Effect of Progesterone Prior to GnRH-PGF2α Treatment on Induction of Oestrus and Pregnancy in Anoestrous Awassi Ewes

An experiment was conducted to examine the effect of progesterone prior to a GnRH‐PGF_2α treatment on oestrus and pregnancy in seasonally anoestrous Awassi ewes. Twenty‐four ewes were randomly assigned to three groups to be pre‐treated with 60 mg medroxyprogesterone acetate sponges (group A), 600 mg progesterone sponges (group B) or blank sponges (group C) for 4 days. All ewes were injected with 100 μg of GnRH 24 h after sponge removal followed, 5 days later, by 20 mg PGF_2α injection. Ewes were exposed to three fertile rams at the time of PGF_2α injection (day 0, 0 h) and were checked for breeding marks at 6‐h intervals for 5 days. Blood samples were collected from all ewes 1 day (day ?10) prior to sponge insertion, at the time of sponge removal (day ?6), 1 day following sponge removal (day ?5, at the time of GnRH injection) and at the time of PGF_2α injection (day 0) for analysis of progesterone. Progesterone concentrations on days ?10 and ?5 were basal and averaged 0.2 ± 0.04 and 0.2 ± 0.2 ng/ml, respectively. Progesterone concentrations on day ?6 were elevated only in group B ewes and were higher (p < 0.0001) than those of groups A and C. Progesterone concentrations on day 0 were higher (p = 0.002) in groups A and B than group C. Oestrous responses occurred only in ewes of groups A and B (p > 0.05). Induced oestrus conception rate was greater (p < 0.01) in group A than groups B and C. Ewes returned to oestrus 17–20 days following day 0 were two of eight, six of eight and three of eight of groups A, B and C, respectively, all of which eventually lambed. The overall lambing rate was 82% in progesterone‐primed ewes compared with only 38% non‐progesterone‐primed ewes (p < 0.05). Progesterone priming apparently sensitizes GnRH‐PGF_2α‐treated seasonally anoestrous ewes and increases their response in oestrus and pregnancy rates.     

Vaginal Histological Changes after Using Intravaginal Sponges for Oestrous Synchronization in Anoestrous Ewes

To characterize the histological and cytological vaginal changes generated by the use of intravaginal sponge (IS) applied in oestrous synchronization treatments in ewes during mid‐non‐breeding season. Thirty‐five multiparous ewes were allocated to three experimental groups according to the moment in which the samples were taken: (i) ewes treated with IS containing 60 mg of medroxyprogesterone acetate for 14 days, sampled the day of IS removal (group ISR; n = 10), (ii) or after sponge removal at time of oestrus or 72 h after removal (group AR; n = 14) and (iii) ewes without sponge treatment that were sampled at the day of IS removal of the other groups (group CG; n = 11). Vaginal biopsies and cytological samples were taken from the anterior vaginal fornix area. The vagina of the CG group had a stratified squamous epithelium with a moderate degree of cellular infiltration with lymphocytes and plasma cells in the lamina propia. Treated ewes (ISR and AR) had epithelial hyperplasia and hypertrophy. ISR ewes had haemorrhage and perivascular infiltrate and an increased number of epithelial cells, neutrophils, macrophages and erythrocytes at IS removal. The use of IS generated histological and cytological alterations in the vaginal wall when used for oestrous synchronization in anoestrous ewes.     

Characterization of Oestrous Induction Response,Oestrous Duration,Fecundity and Fertility in Awassi Ewes During the Non‐breeding Season Utilizing both CIDR and Intravaginal Sponge Treatments

The aim of this study was to investigate characterization of oestrous response, onset of induced oestrus, oestrous duration, fecundity and fertility in Awassi ewes treatment with intravaginal sponges and Controlled Intravaginal Drug Release (CIDR) devices in combination with pregnant mare serum gonadotropin (PMSG) under local environmental conditions during the non‐breeding season. A total of 62 ewes were divided into three groups. Group CIDR (n = 20) was treated with CIDR devices for 12 days and 400 IU PMSG was injected upon removal of the CIDR. For ewes in Group Sponge (SP) (n = 24), 30 mg fluorogestone acetate was administered to the sheep for 12 days and 400 IU PMSG was injected upon withdrawal of the sponge. Group Control (CON) (n = 18) served as a control group and received no treatment. Adult, intact and sexually experienced Awassi rams were introduced to all groups at the time when the intravaginal devices were removed. There were no significant differences in terms of oestrous response (CIDR: 90%, SP: 87.5%), time to onset of oestrus and duration of induced oestrus between the CIDR and SP groups. The oestrous response of treatment groups was significantly greater (p < 0.05) than in the control ewes. There were no significant differences in pregnancy (CIDR: 70%, SP: 70.8%), lambing (CIDR: 85%, SP: 79.2%) and fecundity rates between ewes treated with CIDR and those treated with sponges. However, pregnancy and lambing rates were significantly (p < 0.05) higher in ewes treated with CIDR or sponges when compared with those in the control group. It was concluded that it is possible to induce fertile oestrus, successful pregnancy and lambing with the treatment of either CIDR or intravaginal sponge in combination with PMSG in Awassi ewes during the non‐breeding season.     

