Avaliação "in vitro" da adição fracionada da dimetilformamida na criopreservação de sêmen equino

AUTOR(ES)
DATA DE PUBLICAÇÃO

2005

RESUMO

The aim of this research was to evaluate the efficacy of the manner of the cryoprotectant addition, one or multiple steps, and the cooling rate, on equine semen cryopreservation. The lesion intensity of the spermatozoa, during the process of the cryopreservation, was also studied. One ejaculate of nine stallions was used to test the modified INRA 82 with 5% of dimetyl formamide with the following treatments: addition of the dimethyl formamide in one step and fast cooling rate (T1- control), addition of the dimethyl formamide in multiple steps and fast cooling rate (T2), addition of the dimethyl formamide in one step and slow cooling rate (T3), addition of the dimethyl formamide in multiple steps and slow cooling rate (T4). After centrifugation of ejaculates in INRA 82 extender, sperm pellets were diluted with final extenders to reach the concentration of 100X106 sperm cells per ml. Semen was frozen three centimeters above the nitrogen level, during ten minutes, in 0.5 ml straws. Thawing of samples was done at 52°C for ten seconds followed by immersion of the straw in a water bath at 37°C for thirty seconds. Immediately pos dimethyl formamide addition, total and progressive motility and sperm vigor were evaluated under light microscope (400X). The functional sperm membrane integrity was evaluated by the hypoosmotic swelling test. No differences between treatments were observed, before the cooling of the straw. Immediately post thaw, total and progressive motility and sperm vigor were evaluated under light microscope (400X). Functional and structural sperm membrane integrity were evaluated by the hypoosmotic swelling test and fluorescent dyes, carboxyfluorescein diacetate and propidium iodide, respectively. The spermatozoa was also evaluated in the temperature resistance test. The multiple steps addition of the cryoprotectants was better than the one step addition in all parameters valued, independently from the cooling rate. The slow cooling rate was better than the fast cooling rate in all parameters valued, independently from the manner of the dimethyl formamide addition. It may be concluded that the association of the multiple steps addition of the dimethyl formamide with the slow cooling rate is the best protocol to freeze the equine semen and that the lesions caused by osmotic and cold stress may occur during the cooling and freezing process

ASSUNTO(S)

sêmen análise teses eqüino reprodução teses criopreservação teses veterinária teses

ACESSO AO ARTIGO

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