Luteolysis In Heifers Research Articles

Up-regulation of endometrial oxytocin receptor is associated with the timing of luteolysis in heifers with two and three follicular waves†.

Initiation of luteolysis in ruminants is variable due to ill-defined mechanisms. Cycles of two follicular waves are shorter and have earlier luteolysis than three-wave cycles. This study validated a cytobrush technique for evaluating dynamics of endometrial gene expression and associated changes in mRNA with timing of luteolysis, based on circulating progesterone and ultrasound-determined changes in blood flow and volume of corpus luteum (CL). On day 8 (ovulation=day 0), Holstein heifers were randomized into two groups: cytobrush group (n=9) had an endometrial sample collected every 48h from day 8 until end of luteolysis (CL blood flow ≤ 20%) and control group was sampled only once either before (day 12; n=4) or at the end of luteolysis (n=5). Concentrations of progesterone, CL blood flow, CL volume, and the frequency of two and three-wave cycles were similar between groups. Endometrial mRNA for progesterone receptors and estradiol receptors 1 and 2 was greater on day 8 and decreased thereafter similarly in two and three-wave cycles. Oxytocin receptor mRNA increased earlier in two vs three-wave cycles (day 14 vs 18), and the increase was associated with the onset of luteolysis. In conclusion, the cytobrush technique allowed in vivo collection of multiple endometrial samples during the estrous cycle. Endometrial mRNA expression of steroid receptors did not explain the variability in timing of onset of luteolysis in heifers while the later onset of luteolysis in three-wave cycles was associated with later up-regulation of oxytocin receptor mRNA.

Role of luteal biosynthesis of prostaglandin F2α on function and structure of the corpus luteum during luteolysis in heifers

The role of endogenous prostaglandin F2α (PGF) in the induction of luteolysis by exogenous PGF was studied by simultaneous inhibition of endogenous PGF with flunixin meglumine (FM). Groups were controls (n = 8), PGF treated (n = 8), and FM + PGF treated (n = 9). Treatments were given 10 d postovulation at hours 0, 8, and 16. The protocol was based on (1) the assumption that luteolytic characteristics of exogenous PGF would be altered if the synthesis of endogenous PGF is simultaneously inhibited and (2) the reports that luteolysis involves a direct effect of uterine PGF on large luteal cells followed by an effect of the large cells on the small cells. At hour 48, progesterone concentration was greater in the controls (7.6 ± 0.8 ng/mL) than that in the FM + PGF group (3.0 ± 0.5 ng/mL) and lower in the PGF group (0.7 ± 0.3 ng/mL) than in the FM + PGF group (interaction, P < 0.0001). The effects of each of the 3 groups on percentage change in CL volume were similar to the effects on progesterone. At hour 48, the percentage of CL tissue with color-Doppler signals of blood flow was similar between the controls (56.2% ± 3.8%) and FM + PGF group (50.0% ± 6.4%) and lowest in the PGF group (15.6% ± 7.2%) (interaction, P < 0.0001). A resurgence in progesterone concentration began at hours 24 or 48 in 6 of 9 heifers in the FM + PGF group compared to 0 of 8 heifers in each of the other groups (P < 0.007). The progesterone resurgence in the FM + PGF group was associated with the maintenance of percentage of CL tissue with blood-flow signals. The experimental hypothesis that an inhibitor of endogenous PGF reduces the luteolytic response to exogenous PGF was supported.

Intraovarian factors associated with switching of a future dominant follicle to a subordinate follicle during induced luteolysis in heifers

The factors involved in the switching of a future dominant follicle (DF) to subordinate status were studied (n = 42) by induction of luteolysis with PGF2α (hour 0) when the largest follicle (F1) in follicular wave 2 was 7.0 or 8.5 mm. Combined for 7.0- and 8.5-mm groups, the frequency of switching was greater (P < 0.01) when F1 and CL were ipsilateral (10 of 28, 36%) than when contralateral (0 of 14). The frequency of switching in the ipsilateral relationship was greater (P < 0.002) when F1 and CL were adjacent (<3 mm apart; 10 of 17) than when separated (0 of 11). The difference in diameter between F1 and F2 was less (P < 0.005) when switching occurred (0.4 ± 0.1 mm) than when switching did not occur (ipsilateral, 1.3 ± 0.2 mm; contralateral, 1.1 ± 0.2 mm). Treatment at hour 0 when F1 was 7.0 mm and ipsilateral to the CL resulted in smaller diameter (P < 0.001) of F1 at hour 12 (7.6 ± 0.2 mm) than when treatment was not given (8.3 ± 0.1 mm). The hypotheses were supported that (1) switching from a future DF to a future subordinate is functionally related to luteolysis, and (2) factors involved in switching include an ipsilateral relationship between the largest follicle and CL and close intraovarian proximity of the follicle to the regressing CL. It is proposed that switching of the future DF to subordinate status during spontaneous luteolysis accounts for the reported greater frequency of the contralateral than ipsilateral relationships between the preovulatory follicle and CL in three-wave interovulatory intervals.

