifers with 0.07 mg E2, 0.09 mg E2, or vehicle in four treatments at 6-h intervals (n=6 heifers/group), beginning at the time of expected follicle deviation (largest follicle, 8.5mm). There were no significant differences on follicle diameters or hormone concentrations between the 0.07 and 0.09 mg E2 groups, and heifers were combined into one E2 group (n=12). The E2 treatments induced concomitant preovulatory surges in LH and FSH at 34.0 ± 2.6h after first treatment, compared to 57.6 ± 4.5h in the vehicle group (P<0.0002). The E2 treatments did not affect FSH concentrations during the preovulatory gonadotropin surge. The diameter of the preovulatory follicle at the LH peak was smaller (P<0.0001) in the E2-treated group (10.2 ± 0.2mm) than in the vehicle group (13.1 ± 0.6mm). The hypothesis was not supported that the previously reported increase in circulating E2 in heifers with double preovulatory follicles accounts for the reported lesser concentrations in the preovulatory FSH surge in heifers with double ovulations. Hypotheses were supported that the reported earlier occurrence of the preovulatory LH surge and smaller preovulatory follicles in heifers with double ovulations are attributable to the reported increase in E2 from the double preovulatory follicles.     

The acute effects of oestradiol-17beta and synthetic LH-RH on plasma LH levels in freemartin heifers.

Injection of oestradiol was followed by a surge of plasma LH within 24 h in only 7 of 12 freemartins. Elevations of plasma LH were less than those reported for normal non-cyclic heifers, but some freemartins showed a delayed, or more prolonged, LH response. Responsiveness to oestradiol was not related to degree of chimaerism or plasma androstenedione level, and most of the animals responded similarly in two trials carried out 4 months apart, during which time plasma androstenedione levels had more than doubled. Freemartins which showed an LH surge after oestradiol treatment released greater amounts of LH after the injection of LH-RH than did non-responders.     

Close synchrony of ovulation in superstimulated heifers that have a downregulated anterior pituitary gland and are induced to ovulate with exogenous LH

The synchrony of ovulation was examined in superstimulated heifers that had a downregulated pituitary gland and which were induced to ovulate by injection of exogenous LH. The pituitary was downregulated and desensitized to GnRH by treatment with the GnRH agonist deslorelin. Nulliparous heifers (3.5 yr old) at random stages of the estrous cycle were assigned to 1 of 3 groups, and on Day -7 received the following treatments: Group 1 (control, n = 8), 1 norgestomet ear implant; Group 2 (GnRH agonist, n = 8); Group 3 (GnRH agonist-LH protocol, n = 8), 2 deslorelin ear implants. Ovarian follicle growth in all heifers was superstimulated with twice-daily intramuscular injections of FSH (Folltropin-V): Day O, 40 mg (80 mg total dose); Day 1, 30 mg; Day 2; 20 mg; Day 3, 10 mg. On Day 2, all heifers were given a luteolytic dose of PGF (7 A.M.), Norgestomet implants were removed from heifers in Group 1 (6 P.M.). Heifers in Group 3 were given an injection of 25 mg, i.m. porcine LH (Lutropin) on Day 4 (4 P.M.). Ovarian follicle status was monitored at 8-h intervals from Day 3 (8 A.M.) to Day 6 (4 P.M.) using an Aloka Echo Camera and 7.5 MHz transducer. Heifers in Groups 2 and 3 exhibited estrus earlier (P < 0.05) than heifers in Group 1. Heifers in Group 2 did not have a preovulatory LH surge and they did not ovulate. Individual control heifers in Group 1 ovulated between 12 A.M. on Day 5 and 8 A.M. on Day 6. Heifers with deslorelin implants and injected with LH in Group 3 ovulated between 4 P.M. on Day 5 and 8 A.M. on Day 6. It was confirmed that superstimulated heifers with GnRH agonist implants can be induced to ovulate with LH. It was also demonstrated that ovulation is closely synchronized after injection of LH. Thus, a single, fixed-time insemination schedule could be used in a GnRH agonist-LH superovulation protocol, with significant practical and economic advantages for superovulation and embryo transfer programs.     

Association between surges of follicle-stimulating hormone and the emergence of follicular waves in heifers.

The effects of ablation of a dominant follicle and treatment with follicular fluid on circulating concentrations of follicle-stimulating hormone (FSH) were studied and the temporal relationships between surges of FSH and follicular waves were studied in heifers with two or three follicular waves/interovulatory interval. Cauterization of the dominant follicle on Day 3 or Day 5 (ovulation on Day 0) (six control and six treated heifers/day) resulted in a surge (P less than 0.05) in FSH beginning the day after cautery. The FSH surge prior to wave 2 (first post-treatment follicular wave) occurred 4 days (Day 3 cautery) and 2 days (Day 5 cautery) before the surge in control groups, corresponding to a 4-day and a 2-day advance in emergence of wave 2 compared with controls. It was concluded that the dominant follicle on Day 3 and Day 5 was associated with the suppression of circulating FSH concentrations. Heifers (n = 4/group) were untreated or treated intravenously with a proteinaceous fraction of bovine follicular fluid on Days 0-3, 3-6, or 6-11. Concentrations of FSH were suppressed (P less than 0.05) for the duration of treatment, regardless of the days of treatment. Cessation of treatment was followed within 1 day by the start of a surge in FSH. The FSH surge prior to wave 2 occurred 2 days earlier (treatment on Days 0-3), 1 day later (treatment on Days 3-6), and 6 days later (treatment on Days 6-11) than in controls, corresponding to an equivalent advance or delay, respectively, in the emergence of wave 2 compared with controls. The results suggest that the effects of exogenous follicular fluid on follicular development were mediated, in whole or in part, by altering plasma FSH concentrations. Control heifers combined for the two experiments were separated into those with 2-wave (n = 11) or 3-wave (n = 5) interovulatory intervals. Two-wave heifers had two FSH surges and 3-wave heifers had three apparent FSH surges during the interovulatory interval. Results of the cautery and follicular fluid experiments indicated that a surge in FSH necessarily preceded the emergence of a wave. The FSH surges in treated and control heifers began 2-4 days before the detectable (ultrasound) emergence of a follicular wave (follicles of 4 and 5 mm), peaked 1 or 2 days before emergence and began to decrease approximately when the follicles of a wave begin to diverge into a dominant follicle and subordinate follicles (follicles 6-7 mm).