323 EFFICIENCY OF CLOPROSTENOL-INDUCED LUTEOLYSIS IN SUPEROVULATED COWS

2011 ◽  
Vol 23 (1) ◽  
pp. 258
Author(s):  
J. H. M. Viana ◽  
M. S. B. Vargas ◽  
L. G. B. Siqueira ◽  
B. R. C. Alves ◽  
A. P. Oliveira ◽  
...  

The induction of multiple ovulations is a key procedure for in vivo embryo production. Many corpora lutea (CL) are developed, resulting in abnormally high progesterone concentrations. Luteolysis induction by prostaglandin F2α and its analogues is well described in cows bearing one or few, but not multiple, CL, as occurs after superovulation. The objective of the first experiment was to compare the efficacy of a single prostaglandin F2α treatment on inducing luteolysis in embryo donors immediately after flushing (D7, N = 24) or 4 days later (D11, N = 22). Holstein cows were superovulated with 400 IU of FSH following standard procedures. Embryo flushing was performed 7 days after AI, and cows were randomly allocated into 2 groups to receive either a 0.5 mg of sodium cloprostenol IM treatment immediately after flushing (D7 group) or the same treatment 4 days later (D11 group). Occurrence of luteolysis was monitored by plasma progesterone concentrations (P4), measured by radioimmunoassay in blood samples taken at 4-h intervals. There was no difference in P4 before treatment between D7 and D11 groups (28.6 ± 5.2 v. 36.4 ± 7.4 ng mL–1, respectively; P > 0.05). Although cloprostenol caused a remarkable decline in P4 48 h after treatment in both groups (1.8 ± 0.3 and 1.6 ± 0.4 ng mL–1 for D7 and D11 groups, respectively; P < 0.05), P4 continued decreasing in D11 cows thereafter, remaining smaller than 1 ng mL–1 up to 196 h after treatment, whereas in D7 cows, there was no further reduction in P4. Luteolysis (P4 <1 ng mL–1) was observed in all D11 cows, but failed in 11 of 20 (55%) D7 cows, in which P4 increased after the initial cloprostenol-induced decrease. The second experiment compared luteolysis in superovulated v. nonsuperovulated cows. Non-superovulated (control group, CG, N = 8) and superovulated cows (SOV, N = 6) received a single dose of sodium cloprostenol IM (0.5 mg) on day 11 after oestrus. Morphological and functional luteolysis were monitored daily by ovarian ultrasonography and P4 analysis; also, plasma LH was measured in blood samples taken every 20 min for 1 h, during 5 days. Individual CL size was smaller (1.8 ± 0.1 v. 3.5 ± 0.4 cm2) but total luteal tissue was greater (29.8 ± 7.0 v. 3.5 ± 0.4 cm2; P < 0.05) in SOV than in CG. A considerable decrease in P4 occurred in both groups 24 h after treatment (from 51.1 ± 12.9 to 5.1 ± 0.9 ng mL–1 in SOV and from 5.9 ± 0.6 to 1.1 ± 0.1 ng mL–1 in CG); however, SOV cows did not reach P4 values similar to CG until 96 h after treatment (0.9 ± 0.3 v. 0.4 ± 0.2 ng mL–1, respectively; P > 0.05). There was no difference in initial LH values between SOV and CG (1.5 ± 0.1 and 1.5 ± 0.1 ng mL–1; P > 0.05), but the slower decrease in P4 in the SOV group prevented LH from increasing up to 96 h after luteolysis induction, whereas mean LH values increased (P < 0.05) in CG after 24 h. In conclusion, luteolysis failure may occur when cloprostenol is given at the day of flushing (7 days after AI) in superovulated cows. In addition, luteolysis induction on day 11 after SOV is efficient, but the initial high progesterone concentration results in a slower rate of P4 decrease to basal levels. The authors acknowledge CNPq and FAPEMIG Project CVZ AQP 01654/09.

Author(s):  
Natália Ávila de Castro ◽  
Carlos Eduardo Porciuncula Leonardi ◽  
Jaswant Singh ◽  
Augusto Schneider ◽  
Paulo Bayard Gonçalves ◽  
...  

