Periódicos Brasileiros em Medicina Veterinária e Zootecnia

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Negative pressure in the pre-freezing of Ram semen

Casali, RenataissaarissaarissaarissaCristina Arcego, CarolineDesconsi Mozzaquatro, FabricioMezzalira, Alceu

Background: The effect of controlled stress on cells has been widely studied and there is strong evidence that positive pressure improves the survival of cryopreserved gametes and embryos. Positive pressure resulted in higher motility of frozen boar semen, and increased bull semen cryotolerance. Looking for similar effects, we developed a piece of equipment (Nitrocooler) to apply negative pressure. The Nitrocooler increased the cryotolerance in bovine IVP blastocysts. Nevertheless, until now, negative pressure has not been tested in sperm cells. This study aimed to determine the effects and the best negative pressure intensity to apply in pre-freezing ram semen.Material, Methods & Results: The ram semen obtained with artificial vagina was immediately diluted (1 + 1.5) in Trisegg yolk, and fractioned into 4 experimental groups, Control: Without treatment; P200: Submission to 200 mBar negative pressure; P500: Submission 500 mBar negative pressure; P800: Submission to 800 mBar negative pressure. Then, Tris-yolk glycerol was added to all groups to obtain a dilution of 1 + 3, and a sperm concentration of 180 million per straw. The cooling and freezing procedure was identical for all groups. After thaw, progressive motility (PM) was assessed at zero, 1, 2 and 3 h of TRT and after Percoll selection (PMPP). Acrosome integrity (ACI) was assessed by FITC PNA staining; plasmatic mem

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