Circulating nitric oxide metabolites during luteolysis and the effect of luteinizing hormone on circulating nitric oxide metabolites in heifers

Temporal relationships among circulating concentrations of nitric oxide metabolites (NOM), progesterone (P4), and luteinizing hormone (LH) within the hours of a PGFM pulse were studied during luteolysis in heifers. The peak of a PGFM pulse was designated Hour 0. All of the following increases and decreases were significant. Within a spontaneous PGFM pulse (experiment 1; n = 7), concentrations of P4 and LH decreased between Hours −1 and 0 and increased between Hours 0 and 1; NOM increased between Hours −1 and 2. In experiment 2, PGFM pulses were simulated by intrauterine infusion of PGF2α (PGF group, n = 6), and another group was also treated with acyline to inhibit LH secretion (acyline-PGF group, n = 6). Averaged over the two groups, concentration of P4 decreased between Hours −2 and 0, increased (rebounded) between Hours 0 and 1, and decreased after Hour 2. In the PGF group, concentration of LH decreased between Hours −2 and −0.5 and increased between Hour 0 and Hour 1.5 to a maximum at Hour 1.5; NOM decreased between Hours −2 and −1.5 and increased between Hours 0 and 1.5. In the acyline-PGF group, the effect of hour was not significant for concentrations of LH and NOM. The absence of an increase in NOM concentration when LH was inhibited is a novel finding. The hypotheses were supported that concentrations of LH and NOM are temporally related, and LH has a role in the increase in NOM within the hours of a PGFM pulse.

Life or Death Decisions in the Corpus Luteum

The corpus luteum (CL) is an ephemeral endocrine organ. During its lifespan, it undergoes a period of extremely rapid growth that involves hypertrophy, proliferation and differentiation of the steroidogenic cells, as well as extensive angiogenesis. The growth phase is followed by a period in which remodelling of the tissue ceases, but it engages in unparalleled production of steroids, resulting in extraordinarily high metabolic activity within the tissue. It is during this stage that a critical juncture occurs. In the non-fertile cycle, uterine release of prostaglandin (PG)F(2α) initiates a cascade of events that result in rapid loss of steroidogenesis and destruction of the luteal tissue. Alternatively, if a viable embryo is present, signals are produced that result in rescue of the CL. This review article summarizes the major concepts related to the fate of the CL, with particular focus on recent insights into the mechanisms associated with the ability of PGF(2α) to bring about complete luteolysis. It has become clear that the achievement of luteolysis depends on repeated exposure to PGF(2α) and involves coordinated actions of heterogeneous cell types within the CL. Together, these components of the process bring about not only the loss in progesterone production, but also the rapid demise of the structure itself.

Stimulation of a pulse of LH and reduction in PRL concentration by a physiologic dose of GnRH before, during, and after luteolysis in heifers

A single physiologic dose (5.0μg) of GnRH was given to 9 heifers each day (Hour 0) beginning on Day 15 postovulation until regression of the corpus luteum. Blood samples were taken each day for Hours −3, −2, −1, 0, 0.25, 0.5, 0.75, 1, 1.25, 1.50, 1.75, 2, 3, 4, and 5. Based on daily progesterone concentrations, data were grouped into phases of before (n=4), during (n=8), and after (n=7) luteolysis. The number of LH pulses with a peak at pretreatment Hours −2 or −1 (0.35±0.12pulses/sampling session) was less (P<0.0001) than for a pulse peak at posttreatment Hours 1 or 2 (1.0±0.0pulses/session). The characteristics and effects of LH pulses on progesterone and estradiol were similar between natural (pretreatment) and primarily induced (posttreatment) LH pulses. The same dose of GnRH stimulated an LH pulse with greater (P<0.05) amplitude after luteolysis than during luteolysis. Concentrations of PRL and number and prominence of PRL pulses decreased (P<0.05) between Hours 0 and 2 within each of the phases of before, during, and after luteolysis. The hypothesis that a physiologic dose of GnRH increases the concentration of PRL was not supported; instead, GnRH reduced the concentration of PRL. Results supported the hypotheses that an appropriate dose of GnRH stimulates an LH pulse during luteolysis that is similar to a natural pulse in characteristics and in the effects on progesterone and estradiol.