This study aimed to evaluate the role of prostaglandin F2α (PGF) on ovulation. In Experiment 1, cows were randomly allocated to two treatments to receive 150 μg of d-Cloprostenol (PGF Group, n = 12) or 2 mL of NaCl 0.9% (Control Group, n = 11) and CIDRs, were removed 4 days later. No cow ovulated in Control and PGF groups. In Experiment 2, cows were randomly separated into two experimental groups to receive 4 injections of 150 μg of d-Cloprostenol (n = 9) or 2 mL of NaCL 0.9% (n = 9). In this experiment, ovulation was not observed in any cows. In Experiment 3, ovariectomized cows receive three injections of 300μg of PGF analog (PGF Group, n = 5), 100μg of Lecirelin (GnRH Group, n = 5) or 2 mL of PBS (Control Group, n = 4). The LH concentration was higher (P <0.0001) in cows from the GnRH group than in the PGF and Control groups. In experiment 4, cows with preovulatory follicles (>11.5 mm) were treated with Saline (Control Group, n = 6); Lecirelin (GnRH Group, n = 7) or Cloprostenol Sodium (PGF Group, n = 6). There was a significant increase in the vascular area of follicles from 0 to 24 h in GnRH and PGF treatments. In conclusion, PGF was not able to induce ovulation in cows with high or low plasma progesterone concentration. Additionally, PGF alone was not able to induce LH release and follicle luteinization, but increased follicular vascularization.


2007 ◽  
Vol 19 (1) ◽  
pp. 223
Author(s):  
T. Okazaki ◽  
E. Sasaki ◽  
K. Hasegawa ◽  
T. Takani ◽  
S. Abe

Recent studies have shown that the presence of accessory or multiple corpora lutea (CL) and increased progesterone (P4) concentrations reduced early embryonic mortality in cattle. The objective of this study was to evaluate the effect of equine chorionic gonadotropin (eCG) treatment on the number of CL, the P4 concentrations, and pregnancy rates after embryo transfer (ET). Holstein heifers (n = 120) from 7 dairy farms received an intravaginal progesterone-releasing device (CIDR; InterAg, Hamilton, New Zealand) and 2 mg IM of estradiol benzoate (EB; Gynandol®; Sankyo, Tokyo, Japan) at random stages of the estrous cycle. After 7 to 9 days, CIDRs were removed and 15 mg of prostaglandin F2α (PG; Pronalgon®; Pfizer Japan, Nagoya, Japan) were administered, followed by 100 µg IM GnRH (Conceral®; Takeda Pharmaceutical Co., Ltd., Osaka, Japan) 2 days later (Day 0). The heifers were placed at random into 3 groups for eCG treatment. The eCG was not administered in a control group (n = 53); heifers in other 2 groups received 1000 IU eCG (Peamex®; Sankyo, Japan) IM at the time (0 h group, n = 37) or 48 h before (48 h group, n = 30) PG injection/CIDR removal. On Day 7, heifers were examined by ultrasonography (Aloka SSD500; Aloka, Tokyo, Japan) for number of CL; heifers with at least one functional CL received an in vivo-derived frozen–thawed embryo by direct transfer. At the same time, a blood sample was collected to determine P4 concentration. Pregnancy rates were determined on Days 30 and 60 by ultrasonography and rectal palpation, respectively. The data were analyzed by ANOVA and means were compared with Fisher's PLSD. Proportional data were analyzed by the chi-square test. P4 concentrations (mean ± SD) on Day 7 were 1.8 ± 1.0, 5.6 ± 3.3, and 2.2 ± 1.1 ng mL−1 for the control, 48 h, and 0 h groups, respectively (48 h vs. control and 0 h; P &lt; 0.001). The number of CL on Day 7 were 1.1 ± 0.4, 2.5 ± 1.4, and 1.8 ± 0.9 for the control, 48 h, and 0 h groups, respectively (control vs. 48 h and 0 h, and 48 h vs. 0 h; P &lt; 0.01). Pregnancy rates did not differ between 0 and 48 h groups but both were higher than in the control group (Table 1). Results suggest that the estrus synchronization protocol with administration of eCG at the time of CIDR removal or 48 h earlier significantly increased the number of CL and the P4 concentration, and improved pregnancy rates in Holstein heifers after ET. Table 1.Pregnancy rates of Holstein heifers synchronized with CIDR and PG and treated with eCG


2010 ◽  
Vol 53 (3) ◽  
pp. 575-582 ◽  
Author(s):  
Jacques Natan Grinapel Frydman ◽  
Adenilson de Souza da Fonseca ◽  
Vanessa Câmara da Rocha ◽  
Monica Oliveira Benarroz ◽  
Gabrielle de Souza Rocha ◽  
...  

