Corpus Luteum Size Research Articles

Follicle vascularity coordinates corpus luteum blood flow and progesterone production

Colour Doppler ultrasonography was used to compare the ability of preovulatory follicle (POF) blood flow and its dimensions to predict the size, blood flow and progesterone production capability of the subsequent corpus luteum (CL). Cows (n=30) were submitted to a synchronisation protocol. Follicles ≥7mm were measured and follicular wall blood flow evaluated every 12h for approximately 3.5 days until ovulation. After ovulation, cows were scanned daily for 8 days and similar parameters were evaluated for the CL. Blood samples were collected and plasma progesterone concentrations quantified. All parameters were positively correlated. Correlation values ranged from 0.26 to 0.74 on data normalised to ovulation and from 0.31 to 0.74 on data normalised to maximum values. Correlations between calculated ratios of both POF and CL in data normalised to ovulation and to maximum values ranged from moderate (0.57) to strong (0.87). Significant (P<0.0001) linear regression analyses were seen in all comparisons. In conclusion, higher correlations were observed between the dimensions of POF and/or CL and blood flow of both structures, as well as POF and/or CL blood flow with plasma progesterone concentrations of the resultant CL. These findings indicate that follicle vascularity coordinates CL blood flow and progesterone production in synchronised beef cows.

The Accuracy of Transrectal Ultrasonography Diagnostic for Examination of Ovarian Structures in Cattle

The study aimed to determine the accuracy of ovarian follicle and corpus luteum measurements in vivo using ultrasonography diagnostic and in vitro after slicing of the ovaries. A total of 15 cows were examined using transrectal ultrasonography just before being slaughtered. Ultrasonographic examination was performed using 7.5 MHz transrectal tranducer on ovaries, measured dominant follicle and corpus luteum diameters. Follicleappeared echogenic, black with round shape. Corpus luteum, appeared as low echogenicity structure and grayish in color.Ovaries were taken out soon after animals slaughtered, soaked in 10% buffered formalin for 12 hours,then cut in thin slices and measured follicle and corpus luteum diameters. Results of follicle and corpus luteum diameter measurements in vitro and in vivo compared using two sample or paired t-test, correlation and regression analyses. The two methods of follicle and corpus luteum measurements had linear orrelation (follicle measurement, R2 = 0.933 , r = 0.966, n = 15, P &lt; 0.05) (corpus luteum, R2 = 0.912, r = 0.955, n = 15, P &lt; 0.05).Measurements of dominant follicle diameter and corpus luteum size manifested the real size of the two ovarian structures. Transrectal ultrasonography diagnostic on ovaries proved to be having high accuracy and reliability for identification, measurement of ovarian structure diameters, follicle and corpus luteum in cattle.

Follicle and corpus luteum size and vascularity as predictors of fertility at the time of artificial insemination and embryo transfer in beef cattle

Abstract:Two ultrasound based fertility prediction methods were tested prior to embryo transfer (ET) and artificial insemination (AI) in cattle. Female bovines were submitted to estrous synchronization prior to ET and AI. Animals were scanned immediately before ET and AI procedure to target follicle and corpus luteum (CL) size and vascularity. In addition, inseminated animals were also scanned eleven days after insemination to target CL size and vascularity. All data was compared with fertility by using gestational diagnosis 35 days after ovulation. Prior to ET, CL vascularity showed a positive correlation with fertility, and no pregnancy occurred in animals with less than 40% of CL vascularity. Prior to AI and also eleven days after AI, no relationship with fertility was seen in all parameters analyzed (follicle and CL size and vascularity), and contrary, cows with CL vascularity greater than 70% exhibit lower fertility. In inseminated animals, follicle size and vascularity was positive related with CL size and vascularity, as shown by the presence of greater CL size and vascularity originated from follicle with also greater size and vascularity. This is the first time that ultrasound based fertility prediction methods were tested prior to ET and AI and showed an application in ET, but not in AI programs. Further studies are needed including hormone profile evaluation to improve conclusion.

Modulation of post-partum reproductive performance in dairy cows through supplementation of long- or short-chain fatty acids during transition period.

Thirty-six cross-bred cows were used to study the effect of long-chain (flaxseed) or short-chain (butyric acid) fatty acid supplementation on metabolic status, ovarian function and reproduction performance during transition period. Control cows received a routine feed of transition diet, while the cows in two treatment groups were supplemented with either 750-g crushed flaxseed or 250 g butyric acid per cow per day. Ovarian activity was monitored by transrectal ultrasonography on 10th, 20th and 30th days post-partum. Blood samples were collected by jugular venipuncture into heparinized polystyrene tubes; plasma was prepared and stored under -20 °C until analysis. Results indicated that cows in flaxseed group were in positive energy balance as indicated by lower NEFA and Beta hydroxy Butyrate and higher glucose concentrations. Uterine involution was completed well within 30 days post-partum in all the cows in flaxseed fed group compared to 76.9% in butyric acid supplemented and 61.5% in control groups. The size of dominant follicle and corpus luteum was significantly higher (p < 0.05) for flaxseed group compared to control group, which in turn resulted in higher concentrations of plasma progesterone. Cows fed on diets supplemented with flaxseed exhibited post-partum heat earlier and bred sooner (p < 0.05) than control cows. It has been noticed that supplementation of flaxseed and butyric acid enhanced involution of uterus, early resumption of cyclicity and thereby early breeding. However, in view of the encouraging results obtained for flaxseed supplemented group, its organic nature and easier availability at farmer's gate, we concluded that flaxseed can be safely included in transition diet to modulate reproductive performance of dairy cattle.

