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Cryopreservation of rooster semen is a reproductive technology carried out to boost genetic gain and productivity in commercial flocks of chicken. However, semen freezing significantly reduces the quality and fertilizing potential of spermatozoa. This study examined cryoprotective effects of the mitochondria-targeted antioxidant mitoquinol mesylate added to the freezing extender by assessing post-thaw characteristics of rooster sperm. Semen samples were diluted in the Beltsville extender supplemented with 0, 1, 10, 100 or 1000 nM of mitoquinol mesylate. Following the thawing of cryopreserved semen doses, we evaluated the following sperm parameters: motility and morphology, membrane integrity and mitochondrial function, acrosome integrity, apoptosis status, lipid peroxidation, DNA fragmentation, reactive oxygen species (ROS) accumulation, epigenetic patterns (DNA methylation and histone modifications), and fertilizing ability. Semen diluted in extenders containing 10 or 100 nM of mitoquinol mesylate significantly exceeded all other groups of frozen-thawed semen samples in sperm motility, average path velocity as well as membrane/acrosome integrity, mitochondrial function indices and DNA/histone modification. Moreover, the addition of 10 and 100 nM of mitoquinol mesylate significantly reduced lipid peroxidation, apoptosis rate, DNA fragmentation and ROS concentrations compared with all other dilutions and was associated with a higher (P ≤ 0.05) fertilization rate compared with a non-supplemented control group, albeit it was still significantly lower compared with that obtained using fresh semen. It can be, however, concluded that mitoquinol mesylate significantly improved an array of quality parameters and fertilizing potential of rooster semen, and hence can be recommended for use as a diluent additive in semen cryopreservation procedures employed in commercial poultry operations., Competing Interests: Declaration of competing interest The authors have no conflicts of interest to disclose., (Copyright © 2025 Elsevier Inc. All rights reserved.)

Artificial insemination (AI) and in vivo embryo production (or multiple ovulation and embryo transfer, MOET) programs are both instrumental in accelerating the propagation of genetically and economically superior goats and sheep. The aim of this review was to present the current gestalt of non-surgical AI and embryo recovery (NSER) procedures in small ruminants. Small body size, precluding rectal palpation, and highly limited penetrability of the uterine cervix in ewes are the major reasons for the scarce use of non-surgical assisted reproduction techniques in this species. As a result, AI and embryo recovery techniques in sheep mainly involve laparoscopy or laparotomy (LAP). In does, however, the Embrapa method of AI allows for successful intrauterine deposition of semen, resulting in pregnancy rates from 50 to 80% under field conditions (>3 000 goats inseminated) when frozen-thawed semen is used. After the administration of prostaglandin F 2α (PGF 2α ), non-surgical (transcervical) embryo recovery is also feasible in goats, with the cervical penetration rate approaching 100%. There is a paucity of information on the efficacy of non-surgical AI using frozen semen in sheep, but the results are satisfactory with fresh, cooled, or chilled ram semen. An application of the NSER technique in ewes has greatly improved over the last decade, and cervical penetration rates of ∼90% can be achieved when a hormonal cervical dilation protocol using PGF 2α , oxytocin, and/or estradiol ester (e.g., estradiol benzoate) is applied. In some genotypes of sheep, sufficient cervical dilation can be induced without estradiol ester included in the protocol. Several studies indicated that recovery of transferable quality ovine embryos using NSER is comparable to that employing a ventral midline laparotomy, and NSER is evidently a method of choice when animal welfare is concerned. Considering both the number of retrievable embryos and animal well-being, the NSER is a viable alternative for surgical procedures. With further developments, it has the makings of a primary, if not exclusive, embryo recovery technique in small ruminants worldwide., (Copyright © 2023 The Author(s). Published by Elsevier B.V. All rights reserved.)

The present experiment employed time-lapse (TL) imaging to assess the effects of vitrification on the development of ovine blastocysts and to see if the timing of blastocyst formation and expansion was correlated with the numbers of embryo- and trophoblasts determined through differential staining of the expanded blastocysts. Ovaries were obtained after slaughter from cycling (October-March) Polish Longwool ewes aged 1-3 years and cumulus-oocyte complexes were collected by ovarian scarification. In vitro maturation was performed in TCM 199 medium supplemented with Earle's Salt, 10% of FBS, and 5 μg/mL of LH/FSH at 38 °C for 24 h. After maturation, the oocytes were incubated with thawed ram semen (IVF) for 19 h and all presumptive zygotes were transferred to a 16-well dish containing Cult medium for monitoring with the Primo Vision TL system. A portion of ovine embryos were vitrified (Cryotop system) at the early cavitation stage and TL observations of warmed (n = 30) and non-vitrified (n = 32) blastocysts continued until the attainment of the expanded blastocyst stage, at which point they were differentially stained with bisbenzimide and propidium iodide for microscopic enumeration of embryoblasts (inner cell mass blastomeres) and trophoblast-cells. There were no statistically significant differences in the timing of blastulation (tB) and formation of expanded blastocysts (tEB) between vitrified and non-vitrified ovine embryos, but non-vitrified blastocysts exceeded (P < 0.05) their vitrified and warmed counterparts in the mean number of embryo- and trophoblasts. In addition, the number of trophoblasts was negatively and moderately correlated with tB and tEB, for both vitrified and non-vitrified embryos. It can be concluded that even though vitrification of ovine embryos is associated with a significant reduction in the number of blastomeres, the rate of blastocyst development remains closely linked to the numbers of trophoblastic cells., (Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.)