This work evaluated the effect of in vitro and in vivo treatment with ASA on the morphology of the red blood cells. Blood samples or Wistar rats were treated with ASA for one hour. Blood samples or animals treated with saline were used as control group. Blood smears were prepared, fixed, stained and the qualitative and quantitative morphology of red blood cells were evaluated under optical microscopy. Data showed that the in vitro treatment for one hour with ASA at higher dose used significantly (p<0.05) modified the perimeter/area ratio of the red blood cells. No morphological alterations were obtained with the in vivo treatment. ASA use at highest doses could interfere on shape of red blood cells.


1975 ◽  
Vol 67 (1) ◽  
pp. 81-88 ◽  
Author(s):  
N. L. POYSER ◽  
E. W. HORTON

SUMMARY Five guinea-pigs actively immunized against a prostaglandin F2α(PGF2α)–bovine serum albumin conjugate showed elongated oestrous cycles. During these, corpora lutea were maintained in a functional secretory state as indicated by plasma progesterone levels. The results are compatible with the view that the PGF2α antibodies neutralized the PGF2α released from the uterus and thus prevented its normal luteolytic effect. Similar patterns of progesterone secretion were observed in two hysterectomized animals and in two animals with intra-uterine implants of indomethacin.


1976 ◽  
Vol 56 (1) ◽  
pp. 37-42 ◽  
Author(s):  
R. RAJAMAHENDRAN ◽  
P. C. LAGUË ◽  
R. D. BAKER

Progesterone levels were estimated by radioimmunoassay in blood samples obtained by venipuncture on the day of estrus and every alternate day until the onset of the next estrus in eight cycling dairy heifers. The mean level of progesterone was < 1 ng/ml during the first 2 days of the cycle, increased rapidly over the 4th–12th day period and reached a peak level value of 5.2 ± 1.1 ng/ml on day 14. Thereafter, the level declined rapidly to 2.6 ± 0.6 ng/ml on day 16 and then more gradually to 0.4 ± 0.1 ng/ml on day 21. In the second experiment, eight cycling heifers at diestrus were treated with gonadotrophin (2,000 IU PMSG or 1,000 IU PMSG + 1,000 IU HCG) followed 48 h later by 15 mg prostaglandin (PGF2α). Mid-ventral laparotomies were performed 4 days after the onset of estrus to observe ovarian activity. Progesterone levels were considerably higher in some animals and were slightly higher on the average after gonadotrophin treatments. The number of corpora lutea (CL) in these heifers ranged from 1 to 17. Progesterone levels of three heifers with 4–9 CL did not differ (P > 0.05) from those of three heifers with single CL. Two heifers each with 17 CL had peak progesterone levels of 38.4 and 27.8 ng/ml which were still high (9.6 and 26.5 ng/ml) by day 21. The remaining six heifers had low progesterone levels (< 1 ng/ml) by days 8–14, indicating premature regression of the CL. Thus, progesterone levels were not correlated with the number of CL.


2010 ◽  
Vol 22 (1) ◽  
pp. 379
Author(s):  
H. Kohram ◽  
G. Mohammadi ◽  
E. Dirandeh