Exposure to di(2-ethylhexyl) phthalate inhibits luteal function via dysregulation of CD31 and prostaglandin F2alpha in pregnant mice.

BackgroundDi(2-ethylhexyl) phthalate (DEHP) exposure reduces embryo implantations, increases embryonic loss, and decreases fetal body weights. However, whether it is associated with the alteration of luteal function remains unknown. Thus, our aim in this study was to explore the effect and mechanism of DEHP on luteal function in pregnant mice in vivo.MethodsMice were administered DEHP by gavage at 125, 250, 500 mg/kg/day from gestational days (GD) 1 to 9 or 13. Levels of serum progesterone and estradiol were measured by radioimmunoassay. The numbers and sizes of corpora lutea were calculated by ovarian histomorphology. Steroidogenic enzymes were assessed by qRT-PCR. CD31 protein was detected by immunocytochemistry, and prostaglandin F2alpha (PGF2alpha) levels were evaluated by enzyme immunoassay.ResultsTreatment with DEHP significantly inhibited progesterone secretion in pregnant mice in a dose-dependent manner but did not inhibit estradiol production on GD 9 and 13. Treatment also showed concomitant decreases in transcript levels for key steroidogenic enzymes (CYP11A, 3β-HSD, and StAR) on GD 13. Furthermore, DEHP administration significantly reduced the numbers and sizes of corpora lutea on GD 13. No significant changes in the ratio of ovary weight vs. body weight were observed between the control group and treated animals on GD 9 and 13. In addition, treatment with DEHP significantly inhibited CD31 expression of corpora lutea, whereas plasma PGF2alpha levels in DEHP treatment groups were significantly higher compared with the control groups on GD 9 and 13.ConclusionsThe results show DEHP significantly inhibits luteal function of pregnant mice in vivo, with a mechanism that seems to involve the down-regulation of progesterone and steroidogenic enzymes message RNA, the decrease in CD31 expression, and the increase in PGF2alpha secretion.Electronic supplementary materialThe online version of this article (doi:10.1186/s12958-015-0013-4) contains supplementary material, which is available to authorized users.

Investigation of the Effects of Acupuncture Stimulation on the Size and Blood Flow of Corpus Luteum and Progesterone Levels in Dairy Cows

Investigation of the Effects of Acupuncture Stimulation on the Size and Blood Flow of Corpus Luteum and Progesterone Levels in Dairy Cows

11 FERTILITY RESPONSE IN SUCKLED BEEF COWS SUPPLEMENTED WITH LONG-ACTING PROGESTERONE AFTER TIMED ARTIFICIAL INSEMINATION

Corpus luteum (CL) and progesterone (P4) secretion are affected by preovulatory follicle (POF) size. Increased circulating P4 during early diestrus has a positive effect on embryo development in beef cattle. However, the combined effects of the POF size and P4 supplementation during early diestrus on fertility of beef cows are not known. The objective was to evaluate the effects of POF size and supplementation of long-acting P4 after timed-AI on pregnancy rates (P/AI). Suckled Nelore cows (n = 596) were evaluated twice by transrectal Doppler ultrasonography (US) 10 days apart to detect the cyclic status. In Study 1, anestrous cows (absence of CL on both exams) received an intravaginal P4 device and an oestradiol benzoate (EB) injection on Day –10 (day of the second US). Devices were removed and sodium cloprostenol [prostaglandin F2α (PGF2α)], oestradiol cypionate, and eCG were given on Day –2. Cows were timed-AI on Day 0 and assigned to receive placebo (control group, n = 187) or 150 mg of long-acting P4 on Day 4 (P4 group, n = 189). In Study 2, cyclic cows (presence of CL) received a PGF2α injection on Day –20 (first US). Cows with a new CL on Day –10 received an intravaginal P4 device and an injection of EB and were split to receive an injection of PGF2α [large follicle (LF); n = 109] or not [small follicle (SF); n = 111]. Devices were removed and PGF2α was injected on Day –2. Ovulation was induced with buserelin acetate, and cows were timed-AI on Day 0 and split to receive placebo (LF/control group, n = 55, and SF/control group, n = 55) or 150 mg of long-acting P4 on Day 4 (LF/P4 group, n = 56, and SF/P4 group, n = 54). Ultrasonographic scanning was done on Days 0, 4, and between 35 and 40 to detect the POF and CL sizes and P/AI, respectively. Data were analysed using PROC GLIMMIX (SAS Institute Inc., Cary, NC). In anestrous cows, P/AI was reduced in POF with &lt;11 mm. The P/AI was greater in the P4-treated group than in the control group for all cows (55.6% v. 46.0%; P = 0.05) and for ovulated cows (59%, 105/178 v. 49%, 86/173; P = 0.08). For cyclic cows, POF size (mm) on Day 0 (13.5 ± 0.3 v. 11.2 ± 0.2), ovulation rate (90% v. 77%), and CL area (cm2) on Day 4 (1.46 ± 0.05 v. 1.25 ± 0.05) were greater (P &lt; 0.007) in the LF group than in the SF group. There was a main effect of follicle group on P/AI (54%, LF group v. 38%, SF group; P &lt; 0.01). Moreover, P/AI were greater (P &lt; 0.05) in the LF/control (56%) and LF/P4 (52%) groups than in the SF/control group (31%), whereas no difference was detected between the SF/P4 group (45%) and the other groups. Among cows that ovulated, P/AI was lower (P = 0.05) in the SF/control group (41%, 17/41) compared with the LF/control group (62%, 31/50) and were similar for the SF/P4 group (56%, 25/45) and LF/P4 group (57%, 28/49) compared with others. We suggest that P4-stimulated embryotrophic effects improved fertility in anestrous beef cows supplemented with long-acting P4 on Day 4 after timed-AI. Also, the presence of a functional CL during follicle growth results in smaller POF and CL and reduces the ovulatory and P/AI rates in cyclic cows. Post-AI P4 supplementation may attenuate the negative effects of small POF/CL. Research was supported by CNPq, FAPESP, Ouro Fino Agronegócio, and Innovare.

