Comparison of Two Different Extenders for Cryopreservation of Epididymal Dog Sperm

Abstract

ContentsThe collection of epididymal sperm is an option for preservation of germplasm of genetically superior animals that need to be orchiectomized or have died. The extender type used to freeze sperm is important to avoid spermatozoal membrane damage and to preserve semen quality after cryopreservation. The objective of this study was to verify the effects of a commercial bovine extender (Bovimix ® ; Nutricell, Campinas) and a traditional TRIS-citric acidglucose-egg yolk-7% glycerol extender on cryopreservation of canine epididymal sperm. The testes of 13 adult dogs were kept at 5°C for 24 h in saline solution, and epididymal sperm was recovered in Ringers solution without lactate and were evaluated for motility. Samples with ! 80% motility were pooled and then divided before dilution and packaging in 0.5 ml plastic straws, equilibration at 4°C for 1 h, freezing in nitrogen vapour for 20 min and storing at À196°C. The straws were thawed at 56°C for 10 s and were evaluated for motility by computer assisted analysis (CASA). The semen parameters, sperm movement index, linearity, total motility and rapid progressive motility were statistically higher in Bovimix ® than TRIS. In contrast, amplitude of lateral head displacement, slow sperm and static sperm were lower in Bovimix ® . Despite the high percentage of sperm defects in epididymal cells, regardless of the extender, we concluded that Bovimix ® is a viable alternative for the freezing of canine epididymal sperm. IntroductionDomestic dogs are not only companions but also excellent experimental models because of the similarity of the reproductive physiology with the wild species and humans. Obtaining semen directly from the epididymal tails and vas deferens is a technique that has been frequently used for the purposes of assisted reproduction (Martins et al. 2009). The process of cryopreservation may cause irreversible damage to the sperm membrane interfering with its viability. During the cryopreservation process, sperm cells are exposed to many stressors, which may be linked to thermal shock during cooling of the semen, the formation of intracellular ice crystals or osmotic shock during freezing and thawing or stress and action of cryoprotectants (Watson 2000). Therefore, choosing an extender with optimum properties for preserving sperm morphology and function may be the key to successful semen freezing. The aim of this study was to evaluate the effect on sperm viability using two different extenders, a commercial bovine extender (Bovimix ® ; Nutricell Ltda,Campinas, Brazil) and the Tris extender (Martins 2005) to freeze the epididymal spermatozoa. Materials and MethodsEpididymides were obtained from 13 mixed-breed dogs (age range 2-8 years, body weight <15 kg) by elective orchiectomy. After surgery, the testicles were kept at 5°C for 24 h in saline solution. After that time, the caudal portion of each epididymis and approximately 1.0 cm of the transition of epididymal tail into the deferential duct were squeezed into a Petri dish containing Ringers solution w...

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