This study was done to consider relationships of luteal phase variables (prior to AI) with follicular waves. The estrous cycles of 15 buffaloes were synchronized with 2 i.m. injections of prostaglandin F2α given 11 days apart. The buffaloes were randomly assigned to 1 of 3 treatments. Buffaloes in the control treatment received no treatment, whereas G6 buffalos received a GnRH injection between Day 5 and 7 and G16 buffalos received a GnRH injection between Day 15 and 17 of the estrous cycle (estrus = Day 0). Daily, from estrus Day 0 to the next estrus Day 23, buffaloes had their ovaries scanned by ultrasound. Blood samples were collected by tail following each ultrasound examination from estrus until next estrus (estrus = 0). Concentrations of plasma progesterone were determined by radioimmunoassay kit. The limit of detection of the assay was 0.1 45 ng mL-1 and the intra- and interassay coeffients of variation were 7.4% and 9.2%, respectively. Data were analyzed by using PROC GLM of SAS (SAS Institute, Cary, NC, USA). For comparisons between groups, the 2-sample t-test was used for continuous traits, such as size of CL or hormone concentrations. Prospective comparisons of indices of progesterone indicated that the length of luteal lifespan was longer in 3-wave than in 2-wave buffaloes (P < 0.01). Plasma progesterone concentrations were similar at peak and measured as area under the curve on Day 5 through 17 preceding insemination in 2-wave (5.30 ± 0.40 ng mL-1) and 3-wave buffaloes (5.10 ± 0.20 ng mL-1). Length of the luteal phase (defined as from the day of estrus until the last day on which plasma progesterone remained >2 ng mL-1) was <2 days shorter in 2-wave buffaloes than in 3-wave buffaloes (15.20 ± 0.40 v. 17.10 ± 0.50 d; P < 0.05). In addition, the day of peak progesterone occurred earlier in 2-wave buffaloes (13.50 ± 0.30 v. 15.30 ± 0.70 d; P < 0.05).


2013 ◽  
Vol 25 (1) ◽  
pp. 273
Author(s):  
K. Imai ◽  
S. Sugimura ◽  
M. Ohtake ◽  
Y. Aikawa ◽  
Y. Inaba ◽  
...  

We previously reported that follicular wave synchronization and follicular growth treatment (FGT) before ovum pick-up (OPU) were effective in improving oocyte competence, which was associated with an increase in related embryos obtained by somatic cell nuclear transfer (Sugimura et al. 2012 Cell. Reprogram. 14, 29–37). However, oxygen consumption in oocytes remained unknown. The present study was designed to examine the differences in oxygen consumption between bovine oocytes obtained by OPU with or without FGT after in vitro maturation. Holstein dry cows (n = 8) were reared under the same feeding and environmental conditions. Two OPU sessions were conducted in each cow to collect immature oocytes, as described by Sugimura et al. (2012). The first OPU session (OPU group) was performed in cows on arbitrary days of the oestrous cycle, using a 7.5-MHz linear transducer with the needle connected to an ultrasound scanner. Follicles larger than 8 mm in diameter were then aspirated and a controlled internal drug release device (CIDR) was inserted on Day 5 (the day of the first OPU session = Day 0). Then 30 Armour units (AU) of FSH (Antrin, Kyoritsu Seiyaku, Tokyo, Japan) was administrated to cows twice a day from Day 7 to 10 in decreasing doses (6, 6, 4, 4, 3, 3, 2, 2 AU day–1). Cloprostenol (prostaglandin F2α; 0.75 mg) was administered in the morning of Day 9. The second OPU session (FGT-OPU group) was performed 48 h after prostaglandin F2α administration (Day 11), and only follicles larger than 5 mm in diameter were aspirated. The CIDR was removed from the cows just before OPU. Collected cumulus–oocyte complexes in the OPU and FGT-OPU groups were matured in vitro as described by Imai et al. [2006 J. Reprod. Dev. 52(Suppl.), S19–S29]. To collect in vivo-matured oocytes (control group), the CIDR was inserted into the cows on arbitrary days of the oestrous cycle (= Day 0), and oestradiol benzoate (0.8 mg) was administered on Day 1. The cows received the FGT treatment (as described above) from Day 6 to 10; however, the CIDR was removed in the evening of Day 8. Buserelin (gonadotropin-releasing hormone; 200 µg) was then administrated in the morning of Day 10, and OPU was performed at 24 h after gonadotropin-releasing hormone administration (Day 11). Oxygen consumption of matured oocytes was measured noninvasively with a scanning electron microscopy system (HV-405SP; Hokuto Denko Co., Tokyo, Japan). Data were analysed by ANOVA followed by a Tukey-Kramer test. There was no difference in the mean oxygen consumption between the FGT-OPU group (0.34 ± 0.02 × 10–14 mol–1, mean ± SEM) and control group (0.40 ± 0.01 × 10–14 mol–1). However, oxygen consumption in the FGT-OPU and control groups was significantly lower (P < 0.01) than that in the OPU group (0.50 ± 0.02 × 10–14 mol–1). These results revealed significantly lower oxygen consumption in OPU-derived in vitro-matured bovine oocytes after FGT treatment compared with those obtained without FGT treatment. Oxygen consumption of oocytes obtained from FGT-OPU was similar to that of in vivo-matured oocytes, which may reflect their cytoplasmic maturation status with high developmental competence.


