Ultrasound and endocrine evaluation of the ovarian response to PGF2alpha given at different stages of the luteal phase in ewes

The purpose of this study was to evaluate the ovarian response of ewes to two treatments with PGF2alpha using transrectal ovarian ultrasonography and hormone measurements. Fifteen milligrams of PGF2alpha was given to six cyclic Western White Face (WWF) ewes early in the estrous cycle (Days 4 to 7) and to six late in the cycle (Days 10 to 12 after ovulation), and a second treatment was given 9 days after the first. Ultrasound scanning and blood sampling started 7 days prior to the first PGF2alpha treatment and ended 10 days (scanning) or 19 days (blood sampling) after the second PGF2alpha treatment, for both groups of ewes. Mean ovulation rate (2.6 +/- 0.7) did not differ significantly between the ewes first treated early or late in the cycle, or after the first or second treatments with PGF2alpha. The time from treatment to ovulation was longer in ewes first treated early (4.0 +/- 0.3 days) compared to late (2.8 +/- 0.4 days) in the cycle (P < 0.05). Both the number of ovulations (range: 0-7) and time from treatment to ovulation (range: 1-9 days) were highly variable. This variability appeared to be due to the extension of the life span of ovulating follicles that emerged prior to PGF2alpha administration and also ovulation of some follicles that emerged after treatment. When results for first and second treatments were pooled, the total number of follicles > 5 mm in diameter on the day of treatment that failed to ovulate in response to PGF2alpha was higher in ewes first treated early (0.8 +/- 0.2/ewe) compared to late (0.3 +/- 0.2/ewe) in the cycle (P < 0.05). The proportion of detected luteal structures relative to the number of ovulations was lower in ewes first treated early compared to late in the cycle (60 and 86%, respectively; P < 0.05). Disruption of ovulatory follicle dynamics and normal luteogenesis, and variability in the timing of ovulation after PGF2alpha treatments could all contribute to poor or variable fertility when prostaglandins are used for estrus synchronization.     

Response of the 1-5 day-aged ovine corpus luteum to prostaglandin F2alpha

The hypothesis that, in the ewe, prostaglandin (PG) F2alpha administration on day 3 after ovulation is followed by luteolysis and ovulation was tested using 24 animals. The ewes were treated with a dose of a PGF2alpha analogue (delprostenate, 160 microg) on days 1 (n=8), 3 (n=8) or 5 (n=8) after ovulation, was established by transrectal ultrasonography. Daily scanning and blood sampling were performed to determine ovarian changes and progesterone serum concentrations by radioinmunoassay. The treatment induced a sharp decrease of progesterone concentrations followed by oestrus and ovulation in all ewes treated on days 3 and 5 and in one ewe treated on day 1 (8/8, 8/8, 1/8; P<0.05). Seven ewes treated on day 1 did not respond to PGF2alpha treatment and had an inter-ovulatory cycle of normal length (17.4 +/- 0.5 days). However, the profile of progesterone concentrations during the cycle of these ewes was delayed 1 day (P<0.05) compared with a control cycle. The overall interval between PGF2alpha and oestrus for the 17 responding ewes was 42.4 +/- 2.3 h. In 15 of these ewes the ovulatory follicle was originated from the first follicular wave and the ovulation occurred at 60.8 +/- 1.8 h after PGF2alpha treatment. The other two responding ewes ovulated an ovulatory follicle originated from the second follicular wave between 72 and 96 h after treatment. These results support the hypothesis and suggest that refractoriness to PGF2alpha of the recently formed corpus luteum (CL) may be restricted to the first 1-2 days post-ovulation.     

Effects of a 6-day treatment with medroxyprogesterone acetate after prostaglandin F2 alpha-induced luteolysis at midcycle on antral follicular development and ovulation rate in nonprolific Western white-faced ewes

Medroxyprogesterone acetate (MAP) from intravaginal sponges prolongs the lifespan of large ovarian follicles when administered after prostaglandin F2alpha (PGF2alpha)-induced luteolysis early in the luteal phase of ewes. The present study was designed to determine whether a PGF2alpha/MAP treatment applied at midcycle would alter the pattern of antral follicle growth and increase ovulation rate in nonprolific ewes. A single injection of PGF2alpha (15 mg, i.m.) was given, and an intravaginal MAP (60 mg) sponge was inserted for 6 days, on approximately Day 8 after ovulation, in 7 (experiment 1), 8 (experiment 2) or 11 (experiment 3) ultrasonographically monitored, cycling Western white-faced ewes; seven ewes (experiment 1) served as untreated controls. Blood samples were collected each day and also every 12 min for 6 h, halfway through the period of treatment with MAP (experiment 1), or every 4 h, from 1 day before to 1 day after sponging (experiment 2). Seventeen of 26 treated ewes (experiment 1, n = 6; experiment 2, n = 5; experiment 3, n = 6) ovulated 1 to 6 days after PGF2alpha, but this did not affect the emergence of ensuing follicular waves (experiments 1 and 2). These ovulations, confirmed by laparotomy and histological examinations of the ovaries (experiment 3), were not preceded by an increase in LH/FSH secretion and did not result in corpora lutea, as evidenced by transrectal ultrasonography and RIA of serum progesterone (experiments 1 and 2). Following the removal of MAP sponges, the mean ovulation rate was 3.1 +/- 0.4 in treated ewes and 2.0 +/- 0.3 in control ewes (experiment 1; P < 0.05). In experiments 1 and 2, the ovulation rate after treatment (3.1 +/- 0.4 and 2.8 +/- 0.4) was also greater than the pretreatment rate (1.9 +/- 0.3 and 1.9 +/- 0.1, respectively). Ovulations of follicles from two consecutive waves before ovulation were seen in five treated but only in two control ewes (experiment 1), and in seven ewes in experiment 2. There were no significant differences between the MAP-treated and control ewes in mean daily serum concentrations of FSH and estradiol, and no differences in the parameters of LH/FSH secretion, based on frequent blood sampling. Treatment of nonprolific Western white-faced ewes with PGF2alpha and MAP at midcycle changed follicular dynamics and increased ovulation rate by approximately 50%. These effects of MAP, in the absence of luteal progesterone, may not be mediated by changes in gonadotropin secretion.     