165 EFFECT OF INJECTION OF SEMINAL PLASMA ON OVULATION RATE, CORPUS LUTEUM DEVELOPMENT, AND SENSITIVITY TO PROSTAGLANDIN IN ALPACAS (VICUGNA PACOS)

The seminal plasma (SP) of camelids contains a protein identified as β Nerve Growth Factor with capacity of induced ovulation and develop a corpus luteum. A study was designed to evaluate the effect of application of SP on the interval of time from injection of stimulus to the ovulation and corpus luteum (CL) size (Experiment 1, n = 24) and on the sensitivity of CL to the injection of prostaglandin (196 µg of tiaprost) (PG) at different periods from ovulation (Experiment 2, n = 86). Exp. 1: Adult female alpacas with presence of a follicle ≥7 mm were assigned to the application of 1 mL of SP via IM (T: n = 12) or application of 50 µg of acetate of busereline IM (T2: n = 12). Exp. 2: Alpacas with presence of a follicle ≥7 mm were induced to ovulation with 50 µg of acetate of busereline or 1 mL IM of SP. Animals were evaluated by ultrasound to confirm the ovulation and were assigned to the following treatment: T1 (n = 8): SP + PG Day 4; T2 (n = 8): buserelin acetate + PG Day 4; T3 (n = 8): SP + PG Day 5; T4 (n = 8): buserelin acetate + PG Day 5; T5 (n = 8): SP + PG Day 6; T6 (n = 8): buserelin acetate + PG Day 6; T7 (n = 8): SP + PG Day 7; T8 (n = 8): buserelin acetate + PG Day 7; T9 (n = 8): SP + PG Day 8; T10 (n = 8): buserelin acetate + PG Day 8 and T11 (n = 6) (Control): Application of 1 mL of saline solution. The animals were evaluated by ultrasound with an Aloka SSD500 (Aloka, Tokyo, Japan) and 7.5-MHz linear transducer each 2 h (Exp. 1) and each 12 h (Exp. 2) after of application of PG. In Exp. 1, the ovulation rate was 95.7% to T1 and T2 and an interval of time between injection of stimulus and ovulation was 27.4 ± 2.5 h and 26.8 ± 1.8 to T1 and T2, respectively. In Exp. 2, luteolysis was 0.0%, 0.0%, 25.0%, 0.0%, 100.0%, 100.0%, 100.0%, 100.0%, 100.0%, 100.0%, and 0.0% for the treatments T1, T2, T3, T4, T5, T6, T7, T8, T9, T10, and T11 respectively. The results suggest that no differences exist between in the ovulation rate and interval to the ovulation between the application of buserelin acetate or SP and that the CL was sensible at Day 5 to the prostaglandin respect SP and with similar response to the sensibility of CL from Day 6 to Day 8.