Effects of inhibition of prostaglandin F2α biosynthesis during preluteolysis and luteolysis in heifers

Flunixin meglumine (FM; 2.5 mg/kg) was given to heifers at three 8-h intervals, 16 d after ovulation (first treatment = Hour 0) to inhibit the synthesis of prostaglandin F2α (PGF), based on plasma concentrations of a PGF metabolite (PGFM). Blood samples were collected at 8-h intervals from 15 to 18 d in a vehicle (control) and FM group (n = 16/group). Hourly samples were collected from Hours −2 to 28 in 10 heifers in each group. Heifers that were in preluteolysis or luteolysis at Hour 0 based on plasma progesterone (P4) concentrations at 8-h intervals were partitioned into subgroups. Concentration of PGFM was reduced (P < 0.05) by FM treatment in each subgroup. For the preluteolytic subgroup, the first decrease (P < 0.05) in P4 concentration after Hour 0 occurred at Hours 24 and 40 in the vehicle and FM groups, respectively. Plasma P4 concentrations 32 and 40 h after the beginning of luteolysis in the luteolytic subgroup were greater (P < 0.05) in the FM group. Concentration at the peak of a PGFM pulse in the FM group was greater (P < 0.05) in the luteolytic than in the preluteolytic subgroup. The peak of a PGFM pulse occurred more frequently (P < 0.001) at the same hour as the peak of an LH fluctuation than at the ending nadir of an LH fluctuation. In conclusion, a reduction in prominence of PGFM pulses during luteolysis delayed completion of luteolysis, and treatment with FM inhibited PGFM production more during preluteolysis than during luteolysis.

In vivo intra-luteal implants of prostaglandin (PG) E 1 or E 2 (PGE 1, PGE 2) prevent luteolysis in cows. I. Luteal weight, circulating progesterone, mRNA for luteal luteinizing hormone (LH) receptor, and occupied and unoccupied luteal receptors for LH

Previously, it was reported that chronic intra-uterine infusion of PGE 1 or PGE 2 every four hours inhibited luteolysis in ewes. However, estradiol-17β or PGE 2 given intra-uterine every 8 h did not inhibit luteolysis in heifers, but infusion of estradiol + PGE 2 inhibited luteolysis in heifers. The objective of this experiment was to determine whether and how intra-luteal implants containing PGE 1 or PGE 2 prevent luteolysis in Angus or Brahman cows. On day-13 post-estrus, Angus cows received no intra-luteal implant and corpora lutea were retrieved or Angus and Brahman cows received intra-luteal silastic implants containing Vehicle, PGE 1, or PGE 2 and corpora lutea were retrieved on day-19. Coccygeal blood was collected daily for analysis for progesterone. Breed did not influence the effect of PGE 1 or PGE 2 on luteal mRNA for LH receptors or unoccupied or occupied luteal LH receptors did not differ ( P > 0.05) so the data were pooled. Luteal weights of Vehicle-treated Angus or Brahman cows from days-13–19 were lower ( P < 0.05) than those treated with intra-luteal implants containing PGE 1 or PGE 2. Day-13 Angus luteal weights were heavier ( P < 0.05) than Vehicle-treated Angus cows on day-19 and luteal weights of day-13 corpora lutea were similar ( P > 0.05) to Angus cows on day-19 treated with intra-luteal implants containing PGE 1 or PGE 2. Profiles of circulating progesterone in Angus or Brahman cows treated with intra-luteal implants containing PGE 1 or PGE 2 differed ( P < 0.05) from controls, but profiles of progesterone did not differ ( P > 0.05) between breeds or between cows treated with intra-luteal implants containing PGE 1 or PGE 2. Intra-luteal implants containing PGE 1 or PGE 2 prevented ( P < 0.05) loss of luteal mRNA for LH receptors and unoccupied or occupied receptors for LH compared to controls. It is concluded that PGE 1 or PGE 2 alone delays luteolysis regardless of breed. We also conclude that either PGE 1 or PGE 2 prevented luteolysis in cows by up-regulating expression of mRNA for LH receptors and by preventing loss of unoccupied and occupied LH receptors in luteal tissue.

Circulating hormone concentrations within a pulse of a metabolite of prostaglandin F2α during preluteolysis and early luteolysis in heifers