Ovarian Responses, Hormonal Profiles and Embryo Yields in Anoestrous Ewes Superovulated with Folltropin®-V after Pretreatment with Medroxyprogesterone Acetate-releasing Vaginal Sponges and a Single Dose of Oestradiol-17β

In ruminants, superovulatory treatments started at the time of follicular wave emergence result in greater and less variable ovulatory responses and embryo yields compared with the treatments begun in the presence of a large growing antral follicle(s) from the previous waves. The progesterone–oestradiol treatment is routinely used for follicular wave synchronization in cattle. The main objective of this study was to characterize the ovarian responses, hormonal profiles and in vivo embryo production in anoestrous Rideau Arcott ewes (May‐June), which were superovulated after pretreatment with medroxyprogesterone acetate (MAP)‐releasing intravaginal sponges and a single dose of oestradiol‐17β (E₂‐17β). Six days after insertion of MAP sponges, eight ewes were given an i.m. injection of 350 μg of E₂‐17β (E₂‐17β‐treated ewes); 10 ewes were given an i.m. injection of vehicle (control ewes). Multiple‐dose Folltropin®‐V treatment, followed by the bolus injection of GnRH (50 μg i.m.), began 6 days after E₂‐17β/vehicle injection. Transrectal ovarian ultrasonography revealed that: (i) the interval between E₂‐17β/vehicle injection and regression of all follicles ≥5 to 3 mm in diameter was shorter (p < 0.01; 2.6 ± 0.4 vs 4.8 ± 0.6 days respectively); and (ii) the interval between injection and emergence of the next follicular wave was longer (p < 0.05; 5.4 ± 0.3 vs 1.2 ± 0.4 days, respectively) in E₂‐17β‐treated than in control ewes. During the 6 days after injection, the mean FSH peak concentration and basal FSH concentration were lower (p < 0.01) in E₂‐17β‐treated ewes. The mean ovulation rate and the number of recovered embryos did not differ (p > 0.05) between the two groups of ewes. However, the number of luteinized unovulated follicles per ewe, and the variability in the number of luteal structures and overall embryo yield were less (p < 0.05) in E₂‐17β‐treated compared with control ewes. In conclusion, the MAP–E₂‐17β pretreatment significantly reduced the variability in ovarian responses and embryo yields, without affecting the embryo production in superovulated anoestrous ewes.     

Changes in antiproteolytic activity in the blood and cervical mucus after superovulation stimulation]

The function of proteins, peptides, proteases and inhibitors of proteases in modulations of regulation mechanisms of gonadotrophins during the development of ovarian folicles has not been fully explained up to now. We can see difference reactions of ewes to superovulation stimulations in oestrous and anoestrous periods as shown by the variation of the antiproteolytic activity of blood plasma and cervical mucus. Trypsin is used as a model for serine protease, and trypsin inhibitory activity (TIA) was measured from the reduced rate of trypsin hydrolysis of the chromogenic substrate N-alpha-tosyl-L-arginine-4-nitroanilide (TAPA, Bartík et al., 1974). Full hydrolytic activity was determined as a change in absorbency at 405 nm = 1.0 after ten-minute incubation at 25 degrees C and pH = 8.1, and inhibition was expressed in percentage of full activity. Statistical analyses were performed by Student's t-test. Twenty-three ewes in anoestrus and twenty-eight ewes in oestrus were included in this experiment. They were of the Slovak Merino breed, two to three years old, with the mean live weight of thirty to forty kg. The ewes were treated with Ageline vaginal sponges (20 mg chlorsuperlutin/sponge) to provide for synchronization of ovarian activity, in the interval of eleven to twelve days. After removal of sponges, the ewes were stimulated for superovulation as shown in Tab. I. Heparinized blood plasma samples were stored at -25 degrees C. Low molecular TIA activities were determined in HClO4--treated blood plasma. Samples of cervical mucus were taken on cotton-wool tampons which were evaluated in 0.2 M Tris-HCl buffer, pH = 8.1. Figs. 1-6 shows TIA changes in blood plasma and cervical mucus on particular days. Fig. 7 shows the mean values of controls (I-initial) after synchronization (A) and after stimulation (S). TIA changes were different in anoestrous and oestrous periods. Differences in TIA changes in blood plasma and cervical mucus in oestrous period suggested certain local regulation mechanisms of synthesis and/or secretion of this activity in the cervix. Some values of TIA were maximum at the time of expected heat or ovulation and may be some of the factor(s) which influenced fertility of females after superovulation stimulations.     

