Reproduction in Domestic Animals - Facultad de Ciencias Veterinarias

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16 t h International Congress on Animal Reproduction 72 Poster Abstracts P132 Freezing of goat semen in AndroMed ® , a soybean lecithin based extender Becker-Silva, SC. 1 ; Holtz, W. 2 1Santa Cruz State University, Ilhéus, Brazil; 2 Inst of Animal Husbandry and Genetics, Georg-August-Univ., Goettingen, Germany Introduction The composition of semen extenders containing products of animal origin such as milk or egg yolk remains undefined, making standardization impossible. Furthermore the risk of contamination with infectious agents and introduction of animal diseases into other regions or foreign countries must be considered. Consequently, chemically defined extenders with no animal components are desirable. In the past, several such extenders have been examined with variable results. A commercially available semen extender for the bovine (AndroMed ® , Minitueb, Tiefenbach, Germany), containing soybean lecithin instead of material of animal origin has been shown to be suitable for the freezing of ovine semen. This investigation was designed with the intention of examining in vitro the suitability of AndroMed for the cryopreservation of caprine semen. Materials and Methods From 4 mature Boer goat sires, 26 ejaculates were collected with the aid of an artificial vagina. Each ejaculate was split into half. One part was diluted at 30°C with conventional TRISegg yolk extender containing 6.8% glycerol (TYG), the other with AndroMed extender. Both were treated in a similar way, i.e. cooling to 4°C within 2h, aspiration into 0,25ml straws, freezing in nitrogenvapor at -120°C and storage in liquid nitrogen. After thawing at 38°C, motility (MOT, expressed as percentage of native semen) and membrane integrity (MI) after eosin-nigrosin staining (as percentage of TYG results) were assessed. After 3 h of incubation at 38°C, MOT was assessed once again. Results The post-thaw MOT of goat semen cryopreserved in AndroMed diluent was 54% as compared to 55% in semen frozen in conventional TYG extender. The corresponding MI values were 118 and 100%. After 3h of incubation, the MOT for the AndroMed samples was 31% vs. 37% for the TYG samples. None of these differences were statistically significant (P > 0.05, Student’s t-test). Conclusion On the basis of in vitro assessment, the cryopreservation of caprine semen in AndroMed ® , a diluent free of animal protein originally designed for bovine semen, was just as successful as freezing in conventional TRIS-egg yolk-glycerol extender, and may be considered a microbiologically safer option for storing and shipping of goat semen. A final assessment will require large-scale insemination trials. P133 Effect of pre-ovulatory luteinizing hormone surge on the mRNA expression of angiogeneic growth factors in the ovine ovary Chowdhury, WH 1 *, Scaramuzzi, RJ 2 , Wheeler-Jones, C 2 , Khalid, M 1 1Veterinary Clinical Sciences, The Royal Veterinary College, University of London, United Kingdom; 2 Veterinary Basic Sciences, The Royal Veterinary College, University of London, United Kingdom Introduction Angiogenic factors are essential for neo-vascularisation and development of ovarian follicles. Vascular Endothelial Growth Factor (VEGF) and the angiopoietins (Ang-1, Ang-2) are important and their expression in follicles differs with stage of the oestrous cycle. However, their physiological control in follicles is unclear. This study tests the hypothesis that the mRNA expression for VEGF, Ang- 1 and Ang-2 in antral follicles is regulated by pre-ovulatory LH surge. Methods Twenty eight ewes were made hypogonadotrphic with GnRH agonist (Buserelin) after which a combination of FSH and LH was administrated for normal development of follicles. This was followed by intravenous infusion of 0, 50, 100 or 200μg of LH over 4 or 8h as pre-ovulatory LH surge. Ewes were killed 8h after the LH infusion. The ovaries were serially section (10μm) at -20°C. All follicles ≥2.0mm in diameter were analysed for VEGF, Ang-1 and Ang-2 mRNA by in situ hybridization using ovine riboprobes. mRNA expression was quantified and the data were analysed using a mixed model ANOVA. Results Pre-ovulatory LH surge significantly increased (P