Economic potential of the swine industry hinges upon the reproductive performance of sows, which may be enhanced by improving uterine capacity, a component trait of litter size and piglet productivity. Previous attempts at characterizing morphological traits indicative of high uterine volume have not been completely successful, resulting in the continued need for a reliable method of predicting reproductive value to improve production efficiency of the sow. Hence, the main objective of this study was to scrutinize macro- and micro-morphology of the sow's reproductive tract for quantitative correlations with fertility indices. Reproductive records from Polish Landrace × Polish Large White sows were used to examine the associations between fertility and ovarian/uterine morphology (n = 34) or uterine histomorphometry (n = 10). Several measures related to the ovary, including right and left ovarian weight (r = 0.50, p = 0.005 and r = 0.49, p = 0.006, respectively), were positively correlated with the litter size, while left ovarian number of corpora lutea (r = -0.38, p = 0.04) was negatively correlated with the mean litter size. Analysis of histomorphological characteristics of the uterine wall collected during the luteal phase of the estrous cycle revealed correlations between mean litter size and myometrial vascular content (r = 0.75, p = 0.03), the proportion of myometrial stroma (r = -0.68, p = 0.03), and the variability of endometrial thickness (r = -0.72, p = 0.02) in sows. Eight ovarian, vaginal and uterine characteristics were significantly correlated with mean lifetime numbers of live born and stillborn piglets/litter or the last litter size before slaughter. In conclusion, several anatomical and histomorphological metrics that relate to reproductive performance of swine may be used to inform production protocols and as a tool for selection of elite breeding sows, warranting future research into non-invasive or minimally invasive techniques for obtaining such measures., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Selection of appropriate housing conditions for sows is critical for their physical health and long-term reproductive success. The present objective was to evaluate the influences of housing system postweaning (i.e., individual stalls (IS) or group pens (GP)), season and parity on piglet productivity of sows in a commercial setting. This study utilized 3 053 Polish Large White×Polish Landrace sows that were weaned at a rate of 20-30 animals per week at the median age of 4 weeks; 1 474 sows were moved into GP of seven to eight animals each, while 1 579 were placed in IS after weaning. Starting 2 days postweaning all animals were checked for estrus with a teaser boar and then artificially inseminated using 3×10 9 spermatozoa per dose of an inseminate at the onset of heat and 24 h later. The proportion of sows showing the signs of standing heat at or before 6 days postweaning was greater (P < 0.05) for sows moved to GP compared with IS; this difference manifested mainly in second parity sows weaned in the summer and fall. Conception and farrowing rates were significantly higher (P < 0.01) and the weaning-to-estrus interval shorter in GP compared with IS sows in every season but autumn. Mean litter size was lower (P < 0.05) in IS groups in summer, autumn, and winter, and the number of live-born piglets/sow was lower (P < 0.05) for IS sows in the summer and fall. Beneficial effects of group housing on piglet productivity manifested up until the seventh consecutive farrowing and then began to wane. In summary, there was a significantly greater proportion of sows going estrus "on time" (i.e., <7 days) in group housing compared to single stalls but this effect was confined to the second parity sows during the summer and fall months; these results suggest the existence of a seasonal and age-related aspect to sow fertility worthy of further investigation. While both housing systems have their pros and cons, our present results indicate that, in commercial settings, group housing postweaning improved nearly all reproductive parameters of sows., (Copyright © 2020 The Authors. Published by Elsevier B.V. All rights reserved.)

Time-lapse (TL) imaging provides a practical and safe tool to constantly monitor the development of in vitro-derived embryos. TL may help develop novel methods of predicting the timing of embryo cleavage that will lead to optimizing blastocyst cryopreservation or transfer. The primary objective of the present study was to employ TL imaging to examine associations among the division kinetics of ovine embryos, their quality and rates of development to the blastocyst stage. Oocytes were collected by ovary scarification from 78 Longwool ewes slaughtered in the breeding season (November-March). Cumulus oocyte complexes (COCs) were matured for 24 h in TCM 199 media containing 0.1 IU/mL LH/FSH and 10% FBS. In-vitro fertilization was carried out by co-incubation of semen and COCs for 19 h. Presumptive zygotes were placed in microwells, in droplets of Cult medium (Gynemed, Lensahn, Germany). Digital images of developing embryos were captured every 10 min by Primo Vision TL system (EVO+; Vitrolife, Göteburg, Sweden). The following time intervals were recorded: from IVF to the attainment of two-cell (t2), three-cells (t3) or four-cell (t4) stage, to morula detection (tM), blastulation (tSB) and blastocyst formation (tB). Lastly, the duration of the second cell cycle (cc2; t3-t2) and complete synchronous cell division (s2; t4-t3) were calculated, and the incidence of developmental anomalies noted. Out of 147 embryos selected for TL observations, 55 (37.4%) developed to the blastocyst stage (normally developing embryos, NE) and 92 (62.6%) failed to reach the blastocyst stage (arrested embryos, AE; P < 0.05). Mean t2, tM, s2 and cc2 were all less (P ≤ 0.02) in NE compared with AE. Approximately 61.9% of embryos exhibited developmental anomalies (35.5% in the NE group and 78.2% in the AE group; P < 0.05) and AE exceeded (P < 0.05) NE in the proportion of FRG (blastomeric fragmentation), IRR (blastomeres of irregular size after cleavage), DC (direct cleavage) and MA (multi-morphological aberrations). Of all NE, 63.6% were classified as good quality and 36.4% as poor quality blastocysts (P < 0.05). Good quality ovine blastocysts attained t2, t3, t4, tSB and tB stages earlier (P ≤ 0.03) than poor quality blastocysts and none of the poor quality blastocysts was seen to hatch. To recapitulate, the present results indicate that the kinetics of early ovine embryo development are significant predictors of their potential to develop to the blastocyst stage and the markers of blastocyst quality. Time-lapse imaging may serve as a useful technique for predicting the outcome and enhancing efficacy of in vitro embryo production in sheep., (Copyright © 2020 Elsevier Inc. All rights reserved.)

This study examined the effects of exogenous hCG administration on ovarian function and pregnancy rates in estrous-induced dairy goats during the transition into the breeding season. Eighty-six Toggenburg does received 60 mg of medroxyprogesterone acetate intravaginal sponge for 6 d plus 200 IU of equine chorionic gonadotropin and 30 μg of d-cloprostenol i.m. 24 h before sponge removal, and were then bred for 96 h. Seven days (D7) after first mating the does received either 1 mL of saline (the control group, n = 43) or 300 IU of hCG (the hCG-treated group, n = 43) i.m. Transrectal ovarian ultrasonography (B-mode and color Doppler) was performed on D7, D13, D17, and D21 and ultrasonographic pregnancy detection on D30. Pregnancy rate was higher (P < 0.05) in hCG-treated goats (90.7%; 39/43) than that in control animals (74.4%; 32/43). Accessory luteal structures (ALSs) were detected in 46.5% (20/43) of hCG-treated does. All hCG-treated does that had ALSs and 82.6% of goats without ALS post-treatment remained pregnant. The total luteal area increased (P < 0.05) from D7 to D13 in pregnant animals of both groups, whereas mean vascular area declined (P < 0.05) by D21 in all nonpregnant does. Serum progesterone concentrations increased (P < 0.05) on D21 in pregnant goats of both groups, but they were related to changes in luteal tissue content only in control does throughout the present study. Mean daily numbers of small- and medium-sized antral follicles decreased (P < 0.05) only in pregnant animals of both groups with a decline in medium follicle numbers occurring earlier in hCG-treated (D13) compared with control does (D17). To summarize, a single dose of hCG given on D7 after estrus was followed by a decrease in the number of medium-sized antral follicles in gestating hCG-treated does, induced the formation of ALSs in ~47% of all hCG-treated does, and significantly increased the pregnancy rate in estrous-induced Toggenburg goats in the transition to the breeding season., (Copyright © 2020 Elsevier Inc. All rights reserved.)