Plasma from hourly blood samples from two previous studies in heifers was assayed for hormones that were not considered in the previous reports. The objective was to determine the intrapulse temporal relationships between a prostaglandin F2α metabolite (PGFM) pulse and various hormones during preluteolysis and luteolysis. Hormone concentrations were centralized to the peak of a PGFM pulse (Hour 0) and evaluated from Hours −3 to 3. Experiment 1 ( n = 6) was done during early luteolysis. Progesterone decreased during Hours −3 to 0 and then rebounded, but did not return to prepulse concentrations, and concentration of LH increased between Hours −1 and 2. In Experiment 2 ( n = 7), comparisons were made between the last PGFM pulse of preluteolysis and the first pulse of luteolysis. Intrapulse concentrations of LH increased rapidly between Hours 0 and 1 during preluteolysis and gradually between Hours −2 and 2 during luteolysis. Intrapulse differences in cortisol among hours were not significant for preluteolysis and approached significance ( P < 0.06) during luteolysis, owing primarily to an apparent increase between Hours −2 and 1. Oxytocin concentrations showed only an hour effect ( P < 0.0003) from to an increase between Hours −2 and 0 and a decrease between Hours 0 and 2. Results indicated that oxytocin and PGFM concentrations increased and decreased concomitantly and supported the hypothesis that the reported more prominent rebound in progesterone during the descending portion of a PGFM pulse during preluteolysis than during luteolysis involves a greater transient increase in LH.

Intrapulse temporality between pulses of a metabolite of prostaglandin F 2α and circulating concentrations of progesterone before, during, and after spontaneous luteolysis in heifers

Pulses of the prostaglandin F 2α (PGF) metabolite 13,14-dihydro-15-keto-PGF 2α (PGFM) and the intrapulse concentrations of progesterone were characterized hourly during the preluteolytic, luteolytic, and postluteolytic periods in seven heifers. The common hour of the end of preluteolysis and the beginning of luteolysis was based on a progressive progesterone decrease when assessed only at the peaks of successive oscillations. The end of the luteolytic period was defined as a decrease in progesterone to 1 ng/mL. Blood samples were taken hourly from 15 d after ovulation until luteal regression as determined by color-Doppler ultrasonography. Between Hours −2 and 2 (Hour 0 = PGFM peak) of the last PGFM pulse of the preluteolytic period, progesterone decreased between Hours −1 and 0, and then returned to the prepulse concentration. Concentration did not change significantly thereafter until a PGFM pulse early in the luteolytic period; progesterone decreased by Hour 0 and transiently rebounded after Hour 0, but not to the prepulse concentration. In the later portion of the luteolytic period, progesterone also decreased between Hours −1 and 0 but did not rebound. After the defined end of luteolysis, progesterone decreased slightly throughout a PGFM pulse. Results demonstrated for the first time that the patterns of progesterone concentrations within a PGFM pulse differ considerably among the preluteolytic, luteolytic, and postluteolytic periods.

Luteal blood flow and concentrations of circulating progesterone and other hormones associated with a simulated pulse of 13,14-dihydro-15-keto-prostaglandin F2α in heifers

Progesterone and luteal blood flow effects of an i.u. 2-h infusion of 0.25 mg/h of prostaglandin F(2)(alpha) (PGF) that simulated a natural pulse of 13,14-dihydro-15-keto-PGF (PGFM) were compared to the effects of a single bolus i.u. injection of PGF (4 mg) that induced complete luteolysis in heifers. Blood sampling and an estimate of the percentage of luteal area with colour-Doppler signals of blood flow were performed every 2 min for 20 min and less frequently thereafter for 6 h. After the beginning of PGF infusion or a bolus injection, progesterone increased to a peak at 14 and 10 min respectively, and was accompanied by an increase in blood flow in the bolus group but not in the infusion group. Progesterone then decreased for 1 or 2 h and was accompanied by a continued elevation in blood flow in the PGF bolus group and by a slight increase in the PGF infusion group. Progesterone then rebounded in both groups, but the rebound was greater in the infusion group. Blood flow decreased during the descending arm of the progesterone rebound. Cortisol and prolactin began to increase 6 min after the bolus PGF injection but did not increase during or after PGF infusion. The increases in cortisol, prolactin and blood flow after a PGF bolus treatment but not during a simulated PGFM pulse indicated that the bolus treatment was pharmacologic, and its use may lead to faulty conclusions on the nature of physiologic luteolysis. The comparisons between progesterone and blood flow are novel.

Role of follicular estradiol-17beta in timing of luteolysis in heifers.

