Effects of cortisol on pregnancy rate and corpus luteum function in heifers: an in vivo study

To determine whether glucocorticoids affect the function of the bovine corpus luteum (CL) during the estrous cycle and early pregnancy, we examined the effects of exogenous cortisol or reduced endogenous cortisol on the secretion of progesterone (P4) and on pregnancy rate. In preliminary experiments, doses of cortisol and metyrapone (an inhibitor of cortisol synthesis) were established (n=33). Cortisol in effective doses of 10 mg blocked tumor necrosis factor-induced prostaglandin F(2α) secretion as measured by its metabolite (PGFM) concentrations in the blood. Metyrapone in effective doses of 500 mg increased the P4 concentration. Thus, both reagents were then intravaginally applied in the chosen doses daily from Day 15 to 18 after estrus (Day 0) in noninseminated heifers (n=18) or after artificial insemination (n=36). Pregnancy was confirmed by transrectal ultrasonography between Days 28-30 after insemination. Plasma concentrations of P4 were lower in cortisol-treated heifers than in control heifers on Days 17 and 18 of the estrous cycle (P<0.05). However, the interestrus intervals were not different between control and cortisol-treated animals (P>0.05). Moreover, metyrapone increased P4 and prolonged the CL lifespan in comparison to control animals (P<0.05). Interestingly, in inseminated heifers, cortisol increased the pregnancy rate (75%) compared with control animals (58%), whereas metyrapone reduced the pregnancy rate to 16.7% (P<0.05). The overall results suggest that cortisol, depending on the physiological status of heifers (pregnant vs. nonpregnant), modulates CL function by influencing P4 secretion. Cortisol may have a positive influence on CL function during early pregnancy, leading to support of embryo implantation and resulting in higher rates of pregnancy in heifers.     

Human chorionic gonadotropin influences ovarian function and concentrations of progesterone in prepubertal Angus heifers

The effects of administering hCG on subsequent ovarian structure dynamics and concentrations of progesterone in prepubertal heifers were evaluated. Forty-seven purebred Angus heifers were assigned randomly to 1 of 3 treatments: 1) 100 μg of injection of GnRH (GnRH; n = 16); 2) 1,000 IU injection of hCG (H1000; n = 16); and 3) 500-IU injection of hCG (H500; n = 15). From d -1 to 9 relative to treatment (d 0), daily blood samples were collected to determine concentrations of progesterone and ovaries of each heifer were examined daily by transrectal ultrasonography. Diameter of all follicles ≥ 4 mm and all luteal structures were mapped. Disappearance of the largest follicle occurred within 2 d in a greater percentage (P < 0.05) of all heifers in the H1000 treatment (87.5%) compared with GnRH heifers (43.8%), whereas H500 heifers (73.7%) were intermediate. A new luteal structure formed after follicle disappearance in a greater (P < 0.05) percentage of all heifers treated with H1000 (87.5%) and H500 (73.7%) heifers compared with that in GnRH-treated heifers (18.8%). The largest follicle present in ovaries of H1000 and H500 heifers was smaller (P < 0.05) than that of GnRH heifers from d 2 to 5. Heifers treated with H1000 (1.72 ± 0.19 ng/mL) had peak concentrations of progesterone on d 6 that were greater (P < 0.05) than H500 heifers (1.34 ± 0.20 ng/mL), which were greater than heifers treated with GnRH (0.31 ± 0.19 ng/mL). The mean volume of luteal tissue was greater (P < 0.05) in H1000 heifers (1.54 ± 0.15 cm(3)) than in H500 heifers (1.15 ± 0.15 cm(3)), which was greater (P < 0.05) than in heifers treated with GnRH (0.23 ± 0.15 cm(3)). We conclude that hCG was more effective than GnRH in its ability to cause disappearance of the largest follicle, increase volume of luteal tissue in the subsequent developing luteal structures, and increase concentrations of progesterone in prepubertal heifers. In addition, hCG seems to be more effective when administered at 1,000 IU than at 500 IU.     