The main goal of this study was to examine the utility of measuring systemic concentrations of steroid hormones, namely progesterone (P 4 ) and estrone sulfate (E 1 S), for monitoring the progression of porcine pregnancy and predicting sow fertility. There were 3 subsets of artificially inseminated (AI'd) sows used in the present experiments: (i) animals sacrificed on gestational day 20 (gd20; n = 16) or (ii) gd50 (n = 16; Experiment 1), and (iii) animals maintained throughout pregnancy (n = 24; Experiment 2). Blood samples (10 mL) were drawn from the orbital sinus and the endocrine data determined at different time points around ovulation/artificial insemination (gd0 (first AI), gd1 (second AI), and gd2) and maternal recognition of pregnancy (gd11), as well as on gd20 and gd50 (during 2 periods of increased embryonic/fetal mortality in swine) were examined for correlations with the numbers of healthy, arrested, and reabsorbing embryos (Experiment 1) or with the number of live, stillborn, and mummified piglets recorded at farrowing (Experiment 2). No correlations were recorded between circulating concentrations of both steroids and the numbers of healthy, arresting, or reabsorbing conceptuses on gd20 or 50 (Experiment 1). The number of corpora lutea (CL) was directly related to the number of healthy embryos/conceptuses on gd20 and 50 (r = 0.71, P = 0.007 and r = 0.76, P = 0.0007, respectively) and the number of arresting embryos on gd20 (r = 0.54, P = 0.05), and negatively correlated with the number of reabsorbing embryos on gd20 (r = -0.53, P = 0.05). In Experiment 2, circulating P 4 concentrations on gd11 related directly to the number of live-born piglets (r = 0.46, P < 0.04). Systemic E 1 S concentrations on gd0, gd1, gd2 and gd50 were correlated with the number of mummified conceptuses recorded at farrowing (r = 0.50, P = 0.03; r = 0.59, P = 0.01; r = 0.48, P = 0.04; and r = 0.56, P = 0.01, respectively) and plasma concentrations of E 1 S on gd20 related directly to the number of stillborn piglets (r = 0.60, P = 0.02). In summary, the number of CL on gd20 and 50 is a reliable marker of embryonic/fetal pig status. Measurements of P 4 and E 1 S on gd20 and 50 showed limited diagnostic value (ie, were not indicative of the number of healthy and abnormally developing embryos/fetuses). However, measurements of circulating P 4 and E 1 S concentrations during the periconceptional period and in the early/mid-pregnancy of sows have the makings of a practical method to predict gestational outcomes., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Developmental potential of cryopreserved in vitro matured oocytes is very low in nearly all mammalian species studied to date. Despite relatively high cleavage rates, the vitrified/warmed metaphase II oocytes have a decreased rate of blastocyst formation, which can be attributed to the elevated cytoplasmic lipid content and lipid droplet fragmentation. Secretory products of ampulla oviductal epithelial cells (AECs) at the periovulatory stage of the ovarian cycle enhance the viability of in vitro matured oocytes. The present study was undertaken to determine if co-culture of cumulus-oophorus complexes (COCs) with conspecific AECs or reducing the lipid content of in vitro matured ovine oocytes would improve their cryotolerance and ensuing developmental competence. Ovine COCs aspirated from the slaughterhouse ovaries were matured in the following media or culture conditions: TCM199 + FBS + AECs (T1); TCM199 + FBS (T2); TCM199 + BSA (T3); TCM199 + 0.6 mg/mL of l-carnitine (T4); TCM199+ l-carnitine + FBS (T5), or TCM199 only (Ctr). Subsequently, the oocytes were vitrified and used for in vitro fertilization (IVF). The lowest degree of zona pellucida (ZP) hardening following vitrification of in vitro matured sheep oocytes was observed in T1 and T5 (P < 0.05). Cleavage, blastocyst formation and ensuing development (i.e., total cell numbers) as well as blastocyst hatching rates were all greater (P < 0.05) in T1 compared with the remaining groups; in vitro matured COCs in T4 and Ctr did not develop beyond the cleavage stage. The inner cell mass: trophectoderm cell ratio in T1 (1:3.29) was significantly greater compared with T2 (1:3.39), T3 (1:3.40) and T5 (1:3.44). The present results indicate that the ovine COCs/AECs co-culture system had the most positive influence on cryotolerance, ZP hardening, and developmental competence of in vitro matured oocytes., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Azaperone can reduce stress caused by weaning and relocation of breeding sows, but its effects on reproductive processes are still poorly understood. The primary aim of this study was to describe and compare the endocrine and ovarian activity in ultrasonographically monitored second parity sows, with or without azaperone treatment at weaning (2 mg/kg BW i.m.). The intervals from weaning to the onset of estrus and ovulation were both greater (P<0.05) in azaperone-treated (n=12) than in control sows (n=12) by ~12 h. Mean daily growth rates of identified antral follicles were less (P<0.05) in azaperone-treated than in control sows (1.08±0.17 v.1.23±0.18 mm/day; mean±SD) and treated animals exceeded (P<0.05) controls in the mean ovulation rate (13.7±1.3 v. 12.6±1.2). A transient suppression of cortisol release was observed in the treatment group (at 10 and 30 min after azaperone injections) but circulating cortisol concentrations were variable in both groups of sows for the remainder of the study. The preovulatory rise in LH and estradiol secretion was delayed (P<0.05), and the duration of the LH surge was greater (P<0.001) in azaperone-treated sows compared with their control counterparts. The amplitude of episodic fluctuations in serum cortisol concentrations was correlated with the number of stillborn piglets in control sows (r=0.63, P=0.04). The amplitude and concentration of the preovulatory rise in estradiol secretion were negatively correlated with ovulatory response and litter size (r=-0.63 to -0.82, P<0.05), whereas the time at which the LH surge ended was directly related to the number of live-born piglets (r=0.82, P=0.002) in azaperone-treated animals. The present results indicate that administration of azaperone at weaning had a profound effect on preovulatory LH secretion as well as growth kinetics and estrogenicity of ovarian antral follicles. However, the causative associations among various characteristics of the preovulatory LH discharge, ovarian and adrenal steroid secretion post-weaning, and reproductive variables in sows remain equivocal.

Azaperone treatment can control aggression and decrease stress due to weaning, re-grouping and hierarchical fighting of gilts and sows. However, the effects of this butyrophenone neuroleptic and sedative administered at weaning on pig reproductive function are poorly characterized. In this year-long study, a total of 619 cross-bred sows (Polish Large White×Polish Landrace) kept on a commercial farm received an i.m. injection of azaperone (Stresnil®; 2 mg/kg BW) just before weaning and were artificially inseminated during the ensuing estrus with 3×109 spermatozoa per dose of an inseminate; 1180 sows served as untreated controls. Immediately after weaning, the sows were moved to four pens of seven to nine animals each. A teaser boar was used twice daily to check for estrus and sows were bred at heat detection. Subsequently, all sows stayed in individual stalls until pregnancy testing on day 30 post-artificial insemination and were then re-grouped until farrowing. The proportion of pigs that were in estrus within 6 days post-weaning was significantly lower in azaperone-treated groups of animals than in controls (71.4% v. 84.2%). Overall, the azaperone-treated sows had a significantly longer weaning-to-estrus interval (WEI; 8.7±10.1 v. 6.3±8.1 days; mean±SD) and a significantly larger litter size (LS: 11.8±3.0 v.11.3±3.2; azaperone-treated v. control sows). Treatment of the winter-farrowing sows was associated with increased LS (12.8±2.6 and 11.3±3.1 piglets/sow, respectively; P<0.05) and longer (P<0.05) weaning-to-effective-service intervals (11.7±19.3 and 8.4±12.3 days, respectively) as well as farrowing intervals (155.7±19.7 and 152.2±16.1 days, respectively) compared with untreated controls. In the summer months, significantly longer WEIs (12.1±21.0 v. 8.4±16.9 days) were accompanied by a significant decline in LS only in azaperone-treated sows that were inseminated within 6 days post-weaning (10.8±2.9 v. 11.5±3.3 piglets/sow; azaperone-treated v. controls). Azaperone-treated second parity sows had greater LS (P<0.001) along with prolonged WEIs (P<0.05) in comparison to their respective controls, regardless of the timing of estrus. An application of azaperone at weaning increased the annual piglet productivity of winter-farrowing animals and of second parity sows but depressed it significantly in summer. The extra cost and labor due to delayed onset of estrus may cancel out any reproductive benefits of azaperone treatment.