The hypothesis was tested that estradiol (E2) from the ovarian follicles controls time of luteolysis. Time of luteolysis was evaluated by multiple measures of corpus luteum (CL) structure (area, volume) and function (progesterone [P4], luteal blood flow). The hypothesis for experiment 1 was that repeated ablation of follicles would reduce circulating E2 and delay luteolysis. Heifers were randomly assigned on Day 9 (Day 0 = ovulation) to three groups. All follicles >or=4 mm were ablated on Day 9 (group FA9; n = 6); Days 9-15 (group FA15; n = 6); or Days 9-21 (group FA21; n = 7). As expected, follicular ablation delayed (P < 0.001) the rise in circulating E2 and peak E2 concentrations (FA9, Day 17.6 +/- 0.7; FA15, Day 20.3 +/- 0.3; FA21, Day 24.9 +/- 0.3). Luteolysis (based on each measure) was delayed (P < 0.005) by repeated ablation of follicles, with earlier luteolysis (based on P4 decrease) in FA9 (Day 15.2 +/- 0.8) than FA15 (Day 16.5 +/- 0.4), and a further delay in FA21 (Day 18.3 +/- 0.5). The hypothesis of experiment 2 was that exogenous treatment with E2 would stimulate prostaglandin F(2alpha) (PGF) secretion and prevent the delay in luteolysis associated with follicular ablations. Follicles >or=4 mm were ablated from Day 9 to Day 17 (n = 15). Heifers were treated on Days 13 and 15 with 1.0 mg of estradiol benzoate (FAE2; n = 7) or vehicle (FAV; n = 8). Treatment with E2 induced PGF secretion (detected by PGF metabolite) and induced earlier (P < 0.02) luteolysis in FAE2 than in FAV, whether determined by circulating P4 or by area, volume, or blood flow of CL. In summary, ablation of follicles (>or=4 mm) delayed and treatment with E2 hastened luteolysis in heifers with ablated follicles. Thus, these results are consistent with an essential role for follicle E2 in timing of luteolysis.

Reduction in Circulating Estradiol by Ablation of Follicles Delays Luteolysis in Heifers.

Circulating estradiol (E2) and progesterone (P4) may regulate timing of prostaglandin F2α (PGF2α) release from the uterus. We hypothesized that removal of circulating E2 by ablating follicles >4 mm would therefore delay luteolysis. On Day 9 (Day 0 = ovulation), Holstein heifers were randomized by replicate into three groups. Follicle ablations (all follicles ≥ 4 mm ablated) were done either once on Day 9 (FA9; n=6), on Days 9, 11, 13 and 15 (FA9-15; n=6), or on Days 9, 11, 13, 15, 17, 19 and 21 (FA9-21; n=7). A duplex B-mode and pulsed-wave color-Doppler ultrasound scanner equipped with a linear array 7.5 MHz transrectal transducer was used to estimate corpus luteum (CL) blood flow, based on percentage of the CL with color-Doppler signals during real time scanning. Blood samples were collected daily and plasma assayed by RIA for P4 and E2 concentrations. The CL blood-flow and hormone data were analyzed by SAS MIXED procedure with a REPEATED statement for main effects of group and day and their interaction. Both main effects and the interaction were significant (P<0.01) for P4, E2, and CL blood flow. Mean P4 concentrations first began to decrease (P<0.02) on Day 14 in FA9 and were lower (P<0.05) than in the other two groups on Days 15 and 16. In FA9-15, P4 first began to decline on Day 16 (P<0.01) and was lower (P<0.05) than in FA9-21 on Days 17, 18 and 19. In FA9-21, the first P4 decrease (P<0.01) occurred on Day 19, and there was no significant difference among groups after Day 19. For CL blood flow, the first decrease (P<0.05) began in FA9 on Day 15 and the values were lower (P<0.05) than in FA9-15 on Day 16. In FA9-15, the first decrease (P<0.05) in blood flow occurred on Day 17 and the values were lower (P<0.05) from Day 17 to Day 21 than in FA9-21. The first decrease (P<0.05) in CL blood flow for FA9-21 occurred on Day 19. In FA9, the E2 concentration began to increase (P<0.01) on Day 13 and the values were higher (P<0.05) from Day 14 to Day 17 than in FA9-15 and FA9-21. In FA9-15, the first increase (P<0.01) in E2 occurred on Day 16, whereas in FA9-21 the first increase (P<0.04) occurred on Day 22. Follicular ablation delayed the rise in circulating E2 with earlier (P<0.002) mean peak E2 concentrations in FA9 (17.6±0.7 days after ovulation) than in FA9-15 (20.3±0.3 d) and FA9-21 (24.9±0.3 d). In FA9 and FA9-15 luteolysis did not begin until after the last ablation and 1 day after the first continuous increase in E2 plasma concentration, but in FA9-21 luteolysis began 3 days before the last ablation and first continuous increase in E2 concentration. The first significant decreases in blood flow occurred one day after the first significant decrease in P4 (FA9) or both occurred on the same day (FA9-15, FA9-21). Sequential ablations of all follicles (≥4 mm) delayed both functional (decreased P4) and structural (decreased CL blood flow) indicators of luteolysis. Thus, normal follicle growth was essential for normal timing of luteolysis in heifers. It seems likely that the delay in luteolysis caused by follicular ablation was due to reduced circulating E2, but luteolysis occurred before a detectable increase in plasma E2 concentration when the CL was maintained for long period as in the FA9-21 group.