171 EXPRESSION OF THE OVULATION-INDUCING FACTOR/NERVE GROWTH FACTOR HIGH AFFINITY RECEPTOR IN THE OVARIES OF COWS DURING THE PERIOVULATORY PERIOD

Ovulation-inducing factor/nerve growth factor (OIF/NGF) influences ovulation and corpus luteum (CL) size and function in camelids and, remarkably, cows. The ovulation effect in induced ovulators is mediated at the hypothalamo-pituitary-axis, but the site and mechanism of action of the luteotrophic effect is unknown. This mechanism may be an important aspect of OIF/NGF in spontaneous-ovulators. The objective of this experiment was to detect changes in expression of the high affinity OIF/NGF receptor (TrkA) in the ovary during the periovulatory period in cattle. Cows (n = 14) were examined daily by transrectal ultrasonography to determine the day of ovulation (Day 0), and were assigned randomly to be unilaterally ovariectomized on Day 2, 4, 6, or in the preovulatory period before or after the LH surge. Cows assigned to preovulatory groups were given a luteolytic dose of prostaglandin when the dominant follicle of the second follicular wave was ≥10 mm and growing. Cows assigned to the pre-LH group were ovariectomized 24 h after prostaglandin treatment. Cows assigned to the post-LH group were given LH 24 h after prostaglandin treatment and were ovariectomized 18–20 h later. Cows were allowed to rest for one complete interovulatory interval and re-assigned to a different day-group on which the remaining ovary was removed (n = 4 to 5 ovaries/day-group). Ovaries were fixed in paraformaldehyde, and 5-μm sections of ovarian tissue representing the dominant follicle, largest subordinate follicle, and the CL were treated for enzymatic antigen retrieval and blocked in 1% BSA for 1 h. Slides were incubated overnight with the primary antibody (rabbit anti-human TrkA) and for 2 h with the secondary antibody (goat anti-rabbit IgG). Slides were evaluated by fluorescence microscopy. Images of 3 arbitrarily chosen fields (40×) were captured for each structure for each day-group, and the average proportion of immunoreactive cells in the theca layer and luteal tissue was estimated using ImageJ software (National Institutes of Health, Bethesda, MD, USA). The intensity of individual cell immunofluorescence was also scored as no reaction (0), faint (1), weak to moderate (2), or strong (3). Data were compared among groups by two-way ANOVA. The proportion of immunoreactive cells was higher in the dominant follicle than in the subordinate follicle or the CL (P &lt; 0.001); no effect of day-group or interaction was detected. The intensity score increased from faint on Day 2 to strong on Day 6, and remained strong in the pre- and post-LH groups. A discernible pattern of change in intensity score was not evident for the subordinate follicle or the CL. The greater proportion of TrkA positive cells and greater immunoreactive intensity in the maturing dominant follicle support the hypothesis that the luteotrophic effect of OIF/NGF in cattle is a result of a local increase in the expression of TrkA in the theca layer of the dominant follicle. Research was supported by the Natural Sciences and Engineering Research Council of Canada.

Ovulatory Follicular Dynamics After Estrus Synchronization using Prostaglandin F2a in Dairy Cows

The study aimed to follow development of ovulatory follicular dynamics as well as plasma progesterone profile after estrus synchronization using PGF2 and GnRH. A total of 15 non-pregnant dairy cows, 4-5 years of age, healthy and reproductively sound were used in the present study. Treatment 1, given intramuscular injection of PGF2 25 mg (PGF2), treatment 2 PGF2 25 mg and GnRH 250 g 2 days later (PGF2-GnRH), and treatment 3 with GnRH 250 g (7 days prior to injection of PGF2), PGF2 25 mg and GnRH 250 g (2 days after injection of PGF2) (GnRH-PGF2a-GnRH) (the Ovsynch method). Transrectal ultrasonographic examination using real time, B-mode, with 7.5 MHz tranducer was performed everyday for 12 days to follow ovulatory follicular and luteal dynamics. Blood plasma was taken every day for progesterone determination using EIA technique. Data of follicular, luteal development and progesterone levels were tested using analysis of variance and correlation analysis. The animals showed estrus within 70.70 + 01.90 hours following PGF2 injection. Prostaglandin F2 induced corpus luteum regression, decreased in progesterone plasma levels, followed by ovulatory follicular development and eventually underwent ovulation. Administration of first GnRH increased corpus luteum size, enhanced its regression and decreased plasma progesterone levels, while the second administration induce better ovulatory follicular development. Rate of the corpus luteum regression, progesterone decrease and ovulatory follicular development following PGF2 injection for respective treatments 1, 2 and 3 were 2.53 + 0.24a, 2.73 + 0.36a and 3.53 + 0.28b mm/day; 1.39 + 0.14a, 1.35 + 0.18a dan 1.57 + 0.12b ng/ml/day; and 1.33 + 0.15a, 1.63 + 0.19b and 1.67 + 0.23b mm/day, respectively (P &lt; 0.05). It can be concluded that PGF2 induced corpus luteum regression, decreased in progesterone plasma levels and ovulatory follicular development. Addition of GnRH increased corpus luteum size and plasma progesterone levels, after PGF2 injection corpus luteum regression and progesterone decrease became more prominent, while ovulatory folliculkar development occurred much better. .