Corpus luteum function following spontaneous or prostaglandin-induced estrus in Brahman cows and heifers

Corpus luteum (CL) function following spontaneous or prostaglandin F2 alpha-induced estrus was studied in 27 Brahman cows and 16 Brahman heifers. Females received one injection of 0, .38, .75 or 2.25 mg alfaprostol (PGF)/100 kg body weight at 12 +/- .1 d post-estrus. Serum progesterone (P4) during CL formation, following the subsequent estrus, increased over time (P less than .001, 1 to 13 d post-estrus) and was influenced by PGF (P less than .06) and a PGF X time interaction (P less than .02). Low serum P4 concentrations were observed during CL formation (d 3, 4, 10, and 12) in cattle that received 2.25 mg PGF. On d 13 post-estrus, CL were removed. No differences in weight, P4 content or number of luteinizing hormone (LH) receptors was evident between CL derived 13 d following spontaneous vs PGF-induced estrus. The CL formed following PGF-induced estrus had fewer large (P less than .002) and small (P less than .09) luteal cells in heifers, a lower (P less than .008) percentage composition of histological type I and II luteal cells in cows and a lower (P less than .0001) in vitro P4 response to LH in both cows and heifers when compared with the CL formed following spontaneous estrus. Cows had heavier (P less than .002) CL with a higher (P less than .05) number of large cells, a higher (P less than .09) percentage composition of histological cell type I and II luteal cells, and a higher (P less than .03) in vitro basal and stimulated P4 secretion. It is concluded that 1) the events associated with artificially shortening the estrous cycle with PGF altered subsequent CL function in Brahman females; 2) cows had heavier CL, which were composed of more steroidogenically functional luteal cells, than heifers and 3) age of animal interacted with PGF to alter subsequent CL function in Brahman females.     

Effects of GnRH or progesterone treatment on day 5 post-AI on plasma progesterone, luteal blood flow and leucocyte counts during the luteal phase in dairy cows

This study was designed to test the effects of progesterone or GnRH treatment on day 5 post-AI on fertility and luteal function in dairy cows and heifers. Five days after AI, 32 animals were randomly assigned to a control, intravaginal progesterone for 14 days progesterone releasing intravaginal device (PRID) or GnRH treatment group. On days 5, 7, 12, 14, 17 and 19 post-AI, each animal underwent colour Doppler ultrasonography of the corpus luteum and blood samples were collected for cell counts and plasma progesterone determination. Through general linear model repeated measures analysis of variance, significant effects were observed of treatment, parity, inseminating bull, reduced vascularization of the CL and pregnancy on plasma progesterone concentrations, whereas mean plasma progesterone and time luteal phase day, and treatment and plasma progesterone concentration on day 5 post-AI were found to, respectively, affect neutrophil and lymphocyte counts throughout the luteal phase. Moreover, two binary logistic regression analyses were performed. Based on the odds ratio, the likelihood of pregnancy by days 26-32 post-AI was 23.4 times higher in animals with high mean progesterone levels throughout the study period, compared with animals with low mean progesterone. The likelihood of reduced CL vascularization was 14 times higher in animals treated with PRID, compared with control and GnRH-treated animals. In conclusion, our results indicate that treatment on day 5 post-AI with PRID reduced subsequent CL vascularization, whereas GnRH treatment increased plasma progesterone concentrations on day 12 post-AI, although an effect was identified of the inseminating bull on plasma progesterone levels. Pregnant animals showed higher mean plasma progesterone concentrations than in nonpregnant ones and heifers higher than in lactating cows, whereas blood cell counts differed depending on the treatment and on the mean plasma progesterone concentration on day 5 post-AI.     

Domperidone can ameliorate deleterious reproductive effects and reduced weight gain associated with fescue toxicosis in heifers

The aim of this study was to evaluate the effect of a dopamine antagonist, domperidone, in nonpregnant, reproductively cycling heifers consuming endophyte-infected (EI) fescue diets. Thirty crossbred heifers (Angus x Holstein or Hereford x Holstein) were assigned to one of three treatment groups (n = 10); endophyte-free (EF) fescue diet, EI fescue diet, or endophyte-infected diet and treated with domperidone (EID). Heifers fed EI diets had decreased weight gains compared with heifers fed EF or EID (P < 0.05) during a 21-d treatment period. Ovarian structures were monitored via transrectal ultrasound to determine follicle size and day of ovulation. Blood plasma samples were collected daily and analyzed for progesterone concentration to determine luteal function. Heifers ingesting EI diets had estrous cycles of shorter duration and lower mid-cycle progesterone concentrations than heifers in the EF or EID treatments (P < 0.05). Ovaries from a subset of heifers in each group (n = 3 per group) were harvested and in vitro secretion of progesterone from luteal tissue extracts was determined. No differences in progesterone concentrations were detected among luteal tissue incubates (P > 0.05). These results suggest that domperidone supplementation of heifers consuming EI fescue may ameliorate certain symptoms of fescue toxicosis.     