Patterns of Follicular Growth in Superovulated Sheep and Influence on Endocrine and Ovarian Response

Objectives of this study were to characterize patterns of follicular development in sheep superovulated with purified follicle stimulating hormone (FSH) (OVAGEN^TM, ICP, Auckland, New Zealand) and to determine its influence on preovulatory events (onset of the oestrus behaviour and timing of the preovulatory luteinizing hormone surge) and ovarian response (ovulation rate and embryo yield). Number and size of all ≥ 23 mm follicles from the first FSH injection to withdrawal of progestagen sponges was determined by transrectal ultrasonography just prior to every FSH injection in nine Manchega ewes superovulated with eight decreasing doses (ml) (1.5 × 3, 1.25 × 2 and 1 × 3) of OVAGEN injected twice daily from 60 h before to 24 h after the withdrawal of 40 mg fluorogestone acetate sponges. Oestrous detection and jugular blood sampling for LH radioimmunoassay were performed every 3 h from 14 to 53 h after sponge removal and ovulation rate and number of embryos were determined 4 days after progestagen withdrawal. Administration of OVAGEN induced a significant rise (p < 0.0005) in the number of follicles ≥ 4 mm in size because of an increased growth in size of follicles from the first FSH injection to sponge removal, an increase in the number of newly detected follicles from 12 to 36 h of the first FSH dose (p < 0.005) and a decrease in regression rate from 24 h (p < 0.001). The number of follicles 2–3 mm in size at first FSH dose (10.4 ± 1.5) was positively correlated with the number of ≥ 4 mm follicles at 0 h (19.0 ± 2.7, p < 0.01). A higher number of ≥ 4 mm follicles at 0 h was related with an earlier appearance of oestrus (31.5 ± 1.5 h, p = 0.08) and LH surge (45.0 ± 2.3 h, p < 0.005), and a higher ovulation rate (18.2 ± 3.8, p < 0.005). On the other hand, the rate of embryo recovery was decreased in ewes with earlier preovulatory LH peaks (p < 0.005), with a shorter interval between oestrus and LH peak (p < 0.05).     

Serum Levels of Cardiac Markers NT‐proANP and NT‐proBNP in Brachycephalic bitches at Different Gestational Stages

The aim of this study was to determine serum levels of natriuretic peptide precursors (NT‐proANP and NT‐proBNP) during pregnancy in brachycephalic bitches. Fifteen healthy multiparous bitches were selected for this prospective study. Serum levels of NT‐proANP and NT‐proBNP were measured during anoestrous and at 14, 35, 42, 49 and 56 days (2nd, 5th, 6th, 7th and 8th weeks) of pregnancy. Fourteen animals had normal gestations, and one bitch developed single foetus syndrome. The natriuretic peptide levels of this animal were not included in this study; however, it is important to report that its NT‐proANP levels were four times greater than those of normal patients. There was no significant difference (p = 0.072) in NT‐proBNP levels between anoestrous (0.20 ± 0.10 ng/ml) and the different pregnancy weeks (0.27 ± 0.12 ng/ml). There was a positive correlation (p < 0.0001) between NT‐proANP and gestational age, and the levels of this marker increased significantly (p < 0.0001) during the 6th (0.26 ± 0.06 ng/ml), 7th (0.28 ± 0.04 ng/ml) and 8th weeks (0.29 ± 0.05 ng/ml) when compared to anoestrous (0.18 ± 0.02 ng/ml). NT‐proANP serum levels are correlated with gestational development and may be indicative of cardiovascular adaptation in canine brachycephalic pregnancy.     

Prolactin,Androstenedione and IGF1 Serum Concentrations During Induced Follicular Growth by eCG Administration in the Bitch

The oestrus cycle in the domestic bitch, a monoestrous species, differs considerably from that of other veterinary domestic animals species. In the bitch the combined use of eCG and hCG is effective to induce oestrus predictably and safely (Stornelli et al., Theriogenology, 78, 2012 and 1056). Although several studies were done to describe the hormonal changes during the canine oestrus cycle, to our knowledge none was done to describe the hormonal changes during induced follicular growth after the administration of eCG. The aim of this work was to study prolactin (PRL), insulin‐like growth factor (IGF1) and androstenedione (ANDR) serum concentrations during follicular growth induced by a single dose of eCG administered to late anoestrous bitches. PRL and ANDR concentrations were lower before than after eCG TRT (before eCG vs pro‐oestrus, oestrus and dioestrus; 4.3 ± 1.8 ng/ml vs 6.5 ± 1.6 ng/ml, p < 0.05; 0.08 ± 0.2 ng/ml vs 0.42 ± 0.16 ng/ml, p < 0.05). Conversely, IGF1 concentrations were similar before and after eCG TRT (286.0 ng/ml ±32.2, p > 0.53). Additionally, PRL concentrations were similar before oestrus compared to during oestrus and dioestrus (6.9 ± 1.7 ng/ml, p > 0.19). Furthermore, IGF1 concentrations were higher before and during oestrus compared to first day of dioestrus (286.1 ± 29.8vs 200.4 ± 29.2 ng/ml, p < 0.01). On the contrary, ANDR concentrations were lower before and during oestrus compared to first day of diestrum (0.35 ± 0.17 ng/ml and 0.38 ± 0.15 vs 0.68 ± 0.17 ng/ml, p < 0.05). These results show that treatment with a single injection of 50 IU/kg of eCG in late anoestrous bitches successfully induced changes in follicular growth which were paralleled with changes in PRL, IGF1 and ANDR serum concentration similar to those occurring during a normally occurring oestrous cycle. In addition, our results suggest that IGF1 in the bitch could play an important role in ovarian folliculogenesis.     

