Abstract The objective of this study was to evaluate the effects of the addition of different concentrations of ozone to quarter horse semen submitted to cryopreservation. Six ejaculates from four stallions were collected and were divided in four experimental groups: a control group (BotuCRIO® extender) and three other groups with BotuCRIO® ozonized at concentrations of 6, 8 and 12 μg of O3/mL. The semen samples were diluted (200 x 106 spermatozoa/mL), filled in straws and frozen. After thawing (37 ºC, 30s), the samples were evaluated at 0, 30 and 60 minutes of incubation regarding sperm kinetics by a computer-assisted sperm analysis (CASA), and plasma membrane integrity (PMI), acrosome integrity (ACi) and mitochondrial membrane potential (MMP) by fluorescent probes. There was a reduction in the kinetic parameters total motility (TM), progressive motility (PM), curvilinear velocity (VCL), straight line velocity (VSL) and average path velocity (VAP) in all groups during the thermoresistance test (TT), a pattern also found in PMI and MMP analyses (p<0.05). There was no difference (p>0.05) between the control and treatment (6, 8, and 12 μg of O3/mL) groups, in any of the evaluated times for the kinetic parameters TM, linearity (LIN), straightness (STR), wobble index (WOB), amplitude of lateral head displacement (ALH) and beat cross frequency (BCF). Regarding the VCL, VSL and VAP parameters, the group treated with 6 μg did not differ from the control or from 8 μg, but was higher than 12 μg at 30 and 60 minutes. ACi and PMI did not differ between groups (p>0.05), but PMI was lower in groups 8 μg and 12 μg compared to the control and 6 μg (p<0.05). It was concluded that the addition of ozone does not present beneficial effects for cryopreservation of equine semen at the concentrations used and decreases important parameters of fertility.

Abstract The objective of this study was to investigate the need of seminal plasma removal for short-term cooling of buck semen in soybean lecithin (SL) based extender. Each pool was divided equally, and one half was subjected to centrifugation to remove seminal plasma (SP-), while the other half remained with seminal plasma (SP+). Then, both SP+ and SP- samples were diluted in two SL extenders (extender A = 1% SL; extender B = 2% SL), cooled to 5ºC and stored for 48 hours. The sperm kinetics, evaluated by CASA, and plasma membrane integrity (PMI), acrosomal integrity (ACI) and high mitochondrial membrane potential (HMMP), evaluated by epifluorescence microscopy, were determined within five minutes after reaching 5°C (T0), as well as after 24 (T24) and 48 (T48) hours of storage. Interactions (seminal plasma vs. extender vs. time;) were observed for all variables assessed. Total and progressive motility and other variables of sperm kinetics decreased after 24 hours of cooling in the SP+ group, and after 48 hours of storage, these same variables were lower in SP+/B compared to SP-/B groups. Furthermore, SP+ reduced PMI (extender B, T48), HMMP (A and B extenders, T48) and ACI (extender A, T0) compared to SP- samples. The interactions between seminal plasma and soybean lecithin phospholipids seemed to occur in a time-dependent manner. It was concluded that the removal of seminal plasma improves the quality of goat semen that was cooled in a soybean lecithin-based extender, especially when using 2% soybean lecithin.

Com o objetivo de avaliar o efeito da adição dos antioxidantes Trolox e Glutationa reduzida (GSH) na viabilidade in vitro de espermatozoides criopreservados de cães, amostras de sêmen foram colhidas de animais das raças Buldogue Francês (n=1), Basset Hound (n=2) e Rottweiler (n=1), por meio de manipulação digital. A seguir, procedeu-se à avaliação macroscópica e microscópica e divisão do volume de sêmen em alíquotas, de acordo com os experimentos e grupos experimentais, utilizando-se diferentes concentrações de Trolox (200 e 300 µM/L) ou GSH (2 e 5 µM/L). As amostras foram envasadas em palhetas (0,25 mL), criopreservadas em máquina de congelação e armazenadas em nitrogênio líquido. Após descongelação a 37 ºC (60 seg) e incubação durante 60 minutos, constatou-se que os resultados de motilidade, vigor e espermatozoides com acrossomas íntegros evidenciaram diferença significativa (P<0,05) apenas entre os tempos de incubação, nos três experimentos. No Exp. 2, a adição de 2 e 5 µM/L de GSH (G2 e G3) determinou maior (P<0,05) porcentual de células sem estresse oxidativo. Concluiu-se que GSH, nas concentrações de 2 e 5 µM/L, pode ser adicionada ao diluente de criopreservação do sêmen de cães.

