Antagonist effect of DMSO on the cryoprotection ability of glycerol during cryopreservation of buffalo sperm

The objective of the present study was to investigate the synergistic effect of DMSO and glycerol added at various temperatures on the post-thaw quality of buffalo sperm. Pooled ejaculates from four Nili-Ravi buffalo bulls were divided into 18 aliquots and extended (1:10) in Tris–citric acid extender differing in glycerol:DMSO ratios (0:0, 0:1.5, 0:3; 3:0, 3:1.5, 3:3; and 6:0, 6:1.5, 6:3, respectively; %, v:v) either at 37 or 4 °C. Semen was packaged in 0.5 mL French straws and frozen in a programmable cell freezer. Thawing was performed at 37 °C for 50 s. Post-thaw motion characteristics, plasma membrane integrity and acrosome morphology of buffalo sperm were determined using computer-assisted semen analyzer (CASA), hypoosmotic swelling (HOS) assay and phase-contrast microscopy, respectively. Glycerol (6%) in extender yielded better post-thaw sperm motility, velocities (straight-line and average path), plasma membrane integrity, and normal acrosomes (P < 0.05). Post-thaw sperm motility and plasma membrane integrity declined in the presence of DMSO (P < 0.01). The addition of glycerol (6%) at 37 °C yielded better post-thaw sperm motility, plasma membrane integrity and velocities than addition at 4 °C (P < 0.05). In conclusion, glycerol is still an essential cryoprotectant for buffalo sperm. The addition of DMSO antagonized the cryoprotection ability of glycerol and reduced the post-thaw quality of buffalo sperm. Furthermore, 6% glycerol added at 37 °C, provided better cryoprotection to the motility apparatus and plasma membrane integrity of buffalo sperm.