The acquisition of fertilization ability by oocytes is one of the prerequisites for successful in vitro embryo production. In the present study, we examined the influence of conspecific ampulla oviductal epithelial cells incubated with cumulus-oocyte complexes (COCs) throughout the IVM phase on the developmental competence and maturation-promoting factor (MPF) activity of sheep oocytes. There were six experimental groups in this study, namely four groups with and two groups without oviductal epithelial cells added to IVM media: adult COCs matured in vitro with the ampulla oviductal epithelial cells obtained from adult (AAE; G1) or prepubertal donors (prepubertal sheep ampulla oviductal epithelial cells [PAE]; G4), COCs obtained from prepubertal animals cocultured with AAE (G2) or PAE (G3), and adult (C1) and prepubertal sheep COCs (C2) matured without oviductal epithelial cells. Coincubation of oocytes retrieved from both adult and sexually immature donors with AAE (G1 and G2) resulted in significantly improved rates of metaphase-II (M-II) attainment (G1: 85.1 ± 2.0 and G2: 40.2 ± 1.3) and blastocyst formation (G1: 42.2 ± 1.1 and G2: 21.2 ± 1.0) as well as blastocyst development (total cell count; G1: 130.3 ± 7.8, G2: 70.2 ± 3.5) compared with their respective controls (C1: 94.3 ± 4.1 and C2: 49.7 ± 2.0). Prior to IVM, the activity of MPF was greater (P < 0.05) for oocytes obtained from ewes (G1, G4, and C1) compared with those from ewe lambs (G2, G3, and C2). The greatest increment in MPF activity was recorded in G2 (MPF activity before IVM/MPF activity after IVM = 3.62) followed by C2 and G3 (2.22 and 2.20, respectively), and then all remaining groups of oocytes (C1: 1.89, G1: 1.87, and G4: 1.86). In summary, coincubation with AAE during the 24-hour IVM had a positive impact on ovine oocyte competence and ensuing in vitro embryo production efficiency. A significant increase in MPF activity following IVM of G2 oocytes could be responsible, at least partly, for the improved rate of blastocyst formation after IVF of prepubertal sheep oocytes., (Crown Copyright © 2016. Published by Elsevier Inc. All rights reserved.)

Ovarian antral follicles in the ewe grow in an orderly succession, producing 3 to 4 waves per estrous cycle. In prolific sheep, some large antral follicles from the second-to-last wave of the estrous cycle are added to the ovulatory follicles emerging just before estrus to give a higher ovulation rate; it is feasible that regression of these follicles is prevented by an increase in serum concentrations of FSH or LH pulsatility at proestrus. Prolific sheep tend to have a shorter luteal phase than nonprolific ewes and there is a great deal of evidence that luteal progesterone (P 4 ), in addition to regulating LH release, may govern the secretion of FSH heralding the emergence of follicular waves. The specific purpose of this study was to determine whether or not extending the duration of the luteal phase in prolific sheep to that typically seen in nonprolific breeds would alter the follicle wave dynamics and ovulation rate. In 2 separate experiments, exogenous P 4 (7.5 mg per ewe intramuscularly) was administered on day 11 at PM and day 12 at AM (day 0 = first ovulation of the interovulatory interval studied) in moderately prolific Rideau Arcott × Polled Dorset ewes (experiment 1, n = 8) and highly prolific Olkuska ewes (experiment 2, n = 7; TRT), whereas the equinumerous groups of animals served as controls (CTR). Transrectal ovarian ultrasonography was performed daily, and jugular blood samples were drawn twice a day from day 9 until the next ovulation. Progesterone injections resulted in relatively uniform increments in serum P 4 levels, but the mean duration of the interovulatory interval did not differ (P > 0.05) between TRT and CTR groups of ewes in either experiment. The mean ovulation rate post-treatment was 1.6 ± 0.2 vs 3.2 ± 0.4 (experiment 1, P < 0.001) and 3.2 ± 0.8 vs 4.0 ± 1.0 (experiment 2, P > 0.05) in TRT vs CTR, respectively. The number and percentage of ovulating follicles from the penultimate wave of the interovulatory interval studied was 0.25 ± 0.16 vs 1.75 ± 0.45 (P < 0.01) and 25.0 ± 16.4% vs 75.0 ± 16.4% (P < 0.05) in experiment 1, and 0.50 ± 0.30 vs 1.60 ± 0.40 (P < 0.05) and 13.8 ± 9.0% vs 53.4 ± 16.7% (P < 0.05) in experiment 2, for TRT vs CTR, respectively. In summary, administration of P 4 at the end of diestrus decreased the incidence of ovulations from the penultimate wave of the estrous cycle in both the moderately and highly prolific strains of sheep, but it reduced the ovulation rate only in moderately prolific ewes., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Hormonal ovarian superstimulation has contributed to small ruminant reproduction around the world, impacting genetic improvement and zoosanitary programs, contributing to the conservation of endangered species, and supporting other related biotechnologies. Advanced knowledge surrounding the superovulatory treatments in sheep has resulted in enhanced control of influencing factors and improved the protocols currently used. However, in spite of minimization of some adverse factors, superovulatory responses in ewes still remain variable, preventing the more widespread use of superovulation in commercial embryo transfer programs and reproductive research in this species. Recent evidence demonstrates that changes in antral follicular populations and blood supply, and circulating concentrations of certain reproductive hormones determined at the specific time points just before or during the superovulatory treatment are associated with superovulation success in ewes. This review attempts to compile the data from available literature to identify ovarian and hormonal determinants of the superovulatory outcome in ewes, which can be used to substantially improve the existing protocols and to reduce the extra cost and unnecessary stress imposed on poorly responding animals. An overview of most commonly used and some recently developed, FSH-based ovarian stimulation protocols is given at the outset to highlight variation in the frequency and timing of gonadotropin injections, estrus synchronization methods, and follicular wave synchronization and/or ovulation induction techniques during the superovulatory treatments in ewes., (Copyright © 2016 Elsevier Inc. All rights reserved.)

