The objective was to evaluate the suitability of using natural or lyophilized low density lipoproteins (LDL), in lieu of whole egg yolk, in extenders for cryopreserving ram semen. Once extragonadal sperm reserves were depleted in 10 fertile Santa Inês cross rams, two ejaculates per ram were collected for cryopreservation. Nine extenders were used: Tris-16% egg yolk extender with 5% glycerol as a control (T1), and substitution of whole egg yolk with 8, 12, 16 or 20% natural LDL (T2-T5, respectively), or with 8, 12, 16, or 20% lyophilized LDL (T6-T9). Semen was diluted to 100 × 10(6) sperm/mL, packaged into 0.25 mL straws, cooled, held at 5 °C for 3 h, and then frozen in liquid nitrogen vapor. Immediately after thawing (37 °C for 30 s), sperm total and progressive motility, and kinetic parameters were analyzed with computer assisted semen analysis (CASA). Percentage of sperm with plasma membrane functional integrity was assessed by the hypoosmotic swelling test (HOST), sperm membrane physical integrity with propidium iodide (PI), and acrosome integrity with FITC-PSA using an epifluorescent microscope. For all sperm end points, there was no difference between the control and natural LDL treatments (P > 0.05): total motility (T1: 20.9 ± 11.9 and average of T2-T5: 25.9 ± 13.6%; mean ± SD), progressive motility (T1: 6.6 ± 4.2 and average of T2-T5: 11.7 ± 7.5%), HOST(+) (T1: 23.7 ± 6.9 and average of T2-T5: 23.2 ± 8.7 %) and PI(-)/PSA(-) (T1: 13.8 ± 7.8 and average of T2-T5: 18.1 ± 7.8%). However, lyophilization was apparently unable to preserve the protective function of LDL; every sperm end point was significantly worse than in the control and natural LDL groups. We concluded that natural LDL was appropriate for cryopreserving ram semen, as it yielded results similar to those obtained with whole egg yolk.

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The study aimed at testing the effectiveness of dimethylformamide, alone or combined with glycerol, as cryoprotectant for freezing ram semen. Ejaculates from nine rams were cryopreserved in Tris-based extenders, containing 5% of glycerol, association of dimethylformamide with glycerol, in four proportions achieving 5% of cryoprotectors in the media and pure dimethylformamide (2, 3, 4 and 5%) in replacement to glycerol. The samples were diluted to 100 × 10(6) sptz/ml and stored in 0.25-ml straws in liquid nitrogen. After thawing (37 °C for 30 s), motility was preserved better by the extender containing 5% of glycerol (p < 0.05). The extenders containing pure dimethylformamide, or more than 2% in combination with glycerol, provided sperm motilities close to zero. Plasma and acrosomal membrane integrity were preserved better (p < 0.05) in the extender containing 5% glycerol. It can be concluded that dimethylformamide, alone or combined with glycerol, has no beneficial effects on ovine semen cryopreservation.

-The objective of this work was to evaluate the effects of hormonal synchronization protocols, associated or not with follicular development stimulation, on the recovery of oocytes and on in vitro production of Bos indicus and B. taurus embryos, in different seasons. Ultrasound-guided follicular aspirations (n=237) were performed without pre-treatment (G1, control group) and after follicular wave synchronization (G2), or after follicular wave synchronization and follicle growth induction (G3). Bos indicus produced more oocytes and embryos than B. taurus (18.7±0.9 vs. 11.9±0.6 oocytes and 4.8±0.3 vs. 2.1±0.2 embryos). On average, oocyte and embryo yields were higher in G3 than in G2, and both were greater than in G1, which lead to a higher conversion of oocytes to embryos in these treatments. The hot or the cold season did not affect the B. indicus outcomes, whereas, in B. taurus, both oocyte recovery and embryo production were higher in the cold season. Follicular wave synchronization improves ovum pick-up and in vitro production of embryos in both cattle subspecies evaluated.Index terms: Bos indicus, Bos taurus, artificial insemination, follicle growth induction, follicular wave synchronization, FSH stimulation. Protocolos hormonais para produção in vitro de embriões de zebuínos e taurinosResumo -O objetivo deste trabalho foi avaliar os efeitos de protocolos hormonais de sincronização, associados ou não com o estímulo ao desenvolvimento folicular, na recuperação de oócitos e na produção in vitro de embriões de Bos indicus e B. taurus, em diferentes estações. As aspirações foliculares guiadas por ultrassom (n=237) foram realizadas sem pré-tratamento (G1, grupo controle) e após sincronização da onda folicular (G2), ou após sincronização da onda folicular e estímulo do desenvolvimento folicular (G3). Bos indicus produziu mais oócitos e embriões que B. taurus (18,7±0,9 vs. 11,9±0,6 oócitos e 4,8±0,3 vs. 2,1±0,2 embriões). Em média, as produções de oócitos e embriões foram maiores em G3 do que em G2, e ambas foram maiores que em G1, o que levou a uma maior conversão de oócitos em embriões nesses tratamentos. A estação quente ou fria não afetou os resultados de B. indicus, enquanto, em B. taurus, tanto a recuperação de oócitos quanto a produção de embriões foram maiores na estação fria. A sincronização da onda folicular melhora a aspiração folicular e a produção in vitro de embriões, em ambas as subespécies de bovinos avaliadas.Termos para indexação: Bos indicus, Bos taurus, inseminação artificial, indução de crescimento folicular, sincronização da onda folicular, estimulação com FSH.

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ABSTRACT

The objective of the present study was to evaluate the effect of two diets with different energy levels and two genetic groups (3/4 and 7/8 Holstein × Gir (HG) cows) on the metabolic and hormonal changes and on the production profile of oocytes and embryos in dairy cows during the early postpartum period. The concentrations of oestrogen, progesterone, glucose, insulin, insulin-like growth factor, urea and non-esterified fatty acids in follicular fluid, as well as blood plasma concentrations of glucose, insulin, urea and non-esterified fatty acids, were evaluated. Oocyte collection was performed every 14 days after parturition. After classification, the oocytes were submitted to in vitro embryo production and cleavage, and blastocyst rates were evaluated. Five days after oocyte collection, the dominant follicle was measured and punctured for follicular fluid retrieval. The high-energy diet increased plasmatic insulin and glucose. The 3/4 HG cows presented a higher plasmatic concentration of insulin, glucose and urea. The hormonal and metabolic changes in plasma were not observed in the follicular fluid. The follicular fluid concentration of IGF-I was increased in cows fed the high-energy diet as well as in the 3/4 HG cows. A higher number of total and viable oocytes was recovered in the 3/4 HG cows, but the 7/8 HG cows had a higher cleavage rate. In conclusion, the high-energy diet was more efficient in maintaining the energy status of crossbred cows, as evidenced by their plasma metabolites and follicular fluid, and 3/4 HG cows were more efficient than 7/8 HG cows at producing oocytes in the early postpartum period.