Efecto de la sacarosa, trehalosa y rafinosa en la vitrificación de espermatozoides equinos

Abstract

This research evaluated the effect of different non-penetrating cryoprotective agents (NCPAs) in the vitrification of equine spermatozoa. To achieve this, 16 ejaculates from four Spanish horses were collected using an artificial vagina. Each ejaculate was diluted with the Botusemen-Gold® diluent + 1% BSA and divided into four aliquots to form four respective treatments based on conventional freezing (CF, control) and vitrification with NCPAs: 100 mM sucrose (VIT -Su); 100 mM trehalose (VIT-Tr); and 50 mM raffinose (VIT-Raf). CF was performed using liquid nitrogen vapors (LN2), while vitrifications were carried out by submerging 30 µL drops directly into LN2. the results of sperm samples showed that the CF treatment produced higher (P < 0.01) motility, kinematics, and membrane integrity compared to all vitrification treatments. However, all three vitrification treatments exhibited higher values (P < 0.05) for straight-line velocity, linearity, and oscillation than the CF treatment. Furthermore, VIT-Su and VIT-Ra treatments showed higher values (P<0.05) for total and progressive motility, as well as plasma and acrosomal membrane integrity, compared to the VIT-Tr treatment. Finally, all vitrification and CF treatments increased (P<0.05) the length and area of sperm heads compared to their fresh counterparts. In conclusion, despite conventional freezing resulting in higher sperm cryosurvival, vitrification with sucrose and raffinose produced greater motility and membrane integrity compared to trehalose, making it an attractive method for cryopreserving equine spermatozoa.