Does injection of prostaglandin F(2alpha) (PGF2alpha) cause ovulation in anestrous Western White Face ewes?

In a previous study in our laboratory, treatment of non-prolific Western White Face (WWF) ewes with PGF(2 alpha) and intravaginal sponges containing medroxyprogesterone acetate (MAP) on approximately Day 8 of a cycle (Day 0 = first ovulation of the interovulatory interval) resulted in ovulations during the subsequent 6 days when MAP sponges were in place. Two experiments were performed on WWF ewes during anestrus to allow us to independently examine if such ovulations were due to the direct effects of PGF(2 alpha) on the ovary or to the effects of a rapid decrease in serum concentrations of progesterone at PGF(2 alpha)-induced luteolysis. Experiment 1: ewes fitted with MAP sponges for 6 days (n = 12) were injected with PGF(2 alpha) (n = 6; 15 mg im), or saline (n = 6) on the day of sponge insertion. Experiment 2: ewes received progesterone-releasing subcutaneous implants (n = 6) or empty implants (n = 5) for 5 days. Six hours prior to implant removal, all ewes received a MAP sponge, which remained in place for 6 days. Ewes from both experiments underwent ovarian ultrasonography and blood sampling once daily for 6 days before and twice daily for 6 days after sponge insertion. Additional blood samples were collected every 4 h during sponge treatment. Experiment 1: 4-6 (67%) PGF(2 alpha)-treated ewes ovulated approximately 1.5 days after PGF(2 alpha) injection; these ovulations were not preceded by estrus or a preovulatory surge release of LH, and resulted in transient corpora hemorrhagica (CH). The growth phase was longer (P < 0.05) and the growth rate slower (P < 0.05) in ovulating versus non-ovulating follicles in PGF(2 alpha)-treated ewes. Experiment 2: in ewes given progesterone implants, serum progesterone concentrations reached a peak (1.7 2 ng/mL; P < 0.001) on the day of implant removal and decreased to basal concentrations (<0.17 ng/mL; P < 0.001) within 24 h of implant removal. No ovulations occurred in either the treated or the control ewes. We concluded that ovulations occurring after PGF(2 alpha) injection, in the presence of a MAP sponge, could be due to a direct effect of PGF(2 alpha) at the ovarian level, rather than a sudden decline in circulating progesterone concentrations.     

Ultrasound and endocrine evaluation of the ovarian response to a single dose of 500 IU of eCG following a 12-day treatment with progestogen-releasing intravaginal sponges in the breeding and nonbreeding seasons in ewes

A standard dose of 500 IU of eCG is commonly given to progestogen pre-treated anestrous ewes for induction of estrus. Twelve seasonally anestrous and 12 cyclic Western White Face ewes were treated for 12 days with intravaginal sponges impregnated with medroxyprogesterone acetate (MAP). In trials in both the breeding and nonbreeding seasons, six randomly selected ewes were given 500 IU of eCG at sponge removal to determine the effects of low dose of eCG on ovarian antral follicular dynamics and ovulation. Ultrasound scanning and blood sampling were done daily. Treatment with eCG did not have marked effects on antral follicular growth. All ewes ovulated, except for five of six control anestrous ewes. Luteal structures and progesterone secretion were confirmed in all but the control anestrous ewes. In the breeding season, peak progesterone concentrations were greater (P<0.05) in eCG-treated compared to control ewes. Daily serum estradiol concentrations were greater in the periovulatory period in eCG-treated compared to control ewes (treatment-by-day interaction; P<0.05), particularly in anestrus. Progestogen-treated ewes ovulated follicles from several follicular waves, in contrast to ovulations of follicles from the final wave of the cycle in untreated, cyclic ewes. Anestrous ewes exhibited more frequent follicular waves and FSH peaks compared to cyclic ewes after a progestogen/eCG treatment. In conclusion, 500 IU of eCG given after 12 days of progestogen treatment had limited effects on the dynamics of ovarian follicular waves. However, eCG treatment increased serum concentrations of estradiol during the periovulatory period, particularly in anestrous ewes; this probably resulted in the synchronous estrus and ovulation in anestrous ewes.     