α-Adrenergic control of gonadotropin secretion in the ewe

The effect of the centrally acting α-adrenoceptor agonist, clonidine, on plasma LH and FSH was studied in oestradiol-primed and unprimed ewes and in oestrous ewes. In unprimed anoestrous ewes, clonidine stimulated LH and FSH release after a lag period of 18 h, and noradrenaline intracarotid injection or i.v. infusions immediately stimulated LH release. In oestradiol-infused anoestrous ewes, clonidine produced either a delay or inhibition of the gonadotrophin surge and noradrenaline i.v. infusion advanced the LH surge. In oestrous ewes treated with clonidine, there was marked delay in the LH surge, but the magnitude of the LH and FSH surges were unaffected. Intravenous administration of α-adrenoceptor blockers, phentolamine and phenoxybenzamine, blocked the oestradiol-induced gondotrophin surge in anoestrous ewes. The effect of phenoxybenzamine on gonadotrophin surge was dose dependent in oestrous ewes. Small doses (4 mg/kg i.v.) of phenoxybenzamine delayed the synchronous LH and FSH surges. There was complete blockade of the LH surge and partial blockade of FSH surges in ewes given phenoxybenzamine (8 mg/kg i.v.) before the expected synchronous gonadotrophin surges. After this experiment, the initial rise of plasma progesterone concentrations did not occur until day 6 of oestrous cycle. Administration of phenoxybenzamine before the expected second FSH surge had no effect on the second FSH surge. Gonadotrophin release induced by gonadotrophin-releasing hormone was attenuated by phenoxybenzamine, but not by clonidine. The results suggest that the LH surge is under α-adrenergic control and the first FSH surge is under partial α-adrenergic control, but the second FSH surge is not under α-adrenergic control. The results also suggest oestradiol modulation of α-adrenergic receptor action.     

Ion,Protein, Phospholipid and Energy Substrate Content of Oviduct Fluid During the Oestrous Cycle of Buffalo (Bubalus bubalis)

The aim of this research was to analyse the composition of oviduct fluid (ODF) in buffalo cows at different oestrous cycle phases to fulfil the requirements of buffalo embryos in vitro. ODF was collected by chronic cannulation from three cows that were synchronized by administering a synthetic prostaglandin. Based on hormonal profiles, the pre‐ovulatory, ovulatory, post‐ovulatory and luteal phases of the oestrous cycle were defined. The volume of ODF produced (ml/24 h) was influenced by the oestrous cycle, with values (mean ± SE) around ovulation (1.0 ± 0.2) greater (p < 0.05) than in both the luteal (0.4 ± 0.1) and the post‐ovulatory phases (0.5 ± 0.1), but not different from the intermediate values in the pre‐ovulatory phase (0.8 ± 0.2). Among cycle phases, no differences were found in sodium, potassium, calcium and magnesium concentrations (130.0 ± 1.1, 5.1 ± 0.3, 2.8 ± 0.1 and 0.59 ± 0.04 mmol/l respectively). Interestingly, the chloride secretion (μm /24 h) was higher (p < 0.05) at ovulation (150.2 ± 16.5) than during both the luteal (73.7 ± 22.0) and the post‐ovulatory phases (63.7 ± 11.2), with intermediate values in the pre‐ovulatory phase (113.4 ± 23.5). Glucose concentration (mmol/l) was higher (p = 0.056) in the pre‐ovulatory phase (0.06 ± 0.02) than in the luteal (0.02 ± 0.01) and post‐ovulatory (0.02 ± 0.01) phases but not different from values in the ovulatory phase (0.04 ± 0.02). Concentrations of pyruvate and lactate among oestrous cycle phases were similar (0.08 ± 0.01 and 1.0 ± 0.1 mmol/l respectively). The total quantity of phospholipids (μmol/24 h) was greater (p < 0.05) at ovulation (0.21 ± 0.02) compared with the luteal, pre‐ovulatory and post‐ovulatory phases of the cycle (0.09 ± 0.02, 0.13 ± 0.02 and 0.09 ± 0.01 respectively). No differences were found in either the protein concentration (1.8 ± 0.3 mg/ml) or the quantity of proteins secreted in 24 h (1.8 ± 0.4 mg) among oestrous cycle phases. In conclusion, this study provides the first characterization of buffalo ODF during the oestrous cycle, showing species‐specific differences that may be useful for developing suitable media for buffalo in vitro embryo production.     