Plasma progesterone concentration in beef heifers receiving exogenous glucose, insulin, or bovine somatotropin

Three experiments were conducted to evaluate plasma concentrations of glucose, insulin, IGF-I, and progesterone (P4) in pubertal beef heifers receiving exogenous glucose, insulin, or sometribove zinc. All heifers used had no luteal P4 synthesis but received a controlled internal drug-releasing device containing 1.38 g of P4 to estimate treatment effects on hepatic P4 degradation. In Exp. 1, 8 pubertal, nulliparous Angus × Hereford heifers (initial BW = 442 ± 14 kg; initial age = 656 ± 7 d) were randomly assigned to receive, in a crossover design containing 2 periods of 10 h, intravenous (i.v.) infusions (10 mL) of insulin (1 μg/kg of BW; INS) or saline (0.9%; SAL). Treatments were administered via jugular venipuncture in 7 applications (0.15 μg insulin/kg BW per application) 45 min apart (from 0 to 270 min). Blood samples were collected immediately before each infusion as well as at -120, -60, 330, 390, and 450 min relative to the first infusion. Heifers receiving INS had greater (P < 0.01) plasma insulin, reduced (P ≤ 0.04) plasma glucose and IGF-I, and similar (P = 0.62) plasma P4 concentrations compared with SAL heifers. In Exp. 2, the same heifers were assigned to receive, in a similar experimental design as Exp. 1, i.v. infusions (10 mL) of 1) insulin (1 μg/kg BW) and glucose (0.5 g/kg BW; INS+G) or 2) SAL. Heifers receiving INS+G had greater (P ≤ 0.02) plasma insulin, glucose, and P4 but reduced (P = 0.01) plasma IGF-I concentrations compared with SAL heifers. In Exp. 3, the same heifers were assigned to receive, in a crossover design containing 2 periods of 14 d, subcutaneous (s.c.) injections of 1) 250 mg of sometribove zinc (BST) or 2) SAL. Blood samples were collected 3 h apart (0900, 1200, 1500, and 1800 h) from heifers on d 6, 8, and 10 relative to treatment administration (d 1). Heifers receiving BST had greater (P < 0.01) plasma glucose and IGF-I and similar (P ≥ 0.67) plasma insulin and P4 concentrations compared with SAL heifers. Results from this series of experiments suggested that concurrent increases in glucose and insulin are required to reduce hepatic catabolism and increase plasma concentrations of P4 in bovine females.     

Is nutritional anestrus precipitated by subfunctional corpora lutea in beef cows?

Thirty-four multiparous, lactating, cyclic beef cows which calved in moderate body condition were used to determine effects of restricted nutrition on corpus luteum (CL) development and endocrine status. At 78 d postpartum, six cows were assigned to a control (CON) diet (26.0 Mcal ME), fed to increase bodyweight (BW) and body condition score (BCS), and the remaining 28 cows were fed to lose BW and BCS on a restricted (RES) diet (14.0 Mcal ME). Following a 40-d adjustment period on respective diets, estrous cycles were synchronized and cows bled daily for determination of progesterone (P4), luteinizing hormone (LH) and insulin (INS) beginning at the synchronized estrus. Ultrasonography was used to determine the ovulatory follicle and CL development. Control cows were maintained for one estrous cycle and were ovariectomized on day 11 of their second cycle. Ten cows on restricted diet (RES-C) continued to form a functional CL (P4 > 1.5 ng/ml at day 10 of an estrous cycle) through as many as 5 cycles, after which observations were discontinued. Fourteen cows on restricted diet (RES-A) were ovariectomized on day 11 of a cycle when a CL was identified by ultrasonography, but was subfunctional (P4 < 1.5 ng/ml on day 10 of that cycle). Four additional RES-A cows which had subfunctional CL were not ovariectomized but were bled for an additional 25 d. At ovariectomy, CL and ovarian weights were collected. Luteal tissue was prepared for evaluation of P4 synthesis, LH responsiveness in vitro, and for determination of P4 content and total LH receptors. Bodyweight and BCS increased in CON cows; whereas, RES cows lost BW and BCS (P < .05). In the cycle prior to ovariectomy, serum P4 and LH were not different in 18 RES-A cows which developed subfunctional CL in comparison to CON cows. Four RES-A cows not ovariectomized but bled for an additional 25 d neither exhibited estrus, ovulated, nor had P4 concentrations greater than .3 ng/ml. Serum INS was lower in RES-A cows during the cycle prior to ovariectomy than in CON cows (P < .05). During the 11-d period prior to ovariectomy, mean serum P4 and INS were lower in RES-A cows than in CON cows (P < .05); however, serum LH was not different. Furthermore, CL and ovarian weights, P4 content of CL, secretion of P4 by luteal tissue in response to LH in vitro and LH receptor number were not different between CON and RES-A cows. In conclusion, nutritional anestrus may be preceded by the formation of a CL with lower steroidogenic output in vivo. However, luteal tissue, collected from RES-A cows, did not appear to be subfunctional during in vitro incubation when substrate availability and gonadotropin support were equal between diets.     