The effect of subluteal levels of exogenous progesterone on follicular dynamics and endocrine patterns during early luteal phase of the ewe

Nineteen Corriedale ewes were treated with an im dose of a PGF2alpha during the luteal phase to synchronize estrus. After ovulation had been detected by using ultrasonography (Day 0); the ewes were randomly assigned to 2 different groups. In 11 ewes a CIDR, which had previously been used for 10 d, was inserted on the fourth day after ovulation. The ewes then received a dose of PGF2alpha on Day 5 to induce luteolysis. The CIDR remained in place until the end of the experiment (Day 9). Control ewes (n = 8) received no treatment. Blood samples were taken daily for estradiol, progesterone and FSH determinations. In the untreated ewes, 2 follicular waves were detected in all of the animals throughout the monitoring period, with a mean wave interval of 4.5 d. The total number of follicles which were > or =2 mm decreased from Day 0 to Day 4 (8.8+/-1.0 to 5.3+/-0.6; P< or =0.05) and then increased at Day 7 (7.5+/-0.9; P< or =0.05). The growth profiles of both the largest and the second largest follicles of Wave 1 showed significant divergence, while no divergence was observed in Wave 2. Serum estradiol concentrations decreased significantly from the day before to the day of ovulation and then increased again during the growing phase of the largest follicle of Wave 1. Concentrations of FSH were high on the day of emergence of both waves, but while a significant decline was observed after emergence in Wave 1, the levels remained high in Wave 2. In 8 of the 11 treated ewes, the largest follicle of Wave 1 was still present on the ninth day after ovulation (persistent follicle). In the other 3 ewes, the largest follicle of Wave 1 was already regressing on the day that the treatment was administered, and the largest follicle that was present on Day 9 originated from Wave 2 (nonpersistent follicle). In persistent follicle ewes, the largest follicle of Wave 1 prolonged its lifespan significantly, attaining the maximum diameter (Day 8.1+/-0.8) later than in untreated (Day 3.0+/-0.4) and nonpersisted follicle ewes (Day 2.0+/-0.6). The total number of follicles decreased in persistent follicle ewes between Day 0 and Day 4 (7.9+/-1.5 to 4.5+/-0.5, respectively; P< or =0.05) and remained low until the end of the experiment. Progesterone concentrations (nmol/L) between Days 6 and 9 were significantly different between untreated and persistent follicle ewes (12.8+/-1.0 vs. 9.4+/-1.0, P< or =0.02). The present study confirms that the largest follicle of Wave 1 is dominant in the ewe and that subluteal progesterone concentrations can prolong its lifespan and extend this dominance.     

Follicular growth, ovulation and embryo recovery in dairy cows given FSH at the beginning or middle of the estrous cycle

Variability in the superovulation response is an important problem for the embryo transfer industry. The objective of this study was to determine whether FSH treatment at the beginning of the cycle would improve the ovulation rate and embryo yield in dairy cows. Twenty-eight postpartum cyclic dairy cows were allocated at random to 4 treatment groups (A, B, C and D). Group A cows (n = 10) received FSH (35 mg) at a decreasing dose, starting on Day 9 (Day 0 = day of estrus) for 5 days followed by PGF(2alpha) (35 mg) on Day 12. Cows assigned to Groups B, C and D (n = 6 cows each, respectively) were given 35 mg FSH at a decreasing dose from Days 2 to 6 followed by PGF(2alpha) on Day 7. Group C and D cows received PRID inserts from Day 3 to Day 7. Cows in Group D additionally received 1000 IU hCG 60 hours after PGF(2alpha) treatment. Ovaries were scanned daily using a real time ultrasound scanner from the beginning of FSH treatment until embryo recovery, to monitor follicular development, ovulation and the number of unovulated follicles. Embryos were recovered from the uterus by a nonsurgical flushing technique 7 days after breeding. There were no differences (P>0.01) in the number of follicles > 10 mm at 48 hours after PGF(2alpha) treatment among the 4 groups. The mean numbers of follicles were 10.6 +/- 1.2, 9.3 +/- 1.3, 12.2 +/- 1.3 and 15.0 +/- 2.9 for Groups A, B, C and D, respectively. A significantly (P<0.001) higher number of ovulations was observed and a larger number of embryos was recovered in Group A than in the other groups. The results of this study indicate that superovulation with FSH at the beginning of the cycle causes sufficient follicular development but results in very low ovulation and embryo recovery rates.     

Ovarian synchronization following ultrasound-guided transvaginal follicle ablation in heifers