This study set out to investigate the efficiency of long-term estrus synchronization protocols and ovulatory follicle dynamics in ultrasonographically monitored Santa Inês ewes during lengthening (LD; September-October) and shortening photoperiods (SD; April-May), and the transitional period (TP; January). In addition, the influence of ovarian status (e.g., size of antral follicles and/or presence of corpora lutea) at the outset of the estrus synchronization protocols on the ensuing development of ovulatory follicles was examined. Seventy sexually mature Santa Inês ewes were subjected to one of the two estrus synchronization regimens; on Day 0 (random day of the estrous cycle or anovulatory period), the ewes were fitted with an intravaginal progesterone (P4)-releasing (controlled intrauterine drug release [CIDR]) device, which was left in place for 14 days (G-1CIDR, n = 35) or replaced on Day 7 (G-2CIDR, n = 35), and received an intramuscular injection of 10 mg of PGF2α. The ewes allocated to the G-1CIDR group had mean serum P4 concentrations less than 2 ng/mL during the last 4 days of the synchronization protocol. There were no differences (P > 0.05) in mean ovulation rates between the two protocols tested nor among the ewes varying in ovarian status or studied at different times of the year, but ovulations occurred ∼ 12 hours later in the TP compared with the SD period (P < 0.05). Ovulatory follicles emerged earlier (P < 0.05) in the G-1CIDR group than in the G-2CIDR group (Day 8.3 ± 0.5 vs. 9.2 ± 0.4) and during LD (Day 7.1 ± 0.6) compared with the TP (Day 9.1 ± 0.6) and SD (Day 9.9 ± 0.5 of the protocol). In conclusion, the replacement of CIDR devices prevented the occurrence of lower-than-normal luteal phase levels of P4 at the end of the 14-day estrus synchronization protocol. However, although this procedure and seasonal influences altered certain growth characteristics of ovulatory follicles, there were no effects of these factors on the mean ovulation rate., (Copyright © 2016 Elsevier Inc. All rights reserved.)

The underlying theme of this study involved the evaluation of the dilatory effects of prostaglandin E2 on the ovine cervix and thus the assessment of its potential applicability to transcervical artificial insemination (TCAI) in ewes. A novel method of prostaglandin E2 administration (controlled slow-release vaginal inserts) was examined, and the practical implications of this approach including cervical penetrability and posttreatment pregnancy rates were evaluated. The Guelph method of TCAI was performed during the seasonal anestrus (n = 40) and the breeding season (n = 40) on multiparous Rideau Arcott × Polled Dorset ewes, with or without the pretreatment with Cervidil (for a duration of 12 hours or 24 hours before TCAI). Cervical penetration rates averaged 82.5% (66 of 80), and they varied neither (P > 0.05) between the two seasons nor between Cervidil-treated ewes and their respective controls. Cervidil priming significantly reduced the total time required for TCAI during the breeding season in comparison with controls (54 vs. 98 seconds), especially after the 24-hour exposure (38 vs. 108 seconds). The time taken to traverse the uterine cervix was negatively correlated (P < 0.05) with the breed (percentage of Rideau Arcott genotype) and lifetime lamb production in seasonally anestrous ewes. Four out of 36 (11%) successfully penetrated ewes in the breeding season (three ewes allocated to the 12-hour control group and one ewe that had received Cervidil for 12 hours) became pregnant and carried the lambs to term. Vaginal mucus impedance at TCAI was significantly and positively correlated with the total time required to complete the procedure in cyclic ewes, and the negative correlation between vaginal mucus impedance and total time values at the time of controlled intravaginal drug release device removal approached to significance in anestrous ewes. The present results indicate a moderate benefit of using Cervidil for inducing cervical dilation before TCAI in ewes, mainly in the breeding season. The specific reason(s) for impaired fertility after the TCAI using frozen-thawed ram semen remains to be elucidated., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Follicular wave status at the beginning of exogenous FSH administration is an important contributor to variability in superovulatory responses in ruminants. Studies in ewes have shown a decrease in the number of ovulations when superovulation is initiated in the presence of ostensibly ovulatory-sized ovarian follicles. Hormonal ablation of large antral follicles with the progestin-estradiol (E2-17β) treatment significantly reduces this variability in superovulated anestrous ewes, but the effects of the treatment in cycling ewes have not yet been assessed. Sixteen Rideau Arcott × Polled Dorset ewes (November-December) received either medroxyprogesterone acetate (MAP)-releasing intravaginal sponges (60 mg) or controlled internal drug release (CIDR) devices (containing 300 mg of natural progesterone) for 14 days (Days 0-14), with a single intramuscular injection of 350 μg of E2-17β on Day 6. The superovulatory treatment consisted of six injections of porcine FSH (Folltropin-V) given twice daily, followed by a bolus GnRH injection (50 μg intramuscular) on Day 15. There were no differences (P < 0.05) in the ovulatory responses and embryo yields between the two groups of ewes. In both subsets of animals, the next follicular wave emerged ∼2.5 days after an E2-17β injection (P > 0.05). A decline in maximum follicle size after an E2-17β injection was more abrupt in CIDR- compared with MAP-treated animals, and the ewes pretreated with exogenous progesterone had significantly more 3-mm follicles at the start of the superovulatory treatment. The metabolic clearance rate of exogenous E2-17β appeared to be greater in MAP-treated ewes, but circulating concentrations of porcine FSH failed to increase significantly after each Folltropin-V injection in CIDR-treated animals. The CIDR-treated ewes exceeded (P < 0.05) their MAP-treated counterparts in serum E2-17β concentrations during superovulation. In spite of differences in antral follicle numbers and endocrine profiles between MAP- and CIDR-treated cyclic ewes receiving E2-17β before ovarian superstimulation, there were no differences in superovulatory responses., (Copyright © 2015 Elsevier Inc. All rights reserved.)

The aim of this study was to examine whether leptin (anorexigenic peptide), orexin-A, and ghrelin (orexigenic peptides) could directly (ie, independently of hypothalamic influences) affect the secretion of luteinizing hormone (LH) and growth hormone (GH) by adenohypophyseal (AP) explants obtained from normally fed or fasted (48 h) ewes during the breeding and nonbreeding seasons. In addition, a specific ovine super leptin antagonist (SLAN-3) was used to assess the interactions between leptin and ghrelin and/or orexin-A. Pituitary glands from 16 ovariectomized Polish Longwool ewes that had received estradiol-releasing subcutaneous implants were collected in the breeding (November; n = 8) and nonbreeding (May; n = 8) seasons. The AP explants were incubated for 240 min in a gas-liquid interface and treated with leptin (50 ng/mL), ghrelin (100 ng/mL), orexin-A (100 ng/mL), and SLAN-3 (500 ng/mL) with orexin-A or ghrelin. Treatments with leptin and SLAN-3 + orexin-A increased (P < 0.05) LH concentrations in the cultures of AP explants from fasted animals in the breeding season. Orexin-A increased (P < 0.05) LH secretion by AP explants from both fasted and fed animals in the breeding season. Ghrelin stimulated (P < 0.05) GH secretion by AP explants collected from fasted animals in nonbreeding season and from normally fed ewes in both seasons. Leptin decreased (P < 0.05) GH secretion by AP explants collected from fasted ewes in both seasons and from nonfasted ewes in the breeding season. However, the treatment with SLAN-3 + ghrelin resulted in greater (P < 0.05) GH concentrations compared with leptin treatment of AP explants from fasted ewes in the breeding season and from normally fed ewes in nonbreeding season. In summary, leptin, orexin-A, and ghrelin exerted direct effects on AP secretory function in an ex situ model and both the reproductive season and nutritional status of the animals impinged on the direct effects of the peptides on LH and GH release. Specifically, orexin-A was more potent than leptin in directly stimulating LH secretion in cycling ewes, whereas ghrelin and leptin generally had opposing effects on the secretory function of somatotrophs in sheep., (Copyright © 2014 Elsevier Inc. All rights reserved.)