Relationship between Ultrasonographic Assessment of the Corpus Luteum and Plasma Progesterone Concentration during the Oestrous Cycle in Monovular Ewes

Ultrasonographic observations of the corpus luteum (CL) and collection of blood samples for progesterone radioimmunoassay were performed daily during 15 oestrous cycles in Spanish Merino ewes, a consistently monovular breed. Ultrasonographic image of the CL changed during the oestrous cycle, increasing its echogenic pattern from ovulation to luteolysis. The size of the CL and mean progesterone levels were significantly affected by day of cycle (p < 0.05 and p < 0.001, respectively). Both increased their values from day 1 to day 12 (from 49.6 ± 7.4 to 154.6 ± 11.8 mm² and from 0.2 ± 0.0 to 2.8 ± 0.5 ng/ml, respectively) and then declined sharply until day 0 (28.2 ± 5.3 mm² and 0.1 ± 0.0 ng/ml, respectively). There was a significant correlation between CL area and plasma progesterone concentrations during the entire oestrous cycle, taking the developing and regressing phases of the CL separately (p < 0.05). A central cavity was observed in 33.3% of the CL studied. The presence of this cavity had no effect in total luteal‐tissue area of the CL nor on oestrous cycle length or on progesterone concentrations. Likewise, the cavity did not affect the correlations observed between CL size and progesterone levels, CL size and day of cycle and progesterone levels and day of cycle. It is concluded that ultrasonographic assessment of CL area is a reliable method for estimating peripheral plasma progesterone levels, regardless to the presence or absence of a cavity in the CL.     

Shortening the Postpartum Anoestrous Interval in Suckled Crossbred Dual Purpose Cows Using Progestagen Intravaginal Sponges plus eCG and PGF2α

One hundred and twenty‐six suckled crossbred cows (Bos taurus × Bos indicus), with body condition score ≥3 (1–5 point scale), were employed in the present study to evaluate the effectiveness of intravaginal progestin‐releasing sponges (IVS) for shortening anoestrous interval. Fifty‐four cows were assigned to control group. Seventy‐two cows were treated with IVS impregnated with 250 mg of medroxy‐acetate‐progesterone (MAP) as follows: day 0, IVS plus 5 mg of 17β‐E and 50 mg of MAP i.m.; day 6, 500 IU of equine chorionic gonadotropin and 25 mg prostaglandin F_2α i.m.; day 8, IVS withdrawal and day 9, 1 mg 17β‐E i.m. Cows were also grouped according to postpartum days (dpp) at treatment: MAP <70 days (n = 25); control <70 days (n = 22); MAP >70 days (n = 47); control >70 days (n = 32). From IVS removal, cows were detected in oestrus and inseminated. Cows not detected in oestrus were timed artificial insemination 72 h after sponge removal. Treatment effect on oestrous rate (ER), conception rate (CR), pregnancy rate (PR) and treatment to conception intervals (TCI) and calving to conception intervals (CCI) were evaluated. The ER, CR and PR were analysed using proc logistic , while TCI and CCI with proc glm of SAS. The groups MAP <70 days and MAP >70 days showed higher (p < 0.01) ER than control <70 days and control >70 days (84.0% and 76.6% vs 31.8% and 31.3% respectively). The PR was higher (p < 0.01) in MAP <70 days vs control <70 days (64.0% vs 22.7%) and also higher (p < 0.05) in MAP >70 days vs control <70 days (40.4% vs 18.8%). The TCI and CCI were shorter (p < 0.01) in MAP <70 days vs control <70 days (36.0 and 95.8 days; 95.3 and 158.6 days respectively). In conclusion, only cows treated with IVS before 70 dpp had a CCI shorter than 100 days, consequently this treatment shortened postpartum anoestrous interval in crossbred dual purpose cattle.     

