Congelação e vetrificação de espermatozóides epididimários de gato doméstico (Felis catus): estabelecimento de protocolos para felinos selvagens ameaçados de extinção.

Abstract

The objective of this study was to investigate the effect of different osmolarities of the Tris-citrate thinner plus glycerol or ethylene glycol on the freezing of epididymal sperm and to vitrify the tail of the epididymis, to recover viable sperm from domestic cats (Felis catus). Thus, the work was divided into two phases. Phase 1: The maintenance of sperm parameters in a tris-citrate thinner with osmolarities 275, 325, 375, 425, 475 and 525mOsm was evaluated, at times (T0 = 0, T1 = 30 and T2 = 60 min). And freezing using osmolarities 325 and 375 mOsm with 4% glycerol or 3% ethylene glycol, 6% in sperm from the tail of the domestic cat's epididymis. Phase 2: Develop a protocol for vitrification of the tail of the epididymis of domestic cats for the recovery of viable sperm, using glycerol and ethylene glycol 40% associated with 0.1M sucrose, using that of equilibrium vitrification with the surface vitrification technique solid (SSV). In Phase 1 among osmolarities, the motility of 325 mOsm did not differ statistically with epididymal fluid (fresh control) in the three times and at 375 mOsm in T0 and T1, and both did not present statistical differences between themselves and between times in all parameters spermatic. The use of 4% glycerol in a diluter with 375 mOsm was superior, showing motility of 25% ± 6, vigor 4, plasma membrane integrity of 48% ± 9 after freezing, did not show statistical differences with the cooling step and in morphology, no statistical differences were found between the two osmolarities in all stages of freezing. In Phase 2, Ethylene glycol treatment had better results for all sperm motility parameters (Motility, Vigor, CASA) compared to glycerol (P ˂ 0.05). There were no statistical differences in the parameters of plasma membrane and acrosome integrity (Trypan blue and Giemsa), plasma membrane integrity using also (fluorescent dyes Hoechst and propidium iodide) and in normal sperm morphology before and after vitrification between the experimental. The results of the present study showed that Tris-citrate with 325 and 375 mOsm between the tested osmolarities and post-freeze with 375 mOsm and glycerol 4% maintained the sperm parameters, and was successful in vitrifying the tail of the epididymis with 40% ethylene glycol associated with sucrose (0.1 M), of epididymal sperm from domestic cats.