In order to evaluate the effect of adding Trolox and Glutathione reduced antioxidants on the in vitro viability of dog cryopreserved sperm, semen samples were harvested from animals of French Bulldog (n=1), Basset Hound (n=2) and Rottweiler (n=1) breeds, through digital manipulation. After macroscopic and microscopic evaluation, the volume of semen was divided in aliquots, according to the experiments and experimental groups, using different concentrations of Trolox (200 and 300 µM/L) or GSH (2 and 5 µM/L). The samples were packaged in straws (0.25 mL), cryopreserved in freezing machine and stored in liquid nitrogen. After thawing at 37ºC (60 sec) and incubation during 60 minutes, we verified that motility, vigor and sperm with intact acrosomes showed significant difference (P<0.05) only among incubation times, on the three experiments. In Exp. 2, the addition of 2 and 5 µM/L of GSH (G2 and G3) determined higher (P<0.05) percentage of sperm without oxidative stress. Therefore, we could conclude that GSH, on the concentration of 2 and 5 µM/L, can be added to diluent of dog semen cryopreservation.

Objetivando-se avaliar o efeito de diferentes concentrações dos crioprotetores etileno glicol ou acetamida na viabilidade in vitro de espermatozoides congelados de ovinos, foram utilizados seis pools de sêmen de quatro reprodutores mestiços da raça Santa Inês. Após aprovação, o pool dos ejaculados foi diluído em Tris-gema, acrescido de glicerol (G1=5%; controle), etileno glicol (G2=3%; G3=5%) ou acetamida (G4=2%; G5=7%), envasado em palhetas (0,25mL; 100x106 espermatozoides) e congelado. As amostras descongeladas (37oC/30 segundos) foram avaliadas quanto a motilidade progressiva (MP), vigor, integridade da membrana plasmática (iMP), potencial de membrana mitocondrial (PMM) e integridade do acrossoma (iAC). Constatou-se que a MP do G1 foi superior (P<0,05) à dos G3, G4 e G5, e que o vigor dos G1, G2 e G3 foi maior (P<0,05) do que o dos G4 e G5. A iMP do G1 foi superior (P<0,05) à dos G2, G3, G4 e G5, assim como os G2 e G3 foram superiores (P<0,05) a G4 e G5. O PMM e a iAC não diferiram (P>0,05) entre os grupos. Conclui-se que o etileno glicol (3 e 5%) e a acetamida (2 e 7%) não são eficazes para a proteção dos espermatozoides ovinos submetidos à congelação.

Aiming to evaluate the effect of different concentrations of ethylene glycol or acetamide cryoprotectants on in vitro viability of ram frozen spermatozoa, were used six semen-pools from four Santa Inês crossbred males. After approval, the ejaculate-pool was diluted in Tris egg-yolk plus glycerol (G1=5%; control), ethylene glycol (3%=G2; G3=5%) or acetamide (G4=2%; G5=7%), loaded in straws (0.25mL; 100x106 sperm) and frozeng. Thawed samples (37oC/30 seconds) were evaluated for progressive motility (PM), vigor, plasma membrane integrity (PMi), mitochondrial membrane potential (MMP) and acrosome integrity (ACi). It was found that the PM of G1 was higher (P<0.05) than G3, G4 and G5, and that the vigor of G1, G2 and G3 was higher (P<0.05) than G4 and G5. The PMi of G1 was higher (P<0.05) than G2, G3, G4 and G5, as well as G2 and G3 (P<0.05) where high than G4 and G5. MMP and ACi were not different (P>0.05) among groups. It is concluded that ethylene glycol (3 and 5%) and acetamide (2 and 7%) don't are effectives in protecting ram spermatozoa subjected to freezing.