The effects of time and dose of pregnant mare serum gonadotropin (PMSG) on reproductive efficiency in hair sheep ewes

The aim of this study was to evaluate the effects of dose and application time of pregnant mare serum gonadotropin (PMSG) on reproductive performance of hair sheep ewes synchronized with fluorogesterone acetate (FGA) under tropical conditions of Northeastern Mexico. Ninety-nine hair ewes (63 Blackbelly and 36 Pelibuey) were treated with intravaginal sponges during 10 days. After insertion of FGA sponges, ewes were divided into four groups, and PMSG was injected intramuscularly at doses of 100, 200, and 400 IU. Relative to FGA sponge removal, PMSG was administrated at −48 h, −24 h, and at sponge removal. PMSG was not administered to the control group. Control ewes had similar (P > 0.05) lambing rate, fertility, and fecundity than those treated with 100 IU of PMSG, but lower (P < 0.05) percentages to these variables than those treated with 200 and 400 IU of PMSG. Time to estrus decreased linearly, and ovulation rate increased quadratically as PMSG dose increased (0 to 400 IU). Administration of PMSG before sponge removal increased (P < 0.01) response to estrus and decreased (P < 0.01) interval to estrus compared with control. Ovulation rate, lambing rate, fertility, and fecundity were not affected (P > 0.05) by administration time of PMSG. Both dose and time of PMSG application did not affect (P > 0.05) pregnancy rate, percentage of single and multiple lambing, and prolificacy. In conclusion, results show that the dose of 400 IU of PMSG administered before sponge withdrawal in an estrus synchronization protocol improved reproductive efficiency of hair sheep ewes.     

A study on the effect of GnRH administration on the ovarian response and laparoscopic intrauterine insemination of Awassi ewes treated with eCG to induce superovulation

The effect of GnRH administration on superovulatory response of ewes treated with equine chorionic gonadotrophin (eCG) in breeding and nonbreeding seasons and the contribution of laparoscopic insemination to the improvement of fertilization and embryo recovery were investigated. Twenty-four nonpregnant Awassi ewes of 3–4 years of age were randomly allocated into two groups (n = 12). Each ewe was treated with a progesterone impregnated intravaginal sponge for 12 days. The following superovulation treatment was used: ewes of group 1 received 1,200 IU of eCG once as an intramuscular injection 48 h prior to sponge withdrawal; ewes of group 2 also received 1,200 IU of eCG once as an intramuscular injection, 48 h prior to sponge withdrawal and after 24 h of sponge removal. Ewes were injected with 80 μg of GnRH. Ewes of groups 1 and 2 were further subdivided into four equal groups (n = 6). Subgroups A and C (superovulated with eCG and eCG plus GnRH, respectively) were mated naturally at least two times with Awassi rams of proven fertility at 8-h intervals. Subgroups B and D (same as A and C) had intrauterine insemination at 44–46 h after sponge removal, under laparoscopic visualization of uterine horns, depositing 1 ml of diluted semen containing 100 × 10⁶ motile sperm in the distal portion of each uterine horn. Ovarian response was assessed by determining the number of corpora lutea by laparoscopy at day 6 after mating. Embryo recovery was performed by using a semi-laparoscopic flushing procedure in both uterine horns. Results of the present study showed that ewes treated in breeding season with eCG plus GnRH has a higher number (P < 0.05) of corpora lutea than eCG alone as 7.33 ± 0.54 and 4.33 ± 0.39, respectively. There was no significant difference in the number of corpora lutea in nonbreeding season when ewes treated with eCG and eCG plus GnRH. The number of unovulated follicles was significantly higher (P < 0.05) in eCG treated ewes than in ewes treated with eCG plus GnRH, both in the breeding and nonbreeding seasons. The number of recovered embryos from ewes treated with eCG plus GnRH and eCG differ significantly (P < 0.05) as 4.32 ± 0.56 and 1.06 ± 0.26, respectively, in the breeding seasons. No significant difference was observed when these hormones used for superovulation in the nonbreeding season. A higher number of unfertilized ova (P < 0.05) was observed in ewes when naturally inseminated than in ewes inseminated using the intrauterine laparoscopic technique. Higher rate of embryo recovery (P < 0.05) was achieved when ewes were inseminated via intrauterine (4.66 ± 0.66) compared with ewes naturally mated (2.16 ± 0.74). The fertilization rate in ewes inseminated intrauterine using laparoscopic techniques and naturally mated were 91.5% and 44.8%, respectively. Fertilization failure in ewes inseminated intrauterine using laparoscopic techniques and naturally mated were 8.4% and 55.2%, respectively. It could be concluded that administration of GnRH 24 h after sponge removal increased ovulation rate of Awassi ewes treated with eCG for superovulation in the breeding season. The use of eCG to induce superovulation in Awassi ewes combined with laparoscopic intrauterine insemination increases the fertilization rate.     