2011 ◽  
Vol 23 (1) ◽  
pp. 263
Author(s):  
F. Pereyra-Bonnet ◽  
A. Gibbons ◽  
M. Cueto ◽  
R. Bevacqua ◽  
L. Escobar ◽  
...  

Microinjection of DNA into the male pronucleus is a commonly used method to generate transgenic animals. However, it is only moderately efficient in several species because it requires proper male pronuclear visualisation, which occurs only in a narrow window of time in mice. The cytoplasmic microinjection of exogenous DNA (eDNA) is an alternative method that has not been fully investigated. Our objective was to evaluate if cytoplasmic microinjection of eDNA is capable of producing genetically modified embryos. In vitro and in vivo derived sheep embryos were cytoplasmically microinjected with pCX-EGFP previously incubated (5 min in a PVP droplet) with oolemma-cytoplasm fragments obtained from donor oocytes by microsurgery. A control group using microinjected plasmid alone was included in the in vivo procedure. For in vitro microinjection, IVF embryos were microinjected with circular plasmid with promoter (50 or 500 ng μL–1) or without promoter (50 ng μL–1) at 6 h after fertilization. The IVF was performed following (Brackett and Olliphant 1975 Biol. Reprod. 12, 260–274) with 15 × 106 spermatozoa mL–1, and presumptive zygotes were cultured in SOF. The expression of enhance green fluorescent protein (EGFP) was determined under blue light. For in vivo microinjection, embryos from superovulated sheep (by standard procedures) were recovered and microinjected with 50 ng μL–1 of linearized plasmid without promoter at 12 h after laparoscopic insemination with frozen semen (100 × 106 spermatozoa per sheep). Plasmid without promoter was used to avoid any possible cytotoxic effect produced by EGFP expression. The microinjection of IVF embryos with 50 ng μL–1 of plasmid was the best condition to produce embryos expressing eDNA (n = 96; 46.9% cleaved; 12.2% blastocysts; 53.0 and 4.1% of green embryos and blastocysts, respectively). Variables between the groups with or without promoter IVF were not statistically different (Fisher test: P < 0.05); however, when 500 ng μL–1 was microinjected, no blastocysts were obtained. In the in vivo embryo production group, 111 presumptive zygotes were microinjected (n = 37; with plasmid alone) from 16 donor sheep (11.5 ± 4.0 corpora lutea; 8.4 ± 4.8 presumptive zygotes recovered; 74.3% recovery rate). The mean time from injection to cleavage was 18.0 ± 4.5 h, and the percentage of cleavage and damage (due to the embryo injection) were >70% and <10%, respectively. Fifty-eight good quality embryos were transferred into the oviducts of 19 surrogate ewes; 12 of them are pregnant (63.1%). The presence of green IVF embryos demonstrates that eDNA was transported to the nucleus after cytoplasmic injection. We believe that the multi-fold increase (50- to 100-fold) in plasmid concentration compared with that used by others was the key step to our successful cytoplasmic microinjection. Accordingly, the new/old methodology described in this study provides an easy DNA construct delivery system of interest for the implementation of early reprogramming events. In addition, results obtained in the near future using in vivo cytoplasmic microinjection with high concentrations of eDNA could revalidate this technique for producing genetically modified large animals.


2020 ◽  
Vol 32 (2) ◽  
pp. 189
Author(s):  
M. Younis ◽  
M. Irfan-ur-Rehman Khan ◽  
A. Murtaza ◽  
M. Abbas ◽  
M. Z. Tahir ◽  
...  