Adrenal-gonad interactions in cattle. Corpus luteum function in intact and adrenalectomized heifers

To determine the effect of the hyperadrenal state on corpus luteum (CL) function, we treated intact and adrenalectomized (ADRX) heifers with adrenocorticotropin (ACTH) and hydrocortisone. ACTH treatment of intact heifers by infusion of 1 mg alpha 1-24 ACTH/24 hr from day 2 to 25 of an estrous cycle increased progesterone concentrations in plasma to 4 +/- .52 ng/ml plasma on days 3 to 4. Thereafter, progesterone concentrations declined, indicating severe suppression of CL function as compared to that of control heifers. Similar infusions of ACTH to ADRX heifers did not alter CL function as reflected by progesterone concentrations in jugular plasma. Infusion of ADRX animals with hydrocortisone succinate (100 mg/24 hr) decreased progesterone in plasma during the cycle, with maximum values averaging less than 4 ng/ml, compared to concentrations of 6 to 7 ng/ml in control ADRX heifers. Chronic administration of ACTH or hydrocortisone to heifers caused a decreased progesterone secretion during the luteal phase of the estrous cycle. On the basis of these results, we suggest that stress, as indicated by the hyperadrenal state, could inhibit progesterone secretion by the corpus luteum.     

Changes in bovine luteal progesterone metabolism in response to exogenous prostaglandin F(2alpha)

An experiment was conducted to determine the effect of prostaglandin F2alpha (PGF2alpha) on luteal synthesis of progesterone (P4) and related progestins. Sixteen beef heifers were assigned in equal numbers to four groups in a 2x2 factorial arrangement of treatments. The experiment consisted of two levels of PGF2alpha analog (0 and 500 microg) and two levels of time (4 and 24 h after injection) of corpus luteum collection. All heifers were injected intravenously with saline (2 ml) or PGF2alpha (cloprostenol) on day 8 of the estrous cycle (estrus=day 0). Jugular blood was collected at 0, 1, 2, 3, 4 and 20, 21, 22, 23, and 24 h after injection. Resulting sera were analyzed for P4 by use of radioimmunoassay. Luteal tissue was analyzed by gas chromatography/mass spectrometry for P4, 20beta-hydroxyprogesterone, pregnenolone, and allopregnanolone (3beta-hydroxy-5alpha-pregnan-20-one). Treatment with PGF2alpha reduced serum concentrations of P4 as early as 1 h after injection (P<0.005) and steroid levels remained low over 24 h. Similarly, administration of PGF2alpha caused a decline in luteal P4 (P<0.005), 20beta-hydroxyprogesterone (P<0.10), and pregnenolone (P<0.05). In contrast, treatment with PGF(2alpha) caused an increase in luteal allopregnanolone over time (time x treatment interaction; P<0.05). These data are interpreted to suggest that PGF2alpha promotes conversion of P4 to the metabolite allopregnanolone.     