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

Ultrasonographic images are composed of multiple square picture elements called pixels. Quantitative changes in numerical pixel values (echotexture) determined by computer-assisted analysis of digital images reflect discrete changes in the microscopic structure and physiological status of ovarian antral follicles. The objective of the present study was to determine and compare the ultrasonographic attributes of non-ovulatory antral follicles that grew to an ostensibly ovulatory diameter (> or =5mm) and follicles with different luteal outcomes in response to gonadotropin-releasing hormone (GnRH) in anestrous Western White Face ewes (n=34). All animals received GnRH injections (250ng i.v. every 2h for 24h) followed by a bolus injection of 125microg of GnRH i.v. Ovarian images obtained by repeated transrectal ultrasonography were digitized and subjected to computerized analyses to determine the changes in follicular size and echotexture of the follicular antrum and wall. At the beginning of GnRH treatment, follicles that formed inadequate corpora lutea following ovulation (ICL; n=22) had higher (P<0.001) pixel intensity of the central and peripheral antrum compared with non-ovulatory follicles (n=40). Pixel intensity of the central follicular antrum was greater (P<0.01) in follicles that formed ICL compared with follicles that formed normal (full-lifespan) CL post-treatment (NCL; n=20) and mean pixel heterogeneity of the follicular wall was greater (P<0.05) in non-ovulatory follicles compared with follicles that gave rise to NCL. At the time of GnRH bolus injection (i.e., induction of a synchronous LH surge), the mean diameter of non-ovulatory follicles was greater (P<0.01) than that of all ovulating follicles, and pixel heterogeneity of the central follicular antrum was lowest (P<0.05) in non-ovulatory follicles. The mean diameter of luteinized unovulated follicles (n=9) tended to be greater (P<0.10) at 2.5 and 3 days after emergence, and pixel intensity of the follicular wall was lower (P<0.05) compared with non-luteinized follicles (n=8) at 1.5 and 2.5 days after emergence (beginning of the growth from approximately 3mm onwards). In conclusion, ovarian antral follicles with different outcomes after GnRH treatment (in seasonally anestrous ewes) had distinctive ultrasonographic characteristics.

In the ewe, a rise in circulating concentrations of FSH preceding follicular wave emergence begins in the presence of growing follicles from a previous wave. We hypothesized that prostaglandin F(2alpha) (PGF(2alpha)) given at the time of an endogenous FSH peak in cyclic ewes would result in synchronous ovulation of follicles from two consecutive waves, increasing ovulation rate. Twelve Western White Face (WWF) ewes received a single i.m. injection of PGF(2alpha) (15 mg/ewe) at the expected time of a peak in FSH secretion, from Days 9 to 12 after ovulation. The mean ovulation rate after PGF(2alpha) treatment (2.3+/-0.3) did not differ (P>0.05) from the pre-treatment ovulation rate (1.7+/-0.1). Five ewes ovulated follicles from follicular waves emerging before and after PGF(2alpha) injection (3.0+/-0.6 ovulations/ewe) and seven ewes ovulated follicles only from a wave(s) emerging before PGF(2alpha) treatment (2.0+/-0.3 ovulations/ewe; P>0.05). The mean interval from PGF(2alpha) to emergence of the next follicular wave (1.0+/-0.4 and 4.0+/-0.0 d, respectively; P<0.001) and the interval from PGF(2alpha) treatment to the next FSH peak (0 and 3.5+/-0.4d, respectively; P<0.05) differed between the two groups. Six ewes ovulated after the onset of behavioral estrus, with a mean ovulation rate of 1.7+/-0.2, and six ewes ovulated both before and after the onset of estrus (3.0+/-0.5 ovulations/ewe; P<0.05). None of the ovulations that occurred before estrus resulted in corpora lutea (CL) with a full life span. At 24h before ovulation, follicles ovulating before or after the onset of estrus differed in size (4.1+/-0.3 or 5.5+/-0.4mm, respectively; P<0.05) and had distinctive echotextural characteristics. In conclusion, the administration of PGF(2alpha) at the expected time of an FSH peak at mid-cycle in ewes may alter the endogenous rhythm of FSH secretion and was not consistently followed by ovulation of follicles from two follicular waves. In non-prolific WWF ewes, PGF(2alpha)-induced luteolysis disrupted the normal distribution of the source of ovulatory follicles and may be associated with untimely follicular rupture and luteal inadequacy.

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

The aim of the present study was to investigate the temporal relationship between the secretory pattern of serum LH and FSH concentrations and waves of ovarian antral follicles during the luteal phase of the estrous cycle in sheep. The growth pattern of ovarian antral follicles and CL were monitored by transrectal ultrasonography and gonadotropin concentrations were measured in blood samples collected every 12 min for 6 h/d from 7 to 14 d after ovulation. There were two follicular waves (penultimate and final waves of the cycle) emerging and growing during the period of intensive blood sampling. Mean and basal LH concentrations and LH pulse frequency increased (P < 0.001) with decreasing progesterone concentration at the end of the cycle. Mean and basal FSH concentrations reached a peak (P < 0.01) on the day of follicular wave emergence before declining to a nadir by 2 d after emergence. None of the parameters of pulsatile LH secretion varied significantly with either the emergence of the final follicular wave or with the end of the growth phase of the largest follicle of the penultimate wave of the cycle. However, mean and basal LH concentrations did increase (P < 0.05) after the end of the growth phase of the largest follicle of the final follicular wave of the cycle. Furthermore, the end of the growth phase of the largest follicle of the final wave coincided with functional luteolysis. In summary, there was no abrupt or short-term change in pulsatile LH secretion in association with the emergence or growth of the largest follicle of a wave. We concluded that the emergence and growth of ovarian antral follicles in follicular waves do not require changes in LH secretion, but may involve changes in sensitivity of ovarian follicles to serum LH concentrations.