Modulation of periovulatory endocrine profiles in beef cows: consequences for endometrial glucose transporters and uterine fluid glucose levels

In beef cattle, proestrus estradiol and subsequent progesterone (P4) concentrations can regulate the endometrial characteristics and thereby determine maternal receptivity toward the embryo. However, the underlying mechanisms linking periovulatory endocrine profiles to receptivity, which is crucial to obtain pregnancy, need to be elucidated. We hypothesized that the size of the preovulatory follicle (POF) and subsequent circulating P4 concentrations, during early diestrus, modulate endometrial levels of glucose transporter transcripts and proteins, and subsequently affect the luminal glucose availability in the uterus. Therefore, follicle growth of Nelore cows was manipulated, and cows were assigned to 2 experimental groups: (1) large follicle and large corpus luteum (LF-LCL) group with a large POF and corpus luteum (CL); and (2) small follicle and small corpus luteum (SF-SCL) group with a small POF and CL. At day 7 post gonadotropin-releasing hormone induced ovulation (gonadotropin-releasing hormone treatment = day 0), animals were slaughtered (n = 18 per group), and uterine tissues and washings were collected for characterization of glucose transporters and glucose levels, respectively. The diameter of POF was larger (P < 0.05) in the LF-LCL cows compared with their SF-SCL counterparts (12.8 ± 0.4 vs 11.1 ± 0.4 mm). Furthermore, CL size (17.49 ± 0.88 vs 14.48 ± 0.52 mm) and circulating P4 concentrations at day 7 (4.5 ± 1.0 vs 3.3 ± 1.1 ng/mL, P < 0.05) were significantly higher in the LF-LCL cows compared with the SF-SCL cows. No differences (P > 0.05) were detected in gene expression patterns of SLC2A1, SLC2A3, SLC2A4, SLC2A5, SLC5A1, ATP1A2, ATP1B2, and SLC37A4. However, the protein abundance of endometrial SLC2A1was increased in the LF-LCL group compared with the SF-SCL group (P < 0.05). SLC2A1 and SLC2A4 protein products were mainly identified at the endometrial luminal and glandular epithelium membranes as well as in the endometrial stroma. Glucose concentrations in uterine washings were similar between groups. In conclusion, we provided information on the potential link between endocrine profiles and glucose transport pathways in the bovine endometrium. More specifically, our data reveal that the size of the POF, and subsequent P4 concentrations, do not functionally affect the main endometrial glucose transporter pathways or uterine fluid glucose concentrations during diestrus.

Conceptus-induced changes in the gene expression of blood immune cells and the ultrasound-accessed luteal function in beef cattle: how early can we detect pregnancy?

We aimed to identify the functional characteristics of the corpus luteum (CL) by color Doppler ultrasonography and changes in interferon-stimulated gene (ISG) expression in peripheral blood mononuclear cells (PBMCs) during early pregnancy in beef cows. We then aimed to use these features to establish earlier pregnancy diagnosis methods. In experiment 1, the CL size and blood flow were accessed by Doppler ultrasonography, and the PBMCs were isolated on Days 8, 12, 15, 18, 20, 22, 25, 30, 45, and 60 post-timed artificial insemination (TAI) from pregnant (n = 10) and nonpregnant cows (n = 12). The abundance of ISG (OAS1, MX1, MX2, and ISG15) transcripts was measured by quantitative PCR. Analyses of OAS1 and MX2 expression in isolated PBMCs (ISG-PBMC method) and Doppler imaging of CL (Doppler-US method) were performed to test the accuracy of these methods for the diagnosis of pregnancy on Day 20 post-TAI (n = 110; experiment 2). In experiment 1, the luteal volume and blood flow were reduced in nonpregnant cows during the first weeks post-TAI, but an evaluation of CL vascularization and size was efficient in identifying nonpregnant cows on Day 20 post-TAI. The expression of ISGs in PBMCs can be stimulated by the presence of a viable conceptus between Days 15 and 22 post-TAI, and the expression of these genes reaches a peak on Day 20. In experiment 2, the Doppler-US and ISG-PBMC methods resulted in similar specificity (85.5 and 87.7%, respectively). However, only the Doppler-US method resulted in 100% sensitivity. In conclusion, the greatest abundance of ISGs in PBMCs and a high detection of luteolysis by Doppler imaging on Day 20 post-TAI can be feasibly used for the earlier detection of nonpregnant cows in reproductive programs. The level of accuracy for our described pregnancy methods is high on Day 20 (80%-91%), but only the Doppler imaging method results in an absence of false-negative diagnoses.