16 t h International Congress on Animal Reproduction Poster Abstracts 73 P135 Synchronization of oestrus and ovulation in non-seasonal West African ewes treated with cloprostenol during early stages of luteal development Contreras-Solis, I 1 *; Vasquez, B 2 ; Diaz, T 1 ; Lopez-Sebastian, A 3 ; Gonzalez- Bulnes, A 3 1Facultad de Ciencias Veterinarias, Universidad Central de Venezuela, Venezuela; 2 Instituto Nacional de Investigaciones Agrícolas, Venezuela; 3Instituto Nacional de Investigación y Tecnología Agraria y Alimentaria, Spain Cloprostenol (prostaglandin F2 alpha analogue) has been commonly used to synchronise oestrus in non-seasonal and seasonal ewes during the breeding season. The most usual protocol consist of two injections 9 to 11 days apart; however, the most recent studies indicate the possibility of increasing efficiency of the treatment by applying cloprostenol in early luteal phase (Contreras-Solis et al., in press). On the other hand, information about efficacy of cloprostenol injection during early stages of luteal development is controversial. Therefore, the aim of this study was to assess if a low dose of cloprostenol (43.75 µg; Planate®, Schering Plough) is capable to induce synchronization of oestrus and ovulation during the early stage of luteal growth in non-seasonal hair sheep. Twenty-four adult West African ewes were randomly assigned to three groups treated with cloprostenol at Day 3 (D3 n = 8), 5 (D5 n = 8) and 7 (D7 n = 8) after ovulation. Transrectal ultrasonographies were performed at Day 0 (day of injection), 1 and 10 to detect the presence of luteal tissue and luteolysis. Oestrous behaviour was detected every 4h from 20h after treatment using a trained teaser ram, whilst ovulation was detected by ultrasonographic scanning every 4 h from 16 h of the onset of oestrus. Assessment of luteal function was performed by determining plasma progesterone levels by radioimmunoassay. Seven out of eight ewes (87.5%) from D3 and D5 groups and all ewes from D7 group showed functional corpora lutea at timing of injection (P < 0.001; 1.4 ± 0.3, 3.0 ± 0.4 and 3.9 ± 0.3 ng/mL of mean plasma progesterone concentration, respectively). Cloprostenol was effective to induce luteolysis and oestrus in all ewes. Oestrous activity was observed at 28.6 ± 3.3, 34.3 ± 2.1 and 36.5 ± 3.2 h after treatment for D3, D5 and D7 groups, respectively. Onset of ovulation was detected at 51.4 ± 1.4, 52.0 ± 1.8 and 55.0 ± 1.7 h for D3, D5 and D7, respectively; with oestrusovulation intervals of 22.9 ± 3.1, 18.4 ± 2.7 and 18.5 ± 3.4 h. The number and function of corpora lutea were also similar among groups (D3: 1.9 ± 0.2 and 5.6 ± 0.5 ng/mL; D5: 1.7 ± 0.2 and 5.6 ± 0.4 ng/mL; D7: 1.4 ± 0.2 and 4.5 ± 0.7 ng/mL). Thus, current study indicates that cloprostenol is effective to induce luteolysis, oestrus, ovulation and corpora lutea with normal life span in hair sheep when is administered from Day 3 after ovulation. Funded by FONACIT S1-2002000413 and CDCH-UCV. Contreras- Solis et al., Anim. Reprod. Sci. In press. P136 Conservation in refrigeration of the ovine semen Córdova, A 1 *; Cortés, S 1 ; Córdova, MS 2 ; Córdova, CA 3 ; Guerra, JE 4 ; Tapia, B 5 1Departamento de Producción Agrícola y Animal, Ecodesarrollo de la Producción Animal, Universidad Autónoma Metropolitana-Xochimilco, Cuerpo Académico: Salud y Bienestar Animal. Calz. del Hueso 1100 Col. Villa Quietud CPP. 04960, México, D.F.