A transient increase in gonadotropin secretion between 6 and 20 weeks of age is critical for the onset of puberty in bull calves. To try and hasten the onset of puberty, bull calves were treated (s.c.) with 3 mg of bLH (n = 6) or 4 mg of bFSH (n = 6) once every 2 days, from 4 to 8 weeks after birth; control calves received saline (n = 6). At 4 and 8 weeks of age, mean LH concentrations were higher (P < 0.05) in bLH-treated (2.3 +/- 0.04 ng/ml and 1.20 +/- 0.04 ng/ml) as compared to control calves (0.50 +/- 0.1 ng/ml and 0.70 +/- 0.10 ng/ml). Mean serum FSH concentrations at 4 and 8 weeks of age, were higher (P < 0.05) in bFSH-treated (1.60 +/- 0.20 ng/ml and 1.10 +/- 0.2 ng/ml) as compared to control calves (0.38 +/- 0.07 ng/ml and 0.35 +/- 0.07 ng/ml). The age at which scrotal circumference (SC) first reached > or = 28 cm, occurred earlier (P < 0.05) in bFSH-treated calves as compared to saline-treated calves (39.3 +/- 1.3 and 44.8 +/- 1.3 weeks of age, respectively). Based on testicular histology at 56 weeks of age, treatment with bFSH resulted in greater (P < 0.05) numbers of Sertoli cells (5 +/- 0.2, 6 +/- 0.3 and 5 +/- 0.3 in bLH-, bFSH- and saline-treated calves, respectively); elongated spermatids (42 +/- 2, 57 +/- 8 and 38 +/- 5 in bLH-, bFSH- and saline-treated calves, respectively) and spermatocytes (31 +/- 3, 38 +/- 3 and 29 +/- 2 in bLH-, bFSH- and saline-treated calves, respectively) per seminiferous tubule. We concluded that treatment of bull calves with bFSH from 4 to 8 weeks of age increased testicular growth (SC); hastened onset of puberty (SC > or = 28 cm); and enhanced spermatogenesis.

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

Transrectal ovarian ultrasonographic studies have shown that, in cattle, follicular wave emergence is associated with a large increase in the number of small antral follicles (4-6mm in diameter); an analogous association has not been found for small follicles (2-3mm in diameter) in the ewe. In previous studies in ewes, accurate assessment of the number of follicles has been limited to follicles > or =2 or 3mm in size. Newer, high-resolution equipment allowed us to identify follicles > or =0.4mm and to quantify all antral follicles > or =1mm in diameter in seven cyclic Western White Face ewes. This allowed us to expand the small follicle pool examined, from 1 to 4 follicles/day (2-3.5mm in diameter) in earlier studies, to 8-18 follicles/day (1-3mm in diameter). Total number of small follicles (> or =1 and < or =3mm in diameter) increased between Days -1 and 0 (Day 0=day of ovulation), and declined between Days 1 and 3 (P<0.05). There were no significant changes in the number of small or medium (4mm in diameter) follicles around days of follicle wave emergence (+/-2 days). The 1-3 follicles in the 2-3mm size range, which constituted a follicle wave (i.e. grew to > or =5mm in size before regression or ovulation), were the only small follicles to emerge in an orderly succession during the estrous cycle, approximately every 3-5 days. Thus, unlike in cattle, there is no apparent increase in numbers of small follicles at follicle wave emergence in cyclic sheep, and little evidence for selection of recruited follicles and follicular dominance.

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

This was a study that retrospectively analyzed serum gonadotropin secretion and the ultrasonographic appearance of the testis during development in prepubertal bull calves to determine whether there were differences between early and late maturing bulls. Blood samples were taken every other week from 2 wk of age until puberty. Samples were also taken at 12 minute intervals for 12 hours at 4, 10, 20, 25, 30, 35, 40 and 45 wk of age. The GnRH treatment was administered 10 hours after the start of each period of frequent blood sampling. Bull calves fell into two distinctive groups, with one group maturing between 36.6 and 44.2 wk (n = 12) and the other between 46.4 and 48.9 wk of age (n = 8). In samples taken every other week mean serum LH concentrations were greater in early maturing bulls than in late maturing bulls at 12, 14 and 16 wk of age (P<0.05). In blood samples taken every 12 minutes for 10 hours early maturing bull calves had higher mean serum LH concentrations at 4 and 10 wk of age (P<0.05) and higher LH pulse frequency at 10 and 20 wk of age (P<0.05). Mean serum LH concentrations at 4, 10 and 40 wk of age and LH pulse frequency at 10 and 20 wk of age were negatively correlated with age at puberty in bull calves. Mean pixel units of the right and left testis were higher from 34 to 40 wk of age in early maturing than in late maturing animals (P<0.05). It seems possible that hormone measurements and ultrasonographic characteristics of the testes could be developed into powerful tools for studies on the regulation of reproductive development and may aid in the prediction of reproductive potential.

To characterize the pulsatile secretion of LH and FSH and their relationships with various stages of follicular wave development (follicles growing from 3 to > or =5 mm) and formation of corpora lutea (CL), 6 Western white-faced ewes underwent ovarian ultrasonography and intensive blood sampling (every 12 min for 6 h) each day, for 10 and 8 consecutive days, commencing 1 and 2 d after estrus, respectively. Basal serum concentrations of LH and LH pulse frequency declined, whereas LH pulse duration and FSH pulse frequency increased by Day 7 after ovulation (P<0.05). LH pulse amplitude increased (P<0.05) at the end of the growth phase of the largest ovarian follicles in the first follicular wave of the cycle. The amplitude and duration of LH pulses rose (P<0.05) 1 d after CL detection. Mean and basal serum FSH concentrations increased (P<0.05) on the day of emergence of the second follicular wave, and also at the beginning of the static phase of the largest ovarian follicles in the first follicular wave of the cycle. FSH pulse frequency increased (P<0.05) during the growth phase of emergent follicles in the second follicle wave. The detection of CL was associated with a transient decrease in mean and basal serum concentrations of FSH (P<0.05), and it was followed by a transient decline in FSH pulse frequency (P<0.05). These results indicate that LH secretion during the luteal phase of the sheep estrous cycle reflects primarily the stage of development of the CL, and only a rise in LH pulse amplitude may be linked to the end of the growth phase of the largest follicles of waves. Increases in mean and basal serum concentrations of FSH are tightly coupled with the days of follicular wave emergence, and they also coincide with the end of the growth phase of the largest follicles in a previous wave, but FSH pulse frequency increases during the follicle growth phase, especially at mid-cycle.