Effect of estradiol valerate on ovarian follicle dynamics and superovulatory response in progestin-treated cattle

Three experiments evaluated the effects of estradiol valerate (EV) on ovarian follicular and CL dynamics, intervals to estrus and ovulation, and superovulatory response in cattle. Experiment 1 compared the efficacy of two norgestomet ear implants (Crestar and Syncro-Mate B; SMB) for 9 d (with PGF at implant removal), combined with either 5 mg estradiol-17β and 100 mg progesterone (EP) or 5 mg EV and 3 mg norgestomet (EN) im at the time of implant insertion on CL diameter and follicular wave dynamics. Ovaries were monitored by ultrasonography. There was no effect of norgestomet implant. Diameter of the CL decreased following EN treatment ( P < 0.01). Mean (± S.D.) day of follicular wave emergence (FWE) was earlier ( P < 0.0001) and less variable ( P < 0.0001) in EP- (3.6 ± 0.5 d) than in EN- (5.7 ± 1.5 d) treated heifers. Intervals from implant removal to estrus ( P < 0.001) and ovulation ( P < 0.01) were shorter in EN- (45.7 ± 11.7 and 74.3 ± 12.6 h, respectively) than in EP- (56.4 ± 14.1 and 83.3 ± 17.0 h, respectively) treated heifers. Experiment 2 compared the efficacy of EP versus EN in synchronizing FWE for superovulation in SMB-implanted cows. At random stages of the estrous cycle, Holstein cows ( n = 78) received two SMB implants (Day 0) and were randomly assigned to receive EN on Day 0 or EP on Day 1. Folltropin-V treatments were initiated on the evening of Day 5, with PGF in the morning and evening of Day 8, when SMB were removed. Cows were inseminated after the onset of estrus and embryos were recovered 7 d later. Non-lactating cows had more CL (16.7 ± 11.3 versus 8.3 ± 4.9) and total ova/embryos (14.7 ± 9.5 versus 7.9 ± 4.6) than lactating cows ( P < 0.05). EP-treated cows tended ( P = 0.09) to yield more transferable embryos (5.6 ± 5.2) than EN-treated cows (4.0 ± 3.7). Experiment 3 compared the effect of dose of EV on ovarian follicle and CL growth profiles and synchrony of estrus and ovulation in CIDR-treated beef cows ( n = 43). At random stages of the estrous cycle (Day 0), cows received a CIDR and no further treatment (Control), or an injection of 1, 2, or 5 mg im of EV. On Day 7, CIDR were removed and cows received PGF. Follicular wave emergence occurred within 7 d in 7/10 Control cows and 31/32 EV-treated cows ( P < 0.05). In responding cows, interval from treatment to FWE was longer ( P < 0.05) in those treated with 5 mg EV (4.8 ± 1.2 d) than in those treated with 1 mg (3.2 ± 0.9 d) or 2 mg (3.4 ± 0.8 d) EV, while Control cows were intermediate (3.8 ± 2.0 d). Diameter of the dominant follicle was smaller ( P < 0.05) at CIDR removal and tended ( P = 0.08) to be smaller just prior to ovulation in the 5 mg EV group (8.5 ± 2.2 and 13.2 ± 0.6 mm, respectively) than in the Control (11.8 ± 4.6 and 15.5 ± 2.9 mm, respectively) or 1 mg EV (11.7 ± 2.5 and 15.1 ± 2.2 mm, respectively) groups, with the 2 mg EV group (10.7 ± 1.5 and 14.3 ± 1.7 mm, respectively) intermediate. Diameter of the dominant follicle at CIDR removal was less variable ( P < 0.01) in the 2 and 5 mg EV groups than in the Control group, and intermediate in the 1 mg EV group. In summary, treatment with 5 mg EV resulted in a longer and more variable interval to follicular wave emergence than treatment with 5 mg estradiol-17β, which affected preovulatory dominant follicle size following progestin removal, and may have also affected superstimulatory response in Holstein cows. Additionally, 5 mg EV appeared to induce luteolysis in heifers, reducing the interval to ovulation following norgestomet removal. Conversely, intervals to, and synchrony of, follicular wave emergence, estrus and ovulation following treatment with 1 or 2 mg EV suggested that reduced doses of EV may be more useful for the synchronization of follicular wave emergence in progestogen-treated cattle.