Ultrasonographic Characteristics of Ovulatory Follicles and Associated Endocrine Changes in Cyclic Ewes Treated with Medroxyprogesterone Acetate (MAP)-releasing Intravaginal Sponges and Equine Chorionic Gonadotropin (eCG)

The aim of this study was to assess the ultrasonographic characteristics of ovulatory follicles in cyclic Western White Face ewes (December) that had received intravaginal sponges containing medroxyprogesterone acetate (MAP; 60 mg) for 12 days, with or without an injection of 500 IU of equine chorionic gonadotropin (eCG) at sponge removal. We hypothesized that quantitative echotextural attributes of the follicles in ewes treated only with MAP would differ from those in MAP/eCG-treated ewes, reflecting the increased antral follicular growth and secretory function under eCG influence. Digital images of ovulatory follicles obtained at 0 and 24 h after MAP sponge removal and at 24 h before ovulation in the eCG-treated (five ewes, 13 follicles) and control (six ewes, 9 follicles) animals, were subjected to computerized analyses. The mean diameter of ovulatory follicles increased (p < 0.001) 24 h after eCG treatment. The mean pixel intensity and heterogeneity of the follicular antrum (p < 0.001), as well as mean pixel intensity of the follicular wall and perifollicular ovarian stroma (p < 0.05), were greater in eCG-treated animals compared with control ewes 24 h after sponge removal and at 24 h before ovulation. Mean serum concentrations of oestradiol-17beta tended to increase (p = 0.06) 24 h after eCG treatment and the eCG-treated ewes exceeded (p < 0.05) control animals in progesterone concentrations from days 9-15 after ovulation. Our results support the hypothesis that large antral follicles in eCG-treated ewes exhibit distinctive echotextural characteristics. Follicular image attributes in eCG-treated ewes appear to be indicative of the changes in follicular morphology and secretory activity caused by the administration of the exogenous gonadotropin, which has both FSH- and LH-like activities.     

Supplementation of Slow‐Release Melatonin Improves Recovery of Ovarian Cyclicity and Conception in Summer Anoestrous Buffaloes (Bubalus bubalis)

The role of melatonin as a protective neurohormone against restoring cyclicity in summer anoestrous animals in photoperiod species has gained wider acceptance. This study was designed to uncover the evidence the slow‐release melatonin (MLT) has on initiation of ovarian cyclicity and conception rate (CR) in summer anoestrous buffaloes. Thus, buffaloes diagnosed as summer anoestrous (absence of overt signs of oestrus, concurrent rectal examination and radioimmunoassay for serum progesterone at 10 days interval) were grouped as untreated (Group I, sterilized corn oil, n = 8) and treated (Group II, single subcutaneous injection of MLT @18 mg/50 kg bwt in sterilized corn oil, n = 20). Animals treated and detected in oestrus were artificially inseminated (AI) followed by division into Group III (second dose of MLT on 5th day post‐AI, n = 8) and Group IV (no melatonin administration, n = 10). Blood samples were collected at 4 days interval for estimation of serum MLT, progesterone and oestrogen using radioimmunoassay kit. Mean oestrous induction rate (OIR), oestrous induction interval (OII), interoestrous interval (IOI) and CR were estimated. Compared to control, concentration of melatonin was significantly (p < 0.05) higher in treated group ranging from 14.34 ± 1.72 to 412.31 ± 14.47 pg/ml whereas other two hormones did not show any concentration difference. Melatonin‐administered buffaloes showed significantly (p < 0.05) higher (90%) OIR with OII of 18.06 ± 1.57 days. Results showed improvement in conception rate in buffaloes administered with post‐insemination melatonin. It can be concluded from the study that slow‐release melatonin supplementation restored cyclicity in summer anoestrous animals resulting in improvement in conception rate in buffaloes.     

Response of Corriedale ewes to the "ram effect" after priming with medroxyprogesterone, fluorogestone, or progesterone in the non-breeding season

One hundred eighty-nine Corriedale ewes were used during the non-breeding season to study the "ram effect" stimulus after priming with progestogens. Intravaginal sponges containing either medroxyprogesterone acetate (MAP group, n = 49), fluorogestone acetate (FGA group, n = 49), or progesterone devices (CIDR group, n = 46) were inserted on Day-6 (Day 0 = introduction of the rams). Forty-five ewes were untreated and kept as a control group. On Day 0 the sponges were removed and rams provided with marking harnesses for oestrous detection were placed with the ewes. Onset of estrus was monitored until Day 25, and conception was determined by transrectal ultrasonography. Ewes came into heat during 4 periods: Days 0-3, 5-7, 17-20, and 21-23. The overall number of oestrus ewes were 29%, 53%, 35%, and 50% for the control, MAP, FGA, and CIDR groups, respectively (MAP and CIDR > control, p < 0.05). Control ewes presented oestrus only on Days 17-20 and 21-23. Oestrus in the progestogen-primed ewes was concentrated during Days 0-3 and 17-20, and some ewes came into oestrus on Days 5-7. There were no differences between different primings neither in oestrous response nor in conception rate. The conception rate from matings occurring on Days 0-3 was higher than on those occurring on Days 17-20. We conclude that MAP, FGA, and CIDR is equally effective in improving the response to the ram effect, and the pattern of oestrus in primed ewes was different than previously reported.     