Transrectal ultrasonography of ovaries was performed in 11 ewes from Days 10 to 26 and on Day 30 of pregnancy to record the number, size, and position of ovarian follicles > or = 3 mm in diameter and corpora lutea (CL). Transrectal and/or transabdominal ultrasonography of the uterus was performed on Days 10 to 26, 30, 35, 40, 45 and 50 of gestation to ascertain the number and position of the conceptuses. In a second experiment, ultrasonography was conducted in 15 ewes on Days 10, 25, 30, 45 and 50 of pregnancy and from Days 13 to 29 after parturition. Ovarian data were classified into ovaries without CL (Group 1), ovaries with the CL in 1 ovary (Group 2), and ovaries with the CL in both ovaries during pregnancy (Group 3). In early pregnant ewes, the total number of follicles and the diameter of all follicles > or = 3 mm were smaller (P < 0.05) in the CL-bearing ovaries (both Group 2, n = 7 and Group 3, n = 8) than in the non-CL-bearing ovaries (Group 1, n = 7), while the largest follicle diameter was significantly smaller in Group 3 than in Group 1 or 2 ovaries. The number of 3-mm follicles in Group 2 ovaries was lower (P < 0.05) than in Group 1 or 3 ovaries, but the mean number of follicles > or = 5 mm in diameter was significantly lower in Group 3 than in Group 1. The total luteal volume per ovary was higher (P < 0.001) in Group 2 than in Group 3 ovaries of early pregnant ewes. The total follicle diameter and the number of follicles growing from 3 to > or = 5 mm in diameter was lower (P < 0.05) for Group 2 ovaries of ewes that carried twins (n = 3) compared with Group 2 ovaries from ewes with singletons (n = 4). There were no differences in follicular dynamics between Group 3 ovaries and the ovaries of Group 2 in ewes that carried twins. No follicles > 3 mm were seen in the ovaries of postpartum ewes that contained CL during gestation, until Days 21 and 25 postpartum for Groups 2 (n = 10) and 3 (n = 8), respectively, and follicles reaching > or = 5 mm in diameter were detected in only 2 ovaries (Group 2), on Days 27 and 28 postpartum, respectively. We conclude that during early pregnancy in ewes there is a suppression of antral follicle growth which appears to be exerted primarily by the developing conceptus but remains confined to CL-bearing ovaries. Residual local inhibition of follicular development extends into the postpartum period.

Development and demise of luteal structures were monitored using daily transrectal ultrasonography in 2 breeds of sheep differing in ovulation rates (nonprolific Western white-faced cross-bred, n = 12 and prolific pure-bred Finn sheep, n = 7), during 1 estrous cycle in the mid-breeding season. Jugular blood samples were collected once a day for radioimmunoassay (RIA) of progesterone. The mean diameter of ovulatory follicles was higher in Western white-faced than in Finn ewes (6.4 +/- 0.2 and 5.3 +/- 0.2 mm, respectively; P < 0.001). The mean volume of luteal structures was higher (P < 0.05) in Western white-faced compared with Finn sheep from Days 5 to 15 of the cycle (Day 0 = day of ovulation). This accounted for the higher (P < 0.05) total luteal volumes recorded in Western white-faced ewes on Day 7 and from Days 11 to 15, despite the higher ovulation rate in Finn ewes (2.7 +/- 0.3 and 1.7 +/- 0.2, respectively; P < 0.05). Mean serum progesterone concentrations were higher (P < 0.05) in Western white-faced than in Finn ewes from Days 4 to 14. Daily total luteal volumes were positively correlated with daily serum progesterone concentrations throughout the cycle in Finn sheep (r > or = 0.40, P < 0.02), and during luteal growth and regression (r > 0.60, P < or = 0.00001) but not during mid-cycle in white-faced ewes (r = 0.16; P = 0.22). During the growth of the corpora lutea (CL), luteal tissue volume increased faster (P < 0.05) than serum progesterone concentrations in both breeds of sheep. During luteolysis, the decrease in luteal volumes parallelled that in serum progesterone concentrations in Finn (P = 0.11) but not in Western white-faced ewes, where luteal volumes decreased more slowly (P = 0.02) in relation to progesterone secretion. Increased ovulation rate in prolific Finn ewes resulted in more but smaller CL, and lower serum progesterone levels compared with nonprolific Western white-faced ewes. We conclude that breed-specific mechanisms exist to control the formation of luteal tissue and progesterone secretion in cyclic ewes differing in prolificacy. The mechanisms may involve ovulation of Graafian follicles at different sizes and inhibitory paracrine effects of CL on co-existing CL.

The objective of this experiment was to assess the relationship between electrical resistance of the vaginal mucosa and serum concentrations of estradiol (E2) and progesterone (P4) during the estrous cycle in ewes. Vaginal impedance was recorded daily using a 2-electrode impedometer in 10 nonprolific Western white-faced and 7 prolific Finn ewes, during the mid-breeding season (October to December). Transrectal ultrasonography of ovaries was performed once a day to confirm ovulation and monitor follicle growth (follicles > or =3 mm in diameter) and development of corpora lutea (CL). Jugular blood samples were collected daily for radioimmunoassay (RIA) of estradiol and progesterone. In all ewes, a decline in vaginal impedance (to <40 ohms) was closely associated with the onset of behavioral estrus. In both breeds of sheep, there was no significant correlation between daily serum concentrations of estradiol and vaginal impedance throughout the estrous cycle. Daily serum concentrations of progesterone and the E2:P4 ratio were correlated with vaginal impedance during the period of luteolysis and follicular phase in both breeds (Western white-faced ewes: r = 0.62, P = 0.0002 and r = -0.56, P = 0.0002; Finn ewes: r = 0.61, P = 0.001 and r = -0.45, P = 0.03, respectively) and early in the cycle (Days 0 to 2, Day 0 = day of ovulation) in white-faced ewes (r = 0.61, P = 0.0003 and r = -0.36, P = 0.052, respectively) but not during the remaining portion of the luteal phase in either breed. In conclusion, vaginal mucous impedance appears to be primarily controlled by progesterone, but it also changes in response to shifts in the E2:P4 ratio when progesterone concentrations are low. Impedometric characteristics of the vaginal mucosa in cyclic ewes are an indicator of serum concentrations of progesterone and E2:P4 ratios during the terminal stage of the estrous cycle.

In spring-born ram lambs, the testes (from 2 wk), prostate and vesicular glands (from 4 wk) were examined by ultrasonography every 2 wk up to 26 wk of age. Image analysis was done (numerical pixel values). Ram lambs were treated with a long acting formulation of a GnRH superagonist (Leuprolide acetate; 1.5 mg/kg) at 3 and 7 wk of age. In blood samples taken every 15 min for 8 h, mean serum LH, LH pulse amplitude, and basal and mean serum FSH concentrations were lower at 5 wk of age, and LH pulse frequency was lower at 15 wk of age in animals given Leuprolide acetate compared with those of the controls. There were no differences (P>0.05) in testis, prostate or vesicular gland development between treated and control animals. Testicular diameter of the left and right testes in transverse and longitudinal planes increased slowly to 8 wk of age, more rapidly to 18 wk of age, then more slowly to 26 wk of age (P<0.05). Numerical pixel values of testicular images decreased from 2 to 8 wk of age, increased to 22 wk of age and then plateaued. Width of the prostate increased from 4 to 26 wk of age, but length and width of the vesicular glands increased slowly to 8 wk of age, more rapidly to 18 wk of age and then plateaued (P<0.05). Numerical pixel values for the prostate declined from 4 to 8 wk and for the vesicular glands, declined from 4 to 10 wk of age; numerical pixel values increased to 12 wk and then decreased to a nadir at 18 wk, followed by a steady increase to 26 wk of age (P<0.05). We concluded that developmental patterns of numerical pixel values of the testes, prostate and vesicular glands in ram lambs reflect stages of development, but treatment with a GnRH superagonist at 3 and 7 weeks of age did not affect growth of testes, vesicular or prostate glands.