Characteristics of estrus before and after first insemination and fertility of heifers after synchronized estrus using GnRH,PGF2alpha,and progesterone

Our objectives were to determine fertility of heifers after synchronization of estrus using PGF2alpha, preceded by progesterone (P4), GnRH, or both, and to examine the variability of estrual characteristics in heifers before first and second AI. Dairy (n = 247) and beef (n = 193) heifers were assigned randomly to each of three treatments: 1) 50 microg of GnRH (injected i.m.) administered on d -7 followed by 25 mg of PGF2alpha (i.m.) on d -1 (GnRH + PGF; modified Select Synch protocol); 2) placement of an intravaginal progesterone (P4)-releasing insert on d -7, PGF2alpha on d -1, and insert removal on d 0 (P4+PGF); and 3) 50 microg of GnRH plus a P4 insert on d -7, followed by 25 mg of PGF2alpha on d -1, and insert removal on d 0 (P4+GnRH+PGF). Characteristics of estrus were examined before first AI and before the next eligible AI (18 to 26 d later), including duration of estrus, number of standing events, and total and individual duration of standing events. In addition, all heifers were checked visually at least twice daily for estrus. Blood samples were collected on d -7, -1, and 0 for determination of P4, and pregnancy status was diagnosed by ultrasonography 27 to 34 d after AI. Rates of detected estrus were less (P < 0.05) in dairy than in beef heifers, and greater (P < 0.05) in heifers treated with P4. Pattern of conception and pregnancy rates among treatments differed between beef and dairy heifers (treatment x group interaction; P < 0.05). In dairy heifers, conception and pregnancy rates were greatest with P4+PGF, followed by P4+GnRH+PGF and GnRH+PGF, respectively. The opposite was observed among treatments in beef heifers. Administration of P4 without the preceding injection of GnRH produced the lowest pregnancy rates in beefheifers. Ofthe quantified sexual behavioral characteristics during the synchronized estrus, the number of standing events and total duration of standing events were greater (P < 0.01) than those observed during the next eligible estrus before second AI, whereas duration of estrus was unaffected.     

Influence of inducing luteal regression before a modified controlled internal drug-releasing device treatment on control of follicular development

At the initiation of most controlled internal drug-releasing (CIDR) device protocols, GnRH has been used to induce ovulation and reset follicular waves; however, its ability to initiate a new follicular wave is variable and dependent on stage of the estrous cycle. The objectives of the current studies were to determine 1) if inducing luteal regression before the injection of GnRH at time of insertion of a CIDR resulted in increased control of follicular development, and 2) if removing endogenous progesterone by inducing luteal regression before insertion of the CIDR decreased variation in LH pulse frequency. In Exp. 1 and 2, Angus-cross cycling beef heifers (n = 22 and 38, respectively) were allotted to 1 of 2 treatments: 1) heifers received an injection of PGF(2α) on d -3, an injection of GnRH and insertion of a CIDR on d 0, and a PGF(2α) injection and CIDR removal on d 6 (PG-CIDR) or 2) an injection of GnRH and insertion of a CIDR on d 0 and on d 7 an injection of PGF(2α) and removal of CIDR (Select Synch + CIDR). In Exp. 3, Angus-cross beef heifers (n = 15) were assigned to 1 of 3 treatments: 1) PG-CIDR; 2) PGF(2α) on d -3, GnRH on d 0, and PGF(2α) on d 6 (PG-No CIDR); or 3) Select Synch + CIDR. Follicular development and ovulatory response were determined by transrectal ultrasonography. Across all experiments, more (P = 0.02) heifers treated with PG before GnRH initiated a new follicular wave after the injection of GnRH compared with Select Synch + CIDR-treated heifers. In Exp. 1, after CIDR removal, interval to estrus did not differ (P = 0.18) between treatments; however, the variance for the interval to estrus was reduced (P < 0.01) in PG-CIDR heifers compared with Select Synch + CIDR heifers. In Exp. 3, there was a tendency (P = 0.09) for LH pulse frequency to be greater among PG-CIDR and PG-No CIDR compared with the Select Synch + CIDR, but area under the curve, mean LH concentrations, and mean amplitude did not differ (P > 0.76). In summary, induction of luteal regression before an injection of GnRH increased the percentage of heifers initiating a new follicular wave. Removal of endogenous progesterone tended to increase LH pulse frequency, and the modified treatment increased the synchrony of estrus after CIDR removal.     