Effect of peripheral concentrations of progesterone on follicular growth and fertility in ewes

The effects of progesterone (P₄) on follicular growth and fertility in ewes were examined. In Experiment 1, 22 ewes received either one or three packets of P₄ (5 g/packed) or an empty packet subcutaneously (sc) from Days 5 to 15 of the estrous cycle (estrus = Day 0). On Day 6, P₄-treated ewes received 12.5 mg of prostaglandin F₂α. Follicles ⩾3 mm in diameter were observed via transrectal ultrasonography daily from Day 4 through estrus, corpora lutea (CL) were observed 5 to 7 d after estrus. Ewes with low (LOW; ⩽1 ng/ml; n = 5), intermediate (MED; > 1 and <2 ng/ml; n = 10), or normal (NOR; ⩾2 ng/ml; n = 7) P₄ in jugular plasma on Days 7 through 15 differed in follicular development. The largest follicle at estrus was larger in ewes with LOW vs. MED and NOR P₄ (7.8 ± 0.3 vs. 6.9 ± 0.2 mm; P < 0.05). Treatments differed in proportions of multiple-ovulating ewes, in which the oldest ovulatory follicle was first observed before Day 10 (LOW: 3 of 3, MED: 6 of 10, NOR: 0 of 5, respectively; P < 0.05). Estradiol was higher early in the treatment period in LOW ewes than in MED and NOR ewes (day × treatment; P < 0.05). In Experiment 2, ewes received 5 mg of P₄ in corn oil (low progesterone [LP]; n = 51) or 2 ml of corn oil (CON; n = 49) sc every 12 hr on Days 6 through 14 of the estrous cycle before mating. LP ewes received 15 mg of prostaglandin F₂α on Day 6. Mean serum P₄ on Days 7 through 15 was 0.6 ± 0.1 ng/ml in LP and 1.9 ± 0.1 ng/ml in CON ewes. Eleven LP and 12 CON ewes were scanned daily from Day 4 through mating, and in all ewes (n = 93), CL were counted 10 d after mating and embryos were counted at 25, 40, and 60 d of gestation. In multiple-ovulating ewes, day of cycle of appearance was earlier for the oldest (Day 6.1 ± 0.8 vs. 10.4 ± 0.8) but not second oldest (Day 11.7 ± 1.0 vs. 12.2 ± 0.9) ovulatory follicles in LP compared with CON ewes. The conception rate was lower in LP (72%) than in CON ewes (98%; P < 0.01). However, numbers of CL 10 d after mating, and in pregnant ewes, numbers of embryos 25 d after mating and lambs born, did not differ with treatment. In summary, low P₄ increased the size of the largest follicles and the age of the oldest ovulatory follicles. Embryos resulting from the ovulation of older and younger follicles in the same ewe did not differ in their ability to survive.     

雌酮主动免疫对美利奴母羊发情产羔和生殖内分泌的影响

20头母羊在配种季节开始前6周和3周,用雌酮免疫原免疫2次(E组),13头不作处理为对照(C组)。免疫不影响母羊正常发情,可使母羊产羔率由115.38％提高到156.25％,发情当天的LH水平显著提高(3.06±0.61对1.93±0.88miu/ml);发情周期黄体期的孕酮水平亦增加,周期12天时差异显著(9.31±3.53对3.71±0.92ng/ml);整个发情周期内E组的睾酮水平显著高于C组,而17β-雌二醇水平两组间无差异。     

Inhibitory Effect of Selenium Supplementation on the Reproductive Performance in Synchronized Merino Sheep at Range Conditions in a Selenium-deficient Area

The effect of selenium (Se) supplementation on the reproductive performance of Merino ewes mated out of the normal breeding season was studied in a 2 (no Se supplementation vs Se supplementation) × 2 (ewes mated at natural oestrus vs synchronized oestrus) factorial design with 50 ewes per treatment combination. Synchronization of oestrus was achieved by intravaginal insertion of 40 mg cronolone sponges for 14 days and administration of 440 IU PMSG at sponge withdrawal. Se supplementation was by the addition to the concentrate with 0.5 mg/kg of Se in the form of selenomethionin, for the 3 months prior to the mating and during gestation. Ewes were kept under range conditions in south‐west Spain, and exposed to Merino rams at a female: male ratio of 4 : 1 from 2 days following sponge withdrawal, or the equivalent time, for 21 days. Se supplementation alone did not improve significantly ewe fertility or lamb birth weight, but there was a positive effect of synchronizing ewes mated out of the breeding season as more of these ewes lambed and produced more lambs than those not synchronized. However, a strong interaction was found between synchronization and Se supplementation, causing a deleterious effect on the reproductive performance of ewes. This negative effect, presumably related to high embrionary mortality caused by Se toxicity, should be taken into account for oestrus synchronizing in Se‐deficient areas. The supplementation with Se, for the prevention of nutritional myodystrophy degeneration in lambs, should be conducted in a fashion which does not generate high levels of Se in the ewes around the mating period.