In vivo imaging in the rabbit as a model for the study of ovulation-inducing factors

The study of factors responsible for eliciting ovulation in rabbits has been hampered by the lack of a suitable method of monitoring the ovaries invivo. Ovarian imaging by ultrasound biomicroscopy was used in two experiments designed to determine the effects of seminal plasma on the ovulatory response in rabbits. In Experiment 1, female rabbits were group-housed and treated intramuscularly with saline, gonadotropin releasing hormone (GnRH), or seminal plasma of llamas or rabbits (n = 4 to 6 per group). Rabbits were euthanized eight days later to evaluate the ovarian response by ultrasound biomicroscopy ex situ. No differences among groups were detected in the proportion of rabbits that ovulated or in the number and size of corpora lutea. The high incidence of ovulation in the negative control group was unexpected, and confounded determination of an ovulation-inducing effect of seminal plasma. In Experiment 2, female rabbits were caged individually, and treated as in Experiment 1 (n = 5 to 7 per group). The ovarian response was evaluated invivo by transcutaneous ultrasound biomicroscopy. Ovulation and formation of corpora lutea were detected only in rabbits given GnRH. A preovulatory surge in plasma luteinizing hormone concentration and a post-ovulatory rise in plasma progesterone concentration were detected only in rabbits treated with GnRH. Surgical translocation of the ovaries to a subcutaneous position enabled longitudinal assessment of the ovulatory response by ultrasound biomicroscopy. Results clearly documented the effect of physical/social interaction on ovulation in rabbits, and did not support the hypothesis that seminal plasma elicits ovulation in rabbits.

Effect of combined exogenous progesterone with luteotrophic support via equine chorionic gonadotrophin (eCG) on corpus luteum development, circulating progesterone concentrations and embryo development in cattle.

The aim was to examine the effect of a single intramuscular (i.m.) injection of equine chorionic gonadotrophin (eCG) on Day 3 after oestrus on corpus luteum (CL) development, circulating progesterone and conceptus development in cross-bred beef heifers. In Experiment 1, heifers received: (1) saline, or a single i.m. injection of eCG on Day 3 at (2) 250IU (3) 500IU (4) 750IU or (5) 1000IU. Administration of eCG resulted in increased luteal tissue area and progesterone and oestradiol concentrations compared with controls. In Experiment 2, heifers received (1) a progesterone-releasing intravaginal device (PRID Delta) from Day 3 to 5 or (2) a PRID Delta from Day 3 to 5 plus a single injection of 750IU eCG on Day 3. In vitro-produced blastocysts (n=10 per recipient) were transferred on Day 7 and heifers were slaughtered on Day 14 to assess conceptus development. Administration of eCG reduced the number of short cycles (6.3% vs 31.3%) and increased mean luteal tissue weight (P=0.02). Insertion of a PRID Delta on Day 3 resulted in an elevation (P<0.05) in serum progesterone until removal on Day 5. Administration of eCG at the time of PRID Delta insertion resulted in higher progesterone levels (P<0.05) from Day 10 onwards. Conceptus dimensions were not affected. In conclusion, a single injection of eCG on Day 3 increased CL size and progesterone concentrations and, when given in conjunction with a progesterone-releasing device, appeared to reduce the number of short cycles, presumably due to its luteotrophic nature. The implications of the elevated oestradiol concentrations for embryo quality require further study.

Effects of varying doses of β-nerve growth factor on the timing of ovulation, plasma progesterone concentration and corpus luteum size in female alpacas (Vicugna pacos).

Ovulation in camelids is induced by the seminal plasma protein ovulation-inducing factor (OIF), recently identified as β-nerve growth factor (β-NGF). The present study measured the total protein concentration in alpaca seminal plasma using a bicinchoninic acid (BCA) protein quantification assay and found it to be 22.2±2.0mgmL(-1). To measure the effects of varying doses of β-NGF on the incidence and timing of ovulation, corpus luteum (CL) size and plasma progesterone concentration, 24 female alpacas were synchronised and treated with either: (1) 1mL 0.9% saline (n=5); (2) 4µg buserelin (n=5); (3) 1mg β-NGF protein (n=5); (4) 0.1mg β-NGF (n=5); or (5) 0.01mg β-NGF (n=4). Females were examined by transrectal ultrasonography at 1-2-h intervals between 20 and 45h after treatment or until ovulation occurred, as well as on Day 8 to observe the size of the CL, at which time blood was collected to measure plasma progesterone concentrations. Ovulation was detected in 0/5, 5/5, 5/5, 3/5 and 0/4 female alpacas treated with saline, buserelin, 1, 0.1 and 0.01mg β-NGF, respectively. Mean ovulation interval (P=0.76), CL diameter (P=0.96) and plasma progesterone concentration (P=0.96) did not differ between treatments. Mean ovulation interval overall was 26.2±1.0h. In conclusion, buserelin and 1mg β-NGF are equally effective at inducing ovulation in female alpacas, but at doses ≤0.1mg, β-NGF is not a reliable method for the induction of ovulation.

Clutch size in populations and species of cnemidophorines (Squamata: Teiidae) on the eastern coast of Brazil

We analyzed some reproductive aspects of 16 coastal populations, belonging to five lizard species (A. ocellifera, A. abaetensis, A. nativo, A. littoralis and C. lacertoides) from different restinga habitats along the eastern coast of Brazil. This study aimed to evaluate to what extent the reproductive aspects vary geographically and among species. For each female, we recorded the number of vitellogenic follicles, size and color of the largest follicle, presence and size of corpora lutea, and number and size of oviductal eggs. Clutch size of almost all coastal populations/species of Ameivula had little variation and most clutches were composed of two eggs. There was a significant relationship between female size and the mean clutch size when females from different species were pooled. Mean egg volume, among species, varied from 420 to 655 mm3. Relative clutch mass varied from 0.129 to 0.159 and did not differ significantly among species. We concluded that the five coastal species studied (four bisexuals and one parthenogenetic) had similar reproductive characteristics. Most of them presented multiple clutches, low clutch size and low relative clutch mass, similar to other species in the genus and to unisexual and bisexual species of the Teiidae family.