Fixed-time artificial insemination in replacement beef heifers after estrus synchronization with human chorionic gonadotropin or gonadotropin-releasing hormone

We determined the effects of hCG on ovarian response, concentration of progesterone, and fertility in a fixed-time AI (TAI) protocol. Four hundred forty-four crossbred beef heifers were synchronized with the CO-Synch + CIDR (controlled internal drug-releasing insert) protocol. In addition, heifers were randomly assigned to 1 of 4 treatments in a 2 × 2 factorial arrangement of treatments with main factors being 1) pretreatment, no treatment (control), or treatment with 1,000 IU of hCG 14 d before the initiation of the CO-Synch + CIDR protocol and 2) treatment, administration of 1,000 IU of hCG or 100 μg of GnRH at CIDR insertion of the CO-Synch + CIDR protocol. Blood samples were collected from all heifers on d -21, -14, -7, 0, and 2 relative to PGF(2α) injection. Transrectal ultrasonography was used to examine ovaries in a subset of heifers (n = 362) on d -7 and 0 relative to PGF(2α), and to determine pregnancy status of all heifers on d 33 and 82 relative to AI. Pregnancy rates were similar for heifers pretreated with control (33.0%) or hCG (36.4%), whereas pregnancy rates were greater (P < 0.01) for heifers treated with GnRH (40.1%) compared with hCG (29.0%) at CIDR insertion. Heifers pretreated with hCG had more (P < 0.01) corpora lutea present on the day of CIDR insertion and the day of CIDR removal compared with untreated heifers. A greater proportion (P < 0.01) of heifers ovulated as a result of administration of hCG at the time of CIDR insertion (59.0%) compared with GnRH (38.7%). Heifers treated with hCG at CIDR insertion had greater (P < 0.01) concentrations of progesterone compared with those receiving GnRH at the time of CIDR removal (2.42 ± 0.13 vs. 1.74 ± 0.13 ng/mL; P < 0.01) and at fixed-time AI (0.52 ± 0.03 vs. 0.39 ± 0.03 ng/mL; P < 0.01). Therefore, hCG was more effective than GnRH in its ability to ovulate follicles and to increase concentrations of progesterone in beef heifers. Presynchronization with hCG 14 d before CIDR insertion did not alter pregnancy rates, whereas replacing GnRH with hCG at CIDR insertion decreased pregnancy rates.     

Pulsatile or continuous infusion of luteinizing hormone-releasing hormone and hormonal concentrations in prepubertal beef heifers

An experiment was conducted to determine if exogenous luteinizing hormone-releasing hormone (LHRH) administered iv intermittently as pulses (P) or by continuous sc infusion (I) using osmotic minipumps could sustain pulsatile LH release and induce estrous cyclicity in prepubertal heifers. Prepubertal heifers were assigned randomly to: 1) receive pulses of LHRH (n = 6; 2.5 micrograms LHRH/2 h for 72 h), 2) be infused with LHRH (n = 11; 1.25 micrograms LHRH/h for 72 h), or 3) serve as controls (n = 16). Blood was collected at 20-min intervals for 8 h (0900 to 1700 h) from six heifers in each group on d 1, 2, 3 (during treatment), and on d 4 (during 8 h after terminating LHRH treatments). Heifers given LHRH had higher (P less than .01) LH concentrations than controls. Preovulatory-like LH surges occurred in three I, two P and no control heifers during treatment. Pulse frequencies of LH (no. LH pulses/8 h) were greater (P less than .001) for P heifers than for I and control heifers due to pulsatile LHRH treatment. Serum estradiol was higher (P less than .01) during treatment for LHRH-treated heifers than for controls. Serum follicle-stimulating hormone, cortisol, and progesterone were unchanged during treatment. High levels of cortisol on d 1 declined (P less than .001) to baseline by d 2. Characteristic progesterone rises or short luteal phases occurred within 10 d of treatment initiation in more (P less than .05) LHRH-treated heifers (I = 45%, P = 33%) than controls (6%), although days to first observed estrus and first ovulation were unaffected by treatments. Although both continuous and pulsatile administration of LHRH successfully induced LH and estradiol release as well as preovulatory-like LH surges in some heifers, earlier initiation of estrous cycles was not achieved. Estrous cycles appeared to be delayed by exposure to continuous LHRH infusions during the peripubertal period.     

Effects of melatonin treatment on the attainment of puberty in heifers.