Visando avaliar o efeito da adição de glutationa reduzida (GSH) ao diluente de congelação de sêmen caprino à base de leite desnatado, utilizou-se sêmen de cinco reprodutores Boer. Após colheita e avaliação, procedeu-se à formação do pool dos ejaculados e diluição em leite desnatado e glicerol 7%, acrescido de antioxidantes: G1) Controle; G2) GSH 2mM mL-1; G3) GSH 5mM mL-1 e G4) GSH 7mM mL-1. As amostras foram congeladas em palhetas (0,25mL) e armazenadas a -196°C. Após descongelação, avaliou-se a integridade de membrana plasmática (iMP) e acrossomal (iAc), potencial de membrana mitocondrial (PMM), cinética e ultraestrutura. Os grupos Controle e GSH (2, 5 e 7mM mL-1) não diferiram (P>0,05) em iMP, iAc, PMM e cinética. Na análise ultraestrutural, os porcentuais de membrana plasmática (cabeça e cauda) e acrossoma íntegros não diferiram (P>0,05) entre grupos. Todavia, o grupo Controle apresentou maior porcentual (P<0,05) de gametas com axonema íntegros do que os de GSH (2, 5 e 7mM mL-1). Maior porcentagem (P<0,05) de espermatozoides com mitocôndrias íntegras foi observada no grupo Controle do que nos de GSH (5 e 7 mM mL-1). Conclui-se que a adição de GSH (2, 5 e 7mM mL-1) em diluente de congelação de sêmen caprino, à base de leite desnatado, não preserva a integridade dos espermatozoides.

Aiming to evaluate in vitro effect of different concentrations of glutathione reduced (GSH) in skimmed-milk and glycerol 7% it was used semen from five Boer bucks. After collect and evaluation, a pool of samples was diluted in skimmed-milk and glycerol 7% plus antioxidant: G1) Control; G2) GSH 2mM mL-1; G3) GSH 5mM mL-1 and G4) GSH 7mM mL-1. Samples were frozen in straws (0.25mL) and stored at -196°C. After thawing, samples were subjected to integrity of the plasma membrane (iMP) and acrosomal (iAc), mitochondrial membrane potential (MMP), kinematic and ultrastructure analysis. Control and GSH (2, 5 and 7mM mL-1) groups did no differ (P>0.05) in iMP, iAc, PMM and kinematic parameters. In the ultrastructural analysis, percentages of acrosome and plasma membrane (tail and head region) intact did not differ (P>0.05) between groups. However, Control group had higher percentage (P<0.05) of gametes with intact axonemes than those of GSH (2, 5 and 7mM mL-1) groups. Higher percentage (P<0.05) of sperms with intact mitochondrias were observed on Control group than those of GSH (5 and 7mM mL-1). It can be concluded that the GSH (2, 5 and 7mM mL-1) addition in skimmed-milk diluent to freeze goat semen did not preserve sperm integrity.

Prolapso de reto é afecção comum em ovinos de cauda curta. Neste trabalho relata-se a técnica de colopexia para redução de prolapso retal em trinta ovinos da raça Dorper, dos quais, três vieram a óbito no período pós-operatório e três tiveram que ser sacrificados, pois além de apresentarem recidiva, um deles era idoso, e os outros três por se encontrarem bastante debilitados. Aos 15 dias após a cirurgia, cinco animais apresentaram recidiva do prolapso, sendo a colopexia refeita em três deles tendo bom resultado em apenas um, e os outros dois foram sacrificados. Aos 30 dias de pós-operatório um animal apresentou prolapso retal, os outros dezenove (63,3%) estavam em adequado estado físico. A realização de colopexia é uma alternativa para o tratamento de prolapso retal em ovelhas da raça Dorper, porém recidivas e complicações são comuns.

Rectal prolapse is a common affection in lambs of short tail. This study aimed at reporting the colopexy to reduce the rectal prolapse in Dorper lambs. Thirty animals were submitted to surgery and three of them died in the postoperative period. Three animals had to be sacrificed, because they have shown prolapse recurrence (one of them was old, and the other three were in a strong debilitated state). At 15 days after the surgery, five animals showed prolapse recurrence and the colopexy was performed again in three having good results in only one, the other two were sacrificed. At the 30 postoperative days, an animal showed rectal prolapse signals, the other nineteen animals (63,3%) were healthy. The colopexy use is an option to rectal prolapse treatment in Dorper lambs, although, recurrences and complications are expected.