Early pregnancy diagnosis on days 18 to 21 postinsemination using high-resolution imaging in lactating dairy cows

The aim was to assess the ability of corpus luteum (CL) and uterine ultrasound characteristics on d 18 to 21 to predict pregnancy status in lactating dairy cows. Ultrasound examinations were carried out on cows (n=164) on d 18 to 21 following artificial insemination (AI). Images of the uterus and CL were captured using a Voluson i ultrasound device (General Electric Healthcare Systems, Vienna, Austria) equipped with a 12-MHz, multi frequency, linear array probe. Serum concentrations of progesterone were determined from blood samples collected at each ultrasound examination. Images of the CL were captured and stored for calculation of CL tissue area and echotexture. Images of the CL and associated blood flow area were captured and stored for analysis of luteal blood flow ratio. Longitudinal B-mode images of the uterine horns were stored for analysis of echotexture. Diagnosis of pregnancy was made at each ultrasound examination based on CL blood flow, CL size, and uterine echotexture. Pregnancy was confirmed by ultrasonography on d 30 after AI. The relationship between ultrasound measures and pregnancy outcome, as well as the accuracy of the pregnancy diagnosis made at each ultrasound examination was assessed. Progesterone concentrations and CL tissue area were greater in pregnant compared with nonpregnant cows on all days. The CL blood flow ratio was higher in pregnant compared with nonpregnant cows on d 20 and 21 after AI. Echotexture measures of the CL and uterus were not different between pregnant and nonpregnant cows on any day of examination. The best logistic regression model to predict pregnancy included scores for CL blood flow, CL size, and uterine echotexture on d 21 following AI. Accuracy of pregnancy diagnosis was highest on d 21, with sensitivity and specificity being 97.6 and 97.5%, respectively. Uterine echotexture scores were similar for pregnant and nonpregnant cows from d 18 to 20. On d 21, pregnant cows had higher uterine echotexture scores compared with nonpregnant cows. The logistic regression equation most likely to provide a correct pregnancy diagnosis in lactating dairy cows included the visual score for CL blood flow, CL size, and uterine echotexture on d 21 after AI. In support of this finding, the diagnostic accuracy for visual scores of CL blood flow, CL size, and uterine echotexture were also highest on d 21.

Effectof human chorionic gonadotropin on post-ovulatory mid-luteal profile and subsequent pregnancy rate in anestrus buffalo using two synchronization regimes

The present study was aimed to assess the impact of post-insemination human chorionic gonadotropin (hCG) administration on mid-luteal functions and conception rate in Murrah buffalo. The study was conducted on 2 buffalo treatment groups viz., estradiol + controlled internal drug releaser (CIDR) + gonadotropin releasing hormone (GnRH, n = 27) and ovsynch + CIDR (n = 22). Half of the animals in each group received intramuscular (i.m.) hCG (3000 IU, i.m.) on day 12 post-ovulation while groups served as control. The CG administration on day 12 post-ovulation improved corpus luteum size and progesterone level as well as first service conception rate after timed insemination in buffaloes.

11 STRATEGIES TO MODULATE THE PERI-OVULATORY ENDOCRINE MILIEU BY CONTROLLING PROGESTERONE CONCENTRATIONS PRE- AND POST-INSEMINATION IN BEEF COWS