Pakistan has 30.9 million heads of sheep; however, little information is available on their reproductive aspects. The objective of this study was to document ovarian physiology and endocrinology of Lohi ewes during the oestrous cycle. Nine Lohi ewes, synchronized by administering single prostaglandin F2α (PGF2a; Cyclomate, Star Laboratories), were monitored for ovarian follicular dynamics using transrectal ultrasonography (7.5MHz, HS-1500, Honda) for two consecutive oestrous cycles during the breeding season (September to November 2018). Changes in plasma progesterone and oestradiol-17β concentrations of ewes (n=9) were also determined during the oestrous cycle using radioimmunoassay. The interovulatory interval of Lohi ewes averaged 17.0±0.1 days, and the duration of follicular and luteal phases was 4.6±0.2 and 11.3±0.2 days, respectively. Follicles emerged in either 3- or 4-wave patterns, but the frequency of the 3-wave pattern was higher than that of the 4-wave (87 vs. 13%, respectively; P=0.05). Following ovulation (Day 0), follicles (=3mm) in 3-wave cycles (n=14) emerged on Days 0.7, 5.2, and 10.5, whereas in 4-wave cycles (n=2) follicles emerged on Days 0.1, 4, 8.5, and 11.5. The maximum diameter of preovulatory follicles and corpora lutea (CL) were 5.4±0.3 and 10.4±0.3mm, respectively. Regardless of the wave pattern, single ovulation occurred in each cycle. The CL was first detectable on Day 4±0.1, it reached maximum diameter on Day 9±0.1, and luteolysis began on Day 12.2±0.2 of the cycle. The peak plasma oestradiol-17β concentration (42.5±2.6 pgmL−1) was observed 48h before ovulation and correlated with the diameter of the preovulatory follicle during the follicular phase (r=0.84; P&lt;0.05). The peak plasma progesterone concentration (11.8±1.7ngmL−1) was observed on Day 9±0.1 and coincided with the diameter of CL throughout the oestrous cycle (r=0.93; P&lt;0.05). In conclusion, the majority of oestrous cycles in Lohi ewes had a 3-wave pattern and were mono-ovulatory in nature.


2011 ◽  
Vol 35 (4) ◽  
pp. 817-825
Author(s):  
José Nélio de Sousa Sales ◽  
Lilian Mara Kirsch Dias ◽  
Celso Rodrigues Franci ◽  
Alexandro Aluísio Rocha ◽  
Guilherme Gastão Cardoso ◽  
...  

Two experiments were conducted to evaluate the effect of the intramuscular injection of betacarotene associated to tocopherol on the plasma concentration progesterone of superovulated Holstein heifers (experiment 1) and in crossbred (Bos taurus x Bos indicus) heifers submitted to fixed-time embryo transfer (FTET, experiment 2). In experiment 1, after estrus synchronization and superovulation animals were inseminated 12 and 24 hours after estrus onset and embryos flushed 7 days later. Heifers were allocated randomly to one of three treatments: Control; T800 (800 mg of betacarotene plus 500 mg of tocopherol) and T1200 (1,200 mg of betacarotene plus 750 mg of tocopherol). The treatments were given on the day of ear implant placement and repeated on the first day of superovulation. Blood samples were collected on D0, D5, D9, D12 and D16. In experiment 2, treatments were imposed at intravaginal device insertion (D0). The same experimental design, as in experiment 1, was used. Blood samples were collected on D17 (embryos implanted) for progesterone determination by radioimmunoassay. In experiment 1, average plasma progesterone concentrations after corpora lutea formation (D12 plus D16 means) were 13.7±1.8 ng/ml, 14.5±2.3 ng/ml and 10.8±2.3 ng/ml for control, T800 and T1200, respectively, and did not differ (P=0.44). In experiment 2, progesterone concentrations on D17 in Control (8.88±0.57 ng/ml), T800 (7.48±0.64 ng/ml) and T1200 (5.90±1.33 ng/ml) groups were similar (P=0.11). Results indicate that the supplemental betacarotene and tocopherol injections did not influence peripheral progesterone concentrations in superovulated Holstein donors and crossbreed recipients heifers.


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