Oxytocin modulates the pulsatile secretion of prostaglandin F2αin initiated luteolysis in cattle

Subluteolytic doses of prostaglandin F2αanalogue (oestrophan) given i.m. and oxytocin (OT) antagonist (CAP) and noradrenaline (NA) infused into the abdominal aorta were used to test the importance of lutealOTin pulsatile secretion of prostaglandin F2α(PGF) during luteolysis in heifers (n = 17). In experiment 1, heifers were pre-infused for 30 minutes with saline on either day 17 of the oestrous cycle (group 1; n = 4) or on day 18 of the oestrous cycle (group 2; n = 3), and with CAP (8 mg per animal) on day 17 of the oestrous cycle (group 3; n = 4). Next, heifers were injected with oestrophan (30 μg per animal). Injection of oestrophan in Group 3 increasedOTconcentrations (P < 0.001) to values similar to those observed during spontaneous luteolysis (50 to 70 pg ml–1).PGFMconcentrations in this group also increased (P < 0.001), but were lower (P < 0.05) than the values in groups 1 and 2.CAPgiven prior to oestrophan decreased bothPGFMelevation (P < 0.06) and its area under the curve (P < 0.01), compared to the saline pretreated heifers. In experiment 2NA(4 mg) was infused twice for 30 minutes at five hour intervals to releaseOTon day 17 of the oestrous cycle (n = 6). However, during hormone analysis it appeared that three of six heifers had elevatedPGFMconcentrations (group 1) and three others did not (group 2).NAcaused the correlated increase of progesterone andOTsecretion (r = 0.68; P < 0.05) in both groups but it only influencedPGFsecretion in group 1 only (P < 0.05).We postulate thatOTcan amplify and modulate the course of induced luteolysis as a regulator of the amplitude of pulsatilePGFsecretion.PGFanalogue stimulates secretion of endogenousPGFfrom the uterus in cattle and this may be an important component of the luteolytic response to exogenousPGF.

The effect on luteolysis by intensive oral administration of flunixin granules in heifers.

The objective of the study was to investigate whether oral administration of flunixin meglumine (FM) in the form of granules could prolong the functional life of the corpus luteum in heifers. Previous studies have shown that intensive, i.e. four times daily parental administration of FM can postpone luteolysis. Twelve heifers received an oral dose of 2.2 mg flunixin per kg body weight. Three dosing regimes were used; twice (n = 2), thrice (n = 4) and four times daily (n = 6). The 9-day-treatment period started on Day 14/15 of the oestrous cycle. Blood samples were collected twice daily during the entire experimental period. Frequent samples were withdrawn from Day 14/15 for 10 and 15 days in the control and treatment oestrous cycles, respectively, at 0600, 0800, 1000, 1200, 1400, 1600, 1800 and at 2000 h. The plasma was analysed for the content of the main metabolite of PGF2 alpha, i.e. 15-ketodihydro-PGF2 alpha, and progesterone. A control cycle preceded the treatment cycle so that each heifer acted as its own control. The length of the oestrous cycle was significantly increased when FM was administered thrice, from 18-22 days to 20-24 days (P < 0.05), and four times daily, from 18-21 days to 25-27 days (P < 0.01), but not in the twice daily dosing regime (one-way ANOVA). When FM was administered twice and thrice, luteolysis occurred during treatment. However, when the four times daily regime was used, luteolysis was obtained when the treatment had terminated. No changes in progesterone levels were recorded, although the luteal phase increased when the oestrous cycle length was prolonged. The number of PG-pulses decreased significantly, from 6-12 pulses to 0-3 pulses (P < 0.01), when FM was administered four times daily. A reduction was also observed when heifers received the drug thrice, but the decrease was not significant. The oral route of administration was found to be as effective as the parental one to affect the mechanism responsible for luteolysis in heifers. However, to inhibit and postpone luteolysis, administration of FM four times daily is a necessity. Our results show that oral administration of FM in the bovine species can be of value both in research as well as in the clinic, e.g. to support the luteal function.

The effect of oral administration of flunixin on luteolysis in heifers

The effect of oral administration of flunixin on luteolysis in heifers

Effects of progesterone and oestradiol-17 beta on oxytocin-induced release of prostaglandin F-2 alpha in heifers.

Seven bilaterally ovariectomized heifers were used in 4 experiments and received: (1) saline injections, as control; (2) one injection of oestradiol (3 mg; i.v.); (3) two i.v. injections of oxytocin (100 i.u.) 6 h apart; or (4) one oestradiol injection 30 min after the first oxytocin injection and a second oxytocin injection 6 h later. All experiments were performed without progesterone and then after 7, 14 and 21 days of progesterone treatment. Frequent blood samples were taken for 1 h before and 7 h after the first injection of oxytocin or oestradiol for the measurement of 13,14-dihydro-15-keto-PGF-2 alpha (PGFM) by radioimmunoassay. After 7, 14 and 21 days of progesterone priming, oestradiol caused a significant increase (P less than 0.001) in plasma PGFM after 6 h but not before. After 7, 14 and 21 days of progesterone, there was a significant increase (P less than 0.005) in PGFM after the first oxytocin injection and a similar increase following the second. The oxytocin-induced increase in PGFM after 14 and 21 days of progesterone was significantly higher (P less than 0.001) 6 h after oestradiol injection than before the oestradiol injection. There was no significant effect of oestradiol on the response to oxytocin in animals that received no progesterone or in those animals that received progesterone for only 7 days. These results show that, under the influence of progesterone, oestradiol enhances the oxytocin-induced release of PGF-2 alpha, and suggest a possible synergistic action of these hormones for the induction of luteolysis in heifers.