This experiment determined the effect of treatment of 3- to 4-mo-old heifer calves with exogenous melatonin, in early summer, on the attainment of puberty the following spring. Hereford heifers, born between February 22, and March 16, were paired by age and BW and assigned at random within pair to receive an ear implant of melatonin (MEL, n = 12) or to serve as controls (C, n = 12). The implant period was 5 wk, starting June 10 at a mean age of 105 +/- 5 d and 134.4 +/- 3.5 kg BW. Puberty was evaluated the following year on four occasions: January 26, March 19, May 2, and June 1. One each date, rectal palpation of the ovaries was performed, blood samples were taken to assess concentrations of progesterone (P4), and BW was determined. Puberty was considered to have occurred when concentrations of P4 were greater than or equal to .5 ng/mL concomitant with a palpable corpus luteum (CL), or when concentrations of P4 were greater than or equal to 1 ng/mL and a CL was present on the next date. At implant removal, concentrations of melatonin in plasma collected at 1000 to 1400 were 69.5 +/- 6.3 and 16.7 +/- 2.4 pg/mL for MEL and C heifers, respectively. On that and subsequent occasions, neither BW nor ADG differed between MEL and C heifers. Treatment with melatonin increased the incidence of heifers pubertal by the second observation date (P less than .05).(ABSTRACT TRUNCATED AT 250 WORDS)     

The induction of a delayed post-ovulatory progesterone rise in dairy cows: a novel model

A delayed rise in post-ovulatory progesterone is associated with poor embryo development in the cow, although the underlying cause of this aberrant luteal function is poorly understood. The objective of this study was to develop a novel model, in which a delayed progesterone rise could be induced by manipulating the dynamics of the follicular phase. Luteolysis was induced in 20 dairy cows in the presence of either a larger follicle > 10 mm (LF, n = 11) or a smaller follicle < 10 mm (SF, n = 9) and transrectal ultrasonography was performed to determine follicle and CL growth and timing of ovulation. Plasma progesterone and oestradiol were analysed 3x daily. Cows were slaughtered on either day 4 (n = 4 per group) or day 7 (SF, n = 5; LF, n = 7) after ovulation. The pre-ovulatory follicle was larger in the LF group than the SF group at luteolysis (13.5 +/- 0.4 mm versus 6.7 +/- 0.7 mm, P < 0.001) and ovulation (16.7 +/- 0.3 mm versus 13.6 +/- 0.6 mm, P < 0.001). The LF group experienced a shorter follicular phase and ovulated 36 h earlier than the SF group (P < 0.001). At luteolysis, plasma oestradiol concentrations were greater in the LF group (P < 0.001), although peak concentrations were not different (P > 0.05). Moreover, higher progesterone concentrations were observed in the LF group during the early luteal phase (P < 0.05). Luteal weights were positively correlated with plasma progesterone concentrations on day 5 (P < 0.05) but not day 8. In conclusion, a model has been developed which has shown that the dynamics of follicle development during the pre-ovulatory period is an important determinant of subsequent CL development and function.     

The effect of repeated follicular puncture on ovarian function in dairy heifers

Three dairy heifers were examined during three consecutive oestrous cycles (control period, CP). Subsequently, the animals were subjected to 4 and then 5 weeks of twice-weekly ovum pick-up (OPU) (FPP1 and FPP2, respectively) with a recovery period (RP) of two consecutive oestrous cycles between FPP1 and FPP2. After FPP2, the animals were slaughtered and the ovaries were macroscopically examined. Throughout, ovarian activity was monitored by transrectal ultrasonography and concentrations of plasma progesterone. During CP, all the heifers showed normal cyclicity. During FPPs, the heifers occasionally presented oestrous activity. Corpus luteum (CL)-like structures developed from punctured follicles with diameters and life-spans varying from smaller and shorter than those in the CP (P > 0.05) to equal to those in the CP. There was a tendency for a lower number of emerging and punctured follicles in the presence of a CL-like structure. Subsequently to FPP1, all heifers regained normal cyclicity. A thickening of the ovarian tunica albuginea and a slight hardening of the ovaries were found postmortem. In conclusion, dairy heifers can occasionally show cyclic activity and form CL-like structures during twice-weekly OPU. Further, OPU did not seem to cause any major negative effects on ovarian structure and subsequent ovarian function.     