In Experiments 1 and 2, ultrasound-guided transvaginal follicle aspiration was used as a method of follicle ablation to induce and synchronize subsequent follicular wave emergence and enhance ovulation synchrony following PGF(2alpha) administration. Heifers were at unknown stages of the estrous cycle at the start of both experiments in which all follicles >/=5 mm in diameter were ablated; luteolysis was induced 4 d later with cloprostenol (500 ug/dose, im). In Experiment 1, heifers were randomly assigned to either an ablation (n=17) or a procedural control (no follicle ablation, n=17) group. Ablation-induced wave emergence was indicated by a significant increase in the total number of follicles >/=5 mm within 2 d of ablation (mean, 1.5 d), which was preceded by a significant surge in circulating FSH. Although the mean (+/-SEM) interval from PGF(2alpha) administration to ovulation did not differ between follicle-ablated heifers (5.1+/-0.5 d range, 3 to 9 d) and control heifers (5.1+/-1.0 d; range, 1 to 5 d), the variability of the interval was different (P<0.05). Inequality of variance between the 2 groups was attributed to a greater (P<0.08) degree of ovulation synchrony in the ablation group than in the control group; 13 16 (81%) versus 9 17 (53%), respectively, ovulated within 5 d of cloprostenol administration. Relative asynchrony of ovulations in control heifers was associated with the status of the follicular wave at the time of PGF(2alpha) administration and, in part, to incomplete luteolysis following a single dose of PGF(2alpha). Experiment 2 was designed to examine the efficacy of 2 doses of cloprostenol 12 h apart (n=7) versus a single dose (n=8) to induce complete luteolysis subsequent to follicle ablation-induced wave emergence. Two doses of cloprostenol potentiated ovulation synchrony; more (P<0.05) 2-dose heifers (7 7 , 100%) than single-dose heifers (4 8 , 50%) ovulated within 5 d after PGF(2alpha) administration. In summary, ultrasound-guided transvaginal follicle ablation, done at random during the estrous cycle, induced and synchronized subsequent follicular wave emergence, and resulted in a high degree of ovulation synchrony among heifers after PGF(2alpha) induced luteolysis, especially when 2 doses of PGF(2alpha) were administered 12 h apart.     

Periovulatory changes in the local release of vasoactive peptides, prostaglandin f(2alpha), and steroid hormones from bovine mature follicles in vivo

We previously proposed that an endothelin-angiotensin-atrial natriuretic peptide system may contribute to inducing ovulation of mature bovine follicles by modulating follicular secretion of steroids and prostaglandins (PGs). Thus, this study aimed to determine the real-time changes in the local release of angiotensin II (Ang II), endothelin (ET), atrial natriuretic peptide (ANP), PGF(2alpha), and steroid hormones from bovine mature follicles during the periovulatory period in vivo. Seven cows were treated for superovulation using FSH and PGF(2alpha) injections. Two dialysis capillary membranes per follicle were surgically implanted into the theca layer of mature follicles and connected to a microdialysis system (MDS). Fractions of the perfusate were collected from Day -1 (Day 0 = LH surge) to Day 3. Five out of seven treated cows were normally ovulated, and the newly formed corpora lutea were observed at the end of the experiment. In these five ovulated cows, the release of estradiol, androstenedione, and progesterone in the theca layer increased (P < 0.05) synchronously with the LH surge. Acute increases in PGF(2alpha) and Ang II concentrations in the ovarian venous plasma (OVP) were observed at 24-48 h after the peak of the LH surge, when multiple ovulations were expected to occur. The follicular Ang II release was low during the pre-LH surge period and rose (P < 0.05) at the beginning of the increase in the LH surge. On the other hand, ET-1 release dropped (P < 0.05) when plasma LH started to increase. However, no clear changes in ANP concentration in the MDS perfusate and plasma were observed. The above local changes in Ang II, PGF(2alpha), as well as steroid hormones were not observed in cows (n = 2) that did not show an LH surge and ovulation. The present results demonstrate for the first time the local release of Ang II, ET-1, and ANP from the bovine mature follicle in real-time in vivo and show that Ang II and PGF(2alpha) concentrations in the OVP acutely increase around the time of ovulation. The overall results support the concept of a local functional ET-Ang-ANP system in the bovine mature follicle that may be involved in the ovulatory process.     

Effects of unilateral ovariectomy on follicular development and ovulation in cattle

In a study of 4 cyclic dry cows (Trial I) and 6 cyclic puberal heifers (Trial II), unilateral ovariectomy increased the number of ovulatory follicles, did not alter the hormone profile, cycle length or the number of follicular waves. Ovarian follicular development in all 4 cows was monitored daily using transrectal ultrasonography until the day of ovulation, during which period daily blood samples were also taken from the tail vein for determination of plasma FSH, LH and P4 concentrations. Unilateral ovariectomy was performed on the day after ovulation and ovarian activity was again monitored daily (ultrasonography and blood sampling for FSH, LH and P4) for 2 consecutive cycles (8 cycles in all). Estrus in all 6 heifers was synchronized using 2 injections of PGF2 alpha given 12 d apart. Similarly, ovarian activity in the 6 puberal heifers was monitored daily using ultrasonography and blood sampling for 1 complete control cycle. Following estrus and ovulation the left ovary was removed in all the animals, and thereafter 1 complete cycle was followed. Mean cycle length, FSH, LH and P4 concentrations before and after unilateral ovariectomy were compared using paired sample t-test. The results show that unilateral ovariectomy neither altered the cycle length nor the number of follicular waves in the cows, but it increased the number of ovulatory follicles (2 follicles developed and ovulated in 6 of the 8 cycles). The mean diameter of the largest follicle was 16.1 +/- 0.9 mm and the second largest 12.5 +/- 0.9 mm. No significant (P > 0.05) differences were observed in FSH (0.72 +/- 0.09 vs 0.71 +/- 0.07), LH (0.42 +/- 0.1 vs 0.37 +/- 0.07) and P4 (2.8 +/- 0.6 vs 2.6 +/- 0.4) levels before and after unilateral ovariectomy. Of the 6 heifers, 5 had 2 waves and 1 heifer had 3 waves of follicular growth during the control cycle, and this pattern did not change after the procedure. Mean cycle length (20.7 +/- 0.9 vs 21 +/- 0.9) did not differ before and after unilateral ovariectomy, and 4 of the 6 heifers ovulated twin follicles following ovariectomy. The mean diameter of the largest follicle was 14.5 +/- 0.7 mm and second largest measured 12.1 +/- 0.8 mm. No significant (P > 0.05) differences were observed in FSH (0.16 +/- 0.09 vs 0.21 +/- 0.07), LH (0.11 +/- 0.1 vs 0.15 +/- 0.07) and P4 levels (3.6 +/- 0.26 vs 3.8 +/- 0.29) before and after unilateral ovariectomy. Based on these results, we conclude that unilateral ovariectomy is an ideal method for obtaining twin ovulations in cows and heifers.     