Recent evidence indicates that progesterone (P4) secretion from corpus luteum (CL) is affected by preovulatory follicle (POF) size and that increased circulating P4 during early diestrus has a positive effect on embryo development and pregnancy rates in cattle. Despite the embryotrophic effects, early administration of P4 may impair CL growth and anticipates luteolysis. Owing to develop favourable peri-ovulatory endocrine milieus to pregnancy success, we evaluated in beef cows (1) the effect of CL presence during dominant follicle growth on follicular vascularization, size and ovulation rate and (2) the effects of POF size and supplementation of long-acting P4 post-AI on CL development and regression. Sixty-six Nelore cows received an intravaginal P4 device and an injection of oestradiol benzoate on Day 10 (5–10 days post-oestrus), and were split to receive sodium cloprostenol [prostaglandin F2α (PGF2); large follicle (LF); n = 31) or not [small follicle (SF); n = 35]. Devices were removed and PGF2 was injected on Day 1.75. Ovulation was induced with a gonadotropin-releasing hormone (GnRH; buserelin acetate) on Day 0. Cows were inseminated twice on Days 0.5 and 1. Cows that ovulated were assigned to receive placebo (LF/control group, n = 14; and SF/control group, n = 9) or 150 mg of long-acting P4 on D4.5 (LF/P4 group, n = 13; and SF/P4 group, n = 12). Colour-Doppler ultrasound scanning was done on Days 10, 6, 2, 0.5, 1, 1.5, 4.5, 7.5, and 15.5. Structural luteolysis was detected by 25% decrease in CL area and 50% in luteal blood flow of subsequent scanning. Data were analysed for the main effects of POF size and supplementation and their interaction using the PROC MIXED procedure of SAS (SAS Institue Inc., Cary, NC, USA). Follicle diameter (mm) on Day 2 (11.4 ± 0.4 v. 8.4 ± 0.3) and POF (14.1 ± 0.3 v. 12.5 ± 0.3) were greater (P &lt; 0.05) in LF group. Percentage of luteal blood flow in follicle wall was greater (P &lt; 0.05) on Day 0.5 (33.9 ± 3.4 v. 22.8 ± 3.6) and 1 (48.9 ± 4.8 v. 28.3 ± 4.3) in the LF group. Ovulation rate was lower (P &lt; 0.05) in the SF group (60%, 21/35) than in the LF group (90%, 28/31), suggesting that a functional CL during dominant follicle growth is less favourable to pregnancy success due to 30% reduction in fertilizing potential of non-ovulated follicles. For luteal area (cm2), an effect (P &lt; 0.05) of POF size was detected on Day 4.5 (1.7 ± 0.1 v. 1.3 ± 0.1), 7.5 (2.8 ± 0.2 v. 2.2 ± 0.1), and 15.5 (2.5 ± 0.1 v. 2.2 ± 0.2) in the LF and SF groups, respectively. Luteal blood flow was greater (P &lt; 0.05) in the LF groups (40.4 ± 1.7 and 47.6 ± 3.4) than in the SF groups (35.7 ± 1.5 and 37.9 ± 4.1) on Day 4.5 and 15.5, respectively. This indicates that larger and more vascularized POF may be associated with greater CL vascularization in addition to greater CL size. The frequency of cows with early luteolysis did not differ (P &gt; 0.1) between P4-treated (5/25) and placebo-treated (1/23) cows. We suggest that the P4 supplementation did not severely induce early luteolysis in inseminated cows and that the P4-embryotrophic effects may down-regulate the mechanisms involved in the earlier luteolytic process caused by greater exposure of uterus to P4. Support for this study was provided by the São Paulo Research Foundation (FAPESP, Sao Paulo, Brazil) and Ourofino (Cravinhos, SP, Brazil).

10 INFLUENCE OF ADMINISTRATION OF HUMAN CHORIONIC GONADOTROPIN ON DAY 4 AFTER FIXED-TIME AI ON CORPUS LUTEUM SIZE, FUNCTION, AND PREGNANCY RATE IN BEEF COWS

Successful establishment of pregnancy in the cow depends on early rise of progesterone (P4) to improve embryo development. The present study was carried out to determine the corpus luteum (CL) function and pregnancy rate following the administration of 1.500 IU of hCG in suckled beef cows on Day 4 after fixed-time AI (FTAI). Crossbred Angus cows (n = 429), 40 to 70 days postpartum, were synchronized using 2 mg of oestradiol benzoate (EB) i.m. and an intravaginal progesterone-releasing device containing 1 g of P4 for 7 days. At device removal, prostaglandin F2a (150 mg of D-cloprostenol) was injected i.m.; 24 h later, 1 mg of EB was given and the FTAI were done 30 to 36 h after EB using semen of just 1 bull. Cows were assigned alternatively to receive 1.500 IU of hCG i.m. (n = 200) or 1 mL of saline i.m. (n = 229) on Day 4 after FTAI. Ovaries were examined by transrectal ultrasonography (n = 71) on Days 4 and 7 to detect the presence and to measure the area of CL. Blood samples were collected on Days 4, 7, 10, 14, and 30 post-oestrus to determine P4 concentration by RIA. Pregnancy diagnoses were done 30 days after FTAI. The effects of treatments on CL area and serum P4 levels were analysed by ANOVA (Proc GLM, SAS Institute Inc., Cary, NC, USA) and pregnancy rates compared by chi-square (Proc CATMOT, SAS). The induction of accessory CL determined on Day 7 was 44% in cows treated with hCG and not observed in the control group. The area of the CL on Day 7 was significantly higher (P &lt; 0.01; 3.37 ± 2.56 v. 2.56 ± 0.22 cm2) in the hCG group. Despite the increase in CL size, the mean levels of P4 on Days 4, 7, 10, 14, and 30 post-AI indicated no difference (P &gt; 0.05) in CL function between control and hCG-treated cows. On Day 30, pregnancy rates were not different (P = 0.08), at 46.7 and 55% for control and hCG-treated cows, respectively. In conclusion, administration of 1.500 IU of hCG on Day 4 post-FTAI did not improve pregnancy rate in suckled beef cows.