(Córdova A. aci57@prodigy.net.mx); 2Laborarotorios Brovel, S.A.de C.V., Mexico; 3 Becario de CONACYT-México. Estudiante de Doctorado. Universidad de León, España; 4 Facultad de Agronomía. Universidad Autónoma de Sinaloa, México; 5 Estudiante de Doctorado. Facultad de Veterinaria. Universidad Complutense de Madrid, España Introduction The preservation of semen allows the use of genetic resources for long term, so that the cooling sheep sperm lets more time preserve the ability of sperm fertilization after obtaining an ejaculate. The rate of fertilization is used when cooled semen has a fluctuation between 45 and 65%, when using fresh semen this parameter is about 84%, this is because sperm got harm at the plasma membrane, with the unavoidable reduction of motility and acrosome damage during the process of preservation, which encourages further works on the preservation of semen in the cooling of this kind. Objective Evaluate the effect of the conservation in refrigeration of the ovine semen on the spermatic quality: motility, viability and acrosome integrity (NAR). Methods Four lambs race English Suffolk 2 years were used, obtaining 3 to 4 ejaculated per week and 24 samples were collected from ejaculate with artificial vagina. The progresses of motility, viability, pH, NAR were evaluated. Extenders 1 (Rangel, 1985); extenders 2 (Tris-fructose egg-yolk ram semen - Salamon, 1990) and extensor 3 basis triladyl were used in proportion of 1:3; semen keep on chilling at 4 ° C for 24 hours and the variables were assessed at 0, 2, 4 and 24 hours. Results There was found that the pH remains unchanged during the first 24 hours in the three extenders. The motility and viability were better with diluter than triladyl during the experiment. However, the NAR had a better behave with the extenders 1 and 2. The results were analyzed using descriptive statistics, which showed averages for each variable and there was not found difference statistically significant between the three tests. Conclusion The diluter with the help of triladyl can represent an alternative for the conservation of the ovine semen in refrigeration. P137 Effect of glycerol concentration on the motility and viability of cooled-stored ram semen Crespilho, AC 1 *; Papa, FO 1 ; Dell’Aqua Jr., JA 1 ; Zahn, FS 1 ; Martins Jr, A 2 ; Araujo, GHM 1 ; Oba, E 1 1 Department of Animal Reproduction and Veterinary Radiology, São Paulo State University, Botucatu, Brazil; 2 São Paulo State University, Araçatuba, Brazil Several studies demonstrate the efficiency of glycerol as a sperm membrane protector in many species during cryopreservation procedures. However, few studies investigated its effect on the maintenance of ram sperm viability during refrigeration. The aim of the present study was to evaluate the effect of the addition of different concentrations of glycerol on ram sperm viability under 5°C refrigeration for until 120 hours. One ejaculate of five Santa Inês rams, obtained by electroejaculation, were used. Each ejaculate was divided into 4 equal samples, diluted to a concentration of 10× 10 6 sperms/mL in TRIS (Tris(hydroximethyl)- aminomethane, sodium citrate and fructose) extender without or with 1.5, 3.0 or 6.0% glycerol (G1, G2, G3 and G4, respectively). Samples were packed in 0.5mL straws and cooled in a Minitüb® adjustable refrigerator at -0.25ºC/min until 5ºC. Samples were maintained at 5ºC and were evaluated at moments 0, 24, 48, 72, 96 and 120 hours for sperm motion characteristics by CASA (IVOS – 12) and for integrity of plasma and acrosomal membranes and mitochondrial potential by epifluorescence with propidium iodide, FITC-PSA and JC-1 probes. Data were analyzed by a Student’s t-test and a ‘t’ test with repeated measures to compare each group in a function of time of refrigeration and to evaluate the efficiency of each treatment to maintain sperm viability at 120 hours of refrigeration. There were no significant differences on parameters of motility and integrity when different treatments were compared at moments 0, 24, 48, 72, 96 and 120 hours. However, a significant decrease in total motility (MOT, P=0.018), sperm beat cross frequency (BCF, P= 0.0292) and acrosome integrity (P=0.0088) was observed in G1 samples at 120 hours of refrigeration. The addition of glycerol enhanced the preservation of MOT during the period of refrigeration and this effect was directly correlated to the concentration of cryoprotectant in the extender (G2, P
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Reproduction in Domestic Animals - Facultad de Ciencias Veterinarias

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