Prostaglandin F2alpha-induced estrus in ewes exhibiting estrous cycles of different duration

Effects of prostaglandin F(2alpha) (PGF(2alpha)), administered during the mid-luteal phase of the estrous cycle, were examined in ewes exhibiting estrous cycles classified as short (< or =16.5 days, short-cycle ewes, n = 10) or long (> or =18 days, long-cycle ewes, n = 9) based on the durations of two estrous cycles (cycles -2 and -1) before treatment. The ewes received (i.m.) 20mg of PGF(2alpha) on day 10 of the third estrous cycle (cycle 0) followed, 36 h later, by 25 microg of gonadotropin releasing hormone (GnRH) to time the events of ovulation. Duration of subsequent estrous cycles +1 and +2 were recorded, and then the ewes were treated with the same combination of PGF(2alpha) and GnRH beginning on day 10 of estrous cycle +3. Ovaries were recovered 6h after GnRH administration to assess development of pre-ovulatory follicles. The proportion of ewes that exhibited estrus after PGF(2alpha) and GnRH treatment on cycle 0 was not different (P > 0.05) between short- and long-cycle ewes. Onset of estrus occurred sooner (P < 0.05) after PGF(2alpha) injection in short-cycle ewes than in long-cycle ewes (1.9 +/- 0.1 days and 2.3 +/- 0.1 days, duration of cycle 0 was 11.9 and 12.3 days, respectively). Duration of estrous cycle +1 was 1.2 days longer (P < 0.01) than cycle -1 in short-cycle ewes. However, duration of estrous cycle +1 did not change (P > 0.05) after PGF(2alpha) and GnRH administration in ewes having long cycles. Pre-ovulatory follicles did not differ (P > 0.05) in numbers, diameter, layers of granulosa cells nor concentrations of progesterone and estradiol-17beta in follicular fluid between short- and long-cycle ewes after PGF(2alpha) and GnRH treatment. In conclusion, ewes having short or long estrous cycles responded differently to PGF(2alpha) and GnRH treatment with respect to the interval to onset of estrus and duration of the subsequent estrous cycle.     

Examination of the relative role of FSH and LH in the mechanism of ovulatory follicle selection in sheep

The GnRH-antagonist suppression-ovarian autotransplant model (n = 18) was used to examine the relative roles of temporal changes in FSH and LH stimulation on follicle development and selection. Follicle development was stimulated by infusion with oFSH for 3 days and treatments applied for 60 h after progestagen sponge withdrawal and before delivery of an ovulatory stimulus. In Expt 1, there was continuous infusion of FSH with or without small amplitude high frequency LH pulses, or withdrawal of FSH with or without pulsatile LH. In Expt 2, there was acute or gradual withdrawal of FSH at sponge withdrawal with pulsatile LH. The patterns of follicle development and basal and pulsatile ovarian hormone secretion were determined. The maintenance of FSH throughout the artificial follicular phase resulted in multiple follicle development and ovulation (3.3 +/- 0.3). Pulsatile LH stimulated steroid secretion (P < 0.001) but had little effect on ovulation rates (3.8 +/- 0.8) when FSH was maintained. However, withdrawal of FSH in the absence of LH resulted in atresia of the ovulatory follicles and anovulation whereas, when FSH was withdrawn in the presence of LH, preovulatory follicle development was maintained in some animals (3/6 and 5/9 in Expts 1 and 2, respectively) and these ewes had lower (P < 0.05) ovulation rates (1-2 ovulations per ewe). When FSH was withdrawn gradually in the presence of pulsatile LH, 9/9 animals ovulated with ovulation rates in the normal range. These results indicate that ovulatory follicles can transfer their gonadotrophic dependence from FSH to LH. It is hypothesized that the ability of a follicle to respond to this switch in gonadotrophic support is central to the mechanism of follicle selection.     