Effect of an estrogen antagonist (tamoxifen) on cloprostenol-induced luteolysis in heifers.

Experiments were conducted to determine the role of estrogens on endogenous PGF2 alpha secretion and luteolysis following injection of cloprostenol in heifers. In Exp. 1, eight luteal-phase heifers were used to evaluate tamoxifen (T) as an estrogen antagonist. Heifers received T (35 mg i.v.) or ethanol:saline vehicle (ES) every 4 h for 44 h. All received cloprostenol (500 micrograms i.m.) immediately after the start of T or ES, and received estradiol-17 beta (500 micrograms i.m.) 12 h later. Each ES heifer had a surge of luteinizing hormone (LH) within 48 h of estradiol injection, whereas T-treated heifers did not. Estrus was observed in three ES-treated heifers, but not in T-treated heifers. In Exp. 2, 10 heifers received T (35 mg i.v.) or ES every 4 h for 64 h beginning on d 15 postestrus. Cloprostenol (500 micrograms i.m.) was injected 16 h after the start of treatment. Concentrations of LH were similar (P greater than .05) in both groups. In ES heifers, concentrations of 13,14-dihydro-15-keto-prostaglandin F2 alpha (PGFM) increased; in T-treated heifers, PGFM remained at pre-cloprostenol levels. Luteolysis was induced in all heifers. Progesterone (P4) decreased to less than or equal to 1 ng/ml at similar (P greater than .05) rates in ES-treated and T-treated heifers. Mean concentration of P4 288 h post-cloprostenol was greater (P less than .05) in ES-treated than in T-treated heifers. Three ES-treated heifers, but no T-treated heifers, were in standing estrus. We conclude that T effectively antagonizes estrogen in cattle.(ABSTRACT TRUNCATED AT 250 WORDS)

Changes in Inhibin-Like Bioactivity in Ovulatory and Atretic Follicles and Utero-Ovarian Venous Blood after Prostaglandin-Induced Luteolysis in Heifers*

The objectives of this study were to develop a bioassay for measuring inhibin bioactivity in untreated samples of bovine follicular fluid (BFF) and then examine changes in inhibin bioactivity in ovulatory and atretic follicles and utero-ovarian venous blood during the periovulatory period in heifers. A rat pituitary cell culture system was used to bioassay inhibin-like activity. Addition of 0.005 to 1 microliter untreated (whole), unfiltered charcoal-stripped, or filtered whole BFF to pituitary cultures caused a linear suppression of LHRH-induced FSH release but had no effect on LH secretion. Steroids in BFF did not suppress FSH secretion, since removal of steroids from BFF with charcoal did not remove the FSH-suppressive activity in BFF. Addition of ether extracts of BFF caused a slight but nonparallel suppression of FSH secretion; however, heating these extracts removed most of this suppressive activity. Removal of BFF from pituitary cultures completely restored the capacity of pituitary cultures to respond to LHRH. It was concluded that the inhibin bioassay was specific for detecting inhibin-like activity in fluids from individual follicles without interference of steroids. Within 12 h after a prostaglandin (PG) injection during the luteal phase of heifers, LH levels in serum increased 2- to 4-fold and remained at this level until the occurrence of the preovulatory gonadotropin surge. In contrast, FSH did not change before the gonadotropin surge. Inhibin bioactivity was measured in all follicles (greater than or equal to 6 mm) 0, 12, 24, 36, 48, 60, and 72 h after and in utero-ovarian venous serum 0, 24, and 36 h after PG-induced luteolysis. From 0-36 h after PG administration, inhibin-like activity increased linearly in presumed ovulatory follicles and utero-ovarian venous serum. Then, from 48-72 h after PG treatment, before the preovulatory LH surge, inhibin activity decreased in ovulatory follicles. After the surge but before ovulation, inhibin-like activity increased in ovulatory follicles. Inhibin-like activity in atretic follicles did not change after PG treatment and was lower in atretic than ovulatory follicles. Since a single hypothalamic releasing factor, LHRH, may control the secretion of LH and FSH, increased secretion of inhibin from preovulatory follicles before the preovulatory LH and FSH surges could account for the absence of a presurge rise in FSH in blood, as was observed for LH during this time in heifers. Diminished follicular production of inhibin during the gonadotropin surge could explain the preovulatory release of FSH along with LH during this time.