Nonpuberal estrus in beef heifers

The frequency of occurrence of behavioral estrus without subsequent development of functional luteal tissue (termed nonpuberal estrus, NPE), was determined in 43 Simmental X Hereford-Brahman heifers. Blood samples were collected weekly from the start of the study to first behavioral estrus and then daily from d 1 (d 0 = estrus) through d 14 following first and subsequently observed estrous behaviors. All blood samples were analyzed for serum progesterone (P4) concentrations by radioimmunoassay. More heifers (62.8%) exhibited NPE than had luteal development after their first behavioral estrus (37.2%). There was a tendency for fewer light-weight heifers (less than or equal to 240 kg at the start of the experiment) to exhibit a puberal first estrus compared with the heavy-weight (greater than 240 kg at the start of the experiment) heifers (31.2% vs 68.8%, respectively; P = .12). Heifers that had a puberal first estrus were older (376 +/- 12 d vs 334 +/- 9 d, P less than .05) compared with heifers that had NPE. Weight at first behavioral estrus was similar between heifers that had a puberal first estrus and those that had NPE (298 +/- 8 kg and 289 +/- 6 kg, respectively). More heifers that had a puberal first estrus also had an elevation in serum P4 concentrations before that first estrus (64.3% vs 20.0%, P less than .05), and the serum P4 elevation was greater (2.5 +/- .4 ng vs 1.2 +/- .1 ng, P less than .05) than heifers that had NPE. We have concluded from these results that NPE is a common occurrence in heifers approaching puberty.(ABSTRACT TRUNCATED AT 250 WORDS)     

Uterine influences on the formation of subnormal corpora lutea in seasonally anestrous ewes

The hypothesis that subnormal luteal function after induced ovulation in anestrous ewes was the result of uterine influences exerted during the periovulatory period was tested. Crossbred ewes (n = 27) in seasonal anestrus were induced to ovulate by administration of 12 doses of 250 ng of LHRH at 2-h intervals, followed immediately by a bolus injection of LHRH (250 micrograms; d 0). Ewes were unilaterally hysterectomized on either d -3 (PRELHRH) or 2 (POSTLHRH). Daily blood samples were collected and assayed for progesterone (P4) and 13,14-dihydro-15-keto-prostaglandin F2 alpha (PGFM). All ewes were slaughtered on d 10, and corpora lutea (CL) were collected, weighed, and assayed for concentration of P4. All ewes that ovulated exclusively in the ovary ipsilateral to the remaining uterine horn had a transient increase in plasma P4 of 2 to 3 d (short luteal phase). In ewes with at least one CL in the isolated ovary, elevated plasma P4 was maintained after hysterectomy but was consistently lower (P less than .05) in POSTLHRH ewes than in PRELHRH ewes. Concentrations of PGFM did not differ between treatments. The CL ipsilateral to the remaining uterine horn weighted less (P less than .01) and contained less P4 (P less than .01) than contralateral CL. These data confirm the hypothesis that premature regression of subnormal CL is uterine-dependent in a local fashion. Presence of the uterus during the follicular and(or) early luteal phase inhibited subsequent luteal function in seasonally anestrous ewes.     

Progesterone and Caspase-3 Activation in Equine Cyclic Corpora Lutea

Soon after ovulation, the newly formed corpus luteum (CL) starts secreting progesterone (P(4)), necessary for implantation. The CL, an ovarian transient endocrine organ, undergoes growth and regression throughout its life span. The objective of this study was to evaluate if caspase-3 mediates cell death in the equine cyclic luteal structures and relate it to luteal endocrine function. Blood and luteal tissue were collected during the breeding season after slaughter from 38 randomly assigned cycling mares. Luteal tissues were classified as corpora haemorrhagica (CH; n = 7); mid luteal phase corpora lutea (Mid-CL; n = 17); late or regressing corpora lutea (Late-CL; n = 9) and corpora albicans (CA; n = 5). Plasma P(4) concentration, determined by radioimmunoassay, showed a significant increase from CH to Mid-CL (p < 0.001), followed by a decrease to Late-CL (p < 0.001) and CA (p < 0.001). Caspase-3 processing and poly (ADP) ribose polymerase (PARP) degradation were assessed by western blotting. Active caspase-3 was twofold increased in Mid-CL, Late-CL and CA as compared with CH (p < 0.05). Immunocytochemistry also showed a significant increase in caspase-3 expression in large luteal cells in all structures when compared with CH (p < 0.05). Consistently, the endogenous caspase-3 substrate, PARP, was markedly degraded from CH to CA (p < 0.05). In fact, the ratio of full-length to degraded PARP showed a significant decrease from CH to Mid-CL, Late-CL and CA (p < 0.05). Finally, the decrease in P(4) from Mid- to Late-CL coincided with no further increases in apoptosis. In conclusion, these results suggest that the effector caspase-3 of apoptosis, might play an important role during luteal tissue involution in the mare, even though its relationship with P(4) remains to be elucidated.