Temporal relationships among estrus, body temperature, milk yield, progesterone and luteinizing hormone levels, and ovulation in dairy cows

Estrous cycles of 10 postpartum cyclic Holstein cows were synchronized using prostaglandin f(2alpha) (PGF(2alpha)) given twice 12 d apart to study the relationship of the onset of estrus, body temperature, milk yield, luteinizing hormone (LH) and progesterone concentration to ovulation. Blood samples and body temperatures (vaginal and rectal) were taken every 4 h until ovulation, starting 4 h prior to the second PGF(2alpha) treatment. All cows were observed for estrus following the second administration of PGF(2alpha). Ultrasound scanning of the ovaries commenced at standing estrus and thereafter every 2 h until the disappearance of the fluid filled preovulatory follicle (ovulation). Two cows failed to ovulate and became cystic following the second PGF(2alpha) treatment. The remaining eight cows exhibited a decline in progesterone to <1.0 ng/ml within 28 h, standing estrus and a measurable rise (> 1.0 degrees C) in vaginal but not rectal temperature, and ovulated 90 +/- 10 h after the second PGF(2alpha) treatment. Onset of standing estrus, LH peak and vaginal temperature were highly correlated (P<0.05) with time of ovulation (0.82, 0.81 and 0.74, respectively). Intervals to ovulation tended to depend upon parity. Pluriparous (n = 4) and biparous (n = 4) cows ovulated within 24 and 30 +/- 3 h from the onset of standing estrus; 22 and 31 +/- 2 h from the LH peak; and 22 and 27 +/- 3 h from peak vaginal temperature (mean +/- standard error of the mean), respectively. The results indicated that the onset of standing estrus and rise in vaginal temperature are good practical parameters for predicting ovulation time in dairy cattle.     

Effects of FSH commercial preparation and follicular status on follicular growth and superovulatory response in Spanish Merino ewes

Ovarian follicular development was characterized in 24 Spanish Merino ewes to study effects of the follicular status and the FSH commercial product used on follicular growth and subsequent superovulatory response. Estrus was synchronized using 40 mg fluorogestone acetate sponges. The superovulatory treatment consisted in 2 daily i.m. injections of FSH from 48 h before to 12 h after sponge removal. Sheep were assigned randomly to 2 groups treated with 6 decreasing doses (4, 4, 3, 3, 2, 2 mg) of FSH-P or with 6 doses of 1.25 mL of OVAGEN. Growth and regression of all follicles > or = 2 mm were observed by transrectal ultrasonography, and recorded daily from Day 6 before sponge insertion to the first FSH injection, and then twice daily until estrus was detected with vasectomized rams. Differences were detected in follicular development from the first FSH injection to detection of estrus (-48 to 36 h from sponge removal) between groups. Administration of FSH-P increased the appearance of new follicles with respect to OVAGEN (6.3 +/- 0.7 vs 4.8 +/- 0.4; P < 0.05), and the mean number of medium (4 to 5 mm) follicles (8.9 +/- 1.2 vs 6.6 +/- 0.9; P < 0.05). However, the mean number of follicles that regressed in size after sponge removal (5.9 +/- 0.4 vs 3.3 +/- 0.4) and the number of preovulatory sized follicles that did not ovulate (60 vs 42.4%) were also higher in FSH-P treated ewes (P < 0.05). So, finally, there were no differences in ovulation rate, as determined by laparoscopy on Day 7 after sponge removal, between ewes treated with FSH-P or OVAGEN (6.3 +/- 1.9 vs 7.0 +/- 1.7 CL). In all the ewes, the ovulatory response was related (P < 0.05) both to the number of small follicles (2 to 3 mm in diameter) present in the ovaries at the start of treatment with exogenous FSH and to the number of follicles that reached > or = 4 mm in size at estrus, despite differences in the pattern of follicular development when using different commercial products.     

Follicular and FSH dynamics in ewes with a history of high and low ovulation rates

Daily transrectal ultrasound scanning and twice-daily blood sampling were used to monitor the temporal relationships between FSH concentrations and follicle development during complete interovulatory intervals for ewes in which the ovulation rate in each of the 2 previous years was high or low (> or = 3 and < or = 2 ovulations, respectively). Follicles that reached > or = 5 mm were used to define a follicular wave and were tracked retrospectively to 3 mm (emergence). The hypothesis that FSH surges (identified with a computer program) and follicular waves (retrospectively determined based on ultrasound scanning) are temporally associated was supported in both groups by the emergence of an anovulatory or ovulatory follicular wave near the peak of an FSH surge. Further support for the hypothesis was a significant increase in FSH concentrations before and a significant decrease after follicular-wave emergence in both groups independent of the identification of FSH surges. Ewes with a history of high ovulation rates had smaller follicles (anovulatory and ovulatory) and more ovulations, but the 2 groups were similar in the number of ovulatory follicular waves and associated FSH surges, number and characteristics of the FSH surges, and mean FSH concentrations per interovulatory interval. Surges of FSH were periodic (every 3 or 4 d) regardless of the ovulation-rate group or follicle response. In ewes with a low ovulation rate, the nonovulatory FSH surges were most frequently associated with emergence of detected anovulatory follicular waves. In ewes with a high ovulation rate, more FSH surges were not associated with a detected follicular wave, as defined, presumably because the largest follicle did not reach 5 mm. The results indicated that the factors resulting in a high ovulation rate were not exerted through circulatory patterns or concentrations of FSH but involved a shorter growth phase and smaller maximal diameter of follicles.     

Synchronization of follicular wave emergence in the seasonally anestrous ewe: the effects of estradiol with or without medroxyprogesterone acetate

Fertility is often lower in anestrous compared to cyclic ewes, after conventional estrus synchronization. We hypothesized that synchronization of ovarian follicular waves and ovulation could improve fertility at controlled breeding in anestrous ewes. Estradiol-17beta synchronizes follicular waves in cattle. The objectives of the present experiments were to study the effect of an estradiol injection, with or without a 12-d medroxyprogesterone acetate (MAP) sponge treatment, on synchronization of follicular waves and ovulation in anestrous ewes. Twenty ewes received sesame oil (n=8) or estradiol-17beta (350 microg; n=12). Eleven ewes received MAP sponges for 12d and were treated with oil (n=5) or estradiol-17beta (n=6) 6d before sponge removal. Saline (n=6) or eCG (n=6) was subsequently given to separate groups of ewes at sponge removal in the MAP/estradiol-17beta protocol. Estradiol treatment alone produced a peak in serum FSH concentrations (4.73+/-0.53 vs. 2.36+/-0.39 ng/mL for treatment vs. control; mean+/-S.E.M.) after a short-lived (6 h) suppression. Six of twelve ewes given estradiol missed a follicular wave around the time of estradiol injection. Medroxyprogesterone acetate-treated ewes given estradiol had more prolonged suppression of serum FSH concentrations (6-18 h) and a delay in the induced FSH peak (32.3+/-3.3 vs. 17.5+/-0.5 h). Wave emergence was delayed (5.7+/-0.3 vs. 1.4+/-0.7d from the time of estradiol injection), synchronized, and occurred at a predictable time (5-7 vs. 0-4d) compared to ewes given MAP alone. All ewes given eCG ovulated 3-4d after injection; this predictable time of ovulation may be efficacious for AI and embryo transfer.     

Response of anestrous ewes to the ram effect after follicular wave synchronization with a single dose of estradiol-17beta

Anestrous ewes respond to the introduction of rams with either an ovulation within 2-3 days that may be followed by luteal phases of normal or short length, with delayed ovulations (5-6 days later), or with the luteinization of follicles. The aim of this work was to study the relationship between the growth status of the largest follicle present when rams are introduced and the type of ovarian response in non-treated ewes and in ewes treated with estradiol-17beta before ram introduction. Thirteen anestrous Corriedale ewes were divided into 2 groups: E2 (n = 7) and C (n = 6). The E2 ewes received a single dose of 50 microg estradiol-17beta 5 days before the introduction of the rams to synchronize the onset of their follicle waves, while C ewes remained untreated. When the rams were introduced, all E2 ewes had the largest follicle in a growing stage in contrast with the C ewes (3 out of 6; P < 0.05). Five C and 4 E2 ewes ovulated after the introduction of the rams (Day 3.4 +/- 0.4 for C vs. 4.8 +/- 0.3 for E2 ewes, respectively, P < 0.05). Only one ewe from each group developed a normal luteal phase: 4 C and 3 E2 ewes had short luteal phases. One C ewe and 2 E2 ewes had short luteal phases originating from follicles that did not ovulate. After the first luteal phase, all ewes returned to anesirus without a second ovulation or luteal phase. The remaining E2 ewe did not ovulate or show any changes in progesterone serum concentrations. We conclude that the growth status of the largest follicle alone does not determine the ovarian responding pattern of anestrous ewes to the ram effect.     

Ovarian function in ewes during the transition from breeding season to anoestrus

Transrectal ovarian ultrasonography was conducted in six Western white-faced ewes for 35 days from the last oestrus of the breeding season, to record the number and size of all ovarian follicles > or = 3 mm in diameter and luteal structures. Blood samples were collected once a day for estimation of serum concentrations of follicle-stimulating hormone (FSH), oestradiol and progesterone. Each ewe had five follicular waves (follicles growing from 3 to > or = 5 mm in diameter) over the scanning period. The duration of the growth phase of the largest ovarian follicles did not differ (P > 0.05) between waves, but follicular static and regressing phases decreased significantly (P < 0.05) after the decline in serum progesterone concentrations at the end of the last luteal phase of the breeding season. The intervals between the five follicular waves were: 9.2+/-0.4, 5.2+/-0.7, 8.3+/-0.8 and 5.8+/-0.7 days; the two shorter intervals differed (P < 0.05) from the two longer intervals. Using the cycle-detection program, rhythmic increases in serum FSH concentrations were detected in all ewes; the amplitude, duration and periodicity of FSH fluctuations did not vary (P > 0.05) throughout the period of study. The number of identified FSH peaks (7.8+/-0.5 peaks per ewe, per scanning period) was greater (P < 0.05) than the number of emerging follicular waves. Serum concentrations of oestradiol remained low (< or = 1 pg/ml) on most days, in five out of the six ewes studied, and sporadic elevations in oestradiol secretion above the non-detectable level were not associated with the emergence of follicular waves. The ovulation rate was lower than that seen during the middle portion of the breeding season (November-December) in white-faced ewes but the transitional ewes had larger corpora lutea (CL). Maximal serum concentrations of progesterone appeared to be lower and the plateau phase of progesterone secretion appeared to be shorter during the last luteal phase of the ovulatory season in comparison to the mid-breeding season of Western white-faced ewes. During the transition into anoestrus in ewes, the endogenous rhythm of FSH release is remarkably robust but the pattern of emergence of sequential follicular waves is dissociated from FSH and oestradiol secretion. Luteal progesterone secretion is suppressed because of fewer ovulations and diminished total luteal volume, but it may also result from diminished gonadotropic support. These season-related alterations in the normal pattern of ovine ovarian cycles appear to be due to reduction in ovarian responsiveness to gonadotropins and/or attenuation in secretion of luteinizing hormone (LH) occurring at the onset of the anovulatory season in ewes.     

Use of bovine follicular fluid to increase ovulation rate or prevent ovulation in sheep

Romney ewes were injected intramuscularly once or twice daily for 3 days with 0, 0.1, 0.5, 1 or 5 ml of bovine follicular fluid (bFF) treated with dextran-coated charcoal, starting immediately after injection of cloprostenol to initiate luteolysis on Day 10 of the oestrous cycle. There was a dose-related suppression of plasma concentrations of FSH, but not LH, during the treatment period. On stopping the bFF treatment, plasma FSH concentrations 'rebounded' to levels up to 3-fold higher than pretreatment values. The mean time to the onset of oestrus was also increased in a dose-related manner by up to 11 days. The mean ovulation rates of ewes receiving 1.0 ml bFF twice daily (1.9 +/- 0.2 ovulations/ewe, mean +/- s.e.m. for N = 34) or 5.0 ml once daily (2.0 +/- 0.2 ovulations/ewe, N = 25) were significantly higher than that of control ewes (1.4 +/- 0.1 ovulations/ewe, N = 35). Comparison of the ovaries of ewes treated with bFF for 24 or 48 h with the ovaries of control ewes revealed no differences in the number or size distribution of antral follicles. However, the large follicles (greater than or equal to 5 mm diam.) of bFF-treated ewes had lower concentrations of oestradiol-17 beta in follicular fluid, contained fewer granulosa cells and the granulosa cells had a reduced capacity to aromatize testosterone to oestradiol-17 beta and produce cyclic AMP when challenged with FSH or LH. No significant effects of bFF treatment were observed in small (1-2.5 mm diam.) or medium (3-4.5 mm diam.) sized follicles. Ewes receiving 5 ml bFF once daily for 27 days, from the onset of luteolysis, were rendered infertile during this treatment period. Oestrus was not observed and ovulation did not occur. Median concentrations of plasma FSH fell to 20% of pretreatment values within 2 days. Thereafter they gradually rose over the next 8 days to reach 60% of pretreatment values where they remained for the rest of the 27-day treatment period. Median concentrations of plasma LH increased during the treatment period to levels up to 6-fold higher than pretreatment values. When bFF treatment was stopped, plasma concentrations of FSH and LH quickly returned to control levels, and oestrus was observed within 2 weeks. The ewes were mated at this first oestrus and each subsequently delivered a single lamb.     

Superovulatory response of ovarian follicles of Wave 1 versus Wave 2 in heifers

Experiments were designed to test the hypotheses that ovarian follicular response to superstimulatory treatment initiated during Wave 1 is equivalent to that of Wave 2, and recovery rate and quality of ova embryos derived from follicles of Wave 1 are equivalent to those derived from follicles of Wave 2. In a preliminary experiment (Experiment 1), heifers were given Folltropin-V (20 mg NIH-FSH-P1, im, bid for 5 d) beginning the day after emergence of the first (n=10) or second (n=10) follicular wave of the estrous cycle, equivalent to approximately Day 1 and Day 10, respectively (Day 0=ovulation). Luteolysis was induced with cloprostenol (500 mug im, bid) on the fourth day of treatment. Fewer (P<0.05) ovulations per heifer were induced in the Wave 1 group than in the Wave 2 group (4.6+/-1.0 vs 9.1+/-1.3). However, the interval from wave emergence to initiation of treatment was found, in retrospect, to have been longer (P<0.05) in the Wave 1 group, i.e., treatment was initiated relatively later with respect to wave emergence. Experiment 2 was designed to correct this disparity and to initiate the same treatment protocol on the day of wave emergence rather than the day after (n=21 per Wave group). There was no difference between Wave 1 and Wave 2 groups in the interval from wave emergence to initiation of treatment (0.4+/-0.1 d), the number of ovulations detected by ultrasonography (6.6+/-1.0 vs 8.2+/-1.7), the number of CL detected at slaughter (6.5+/-0.9 vs 8.1+/-1.8), the total number of ova embryos recovered (5.2+/-0.7 vs 5.1+/-0.8), or the number of fertilized embryos collected (2.8+/-0.6 vs 3.0+/-0.6). In addition, there was no difference between groups in the proportion of heifers that ovulated in either experiment; collectively, luteolysis and ovulation was induced in 58 of 60 heifers. The results supported the general hypothesis that follicles and oocytes of the first and second follicular waves are equivalent in the response to superstimulatory treatment. Regardless of which follicular wave, initiation of treatment near the time of wave emergence appears critical for maximal superovulatory response. Because of the consistency in the time of emergence of Wave 1 (day of ovulation) and equivalence in superovulatory response, use of Wave 1 rather than subsequent follicular waves may be more convenient and time-sparing in superovulation programs; the day of estrus (day before ovulation) may be used as a consistent point of reference for the start of treatment.