Glutathione-supplemented tris-citric acid extender improves the post-thaw quality and in vivo fertility of buffalo (Bubalus bubalis) bull spermatozoa

Abstract

In this study we evaluated the effects of semen extender supplementation with different concentrations of glutathione (GSH) on buffalo (Bubalus bubalis) bull sperm motility, plasma membrane integrity, viability and DNA integrity as well as in vivo fertility. Semen from three Nili-Ravi buffalo bulls was collected, and qualified semen ejaculates (n=18) were split into five aliquots for dilution (37°C; 50×106spermatozoaml−1) with experimental tris-citric acid extender containing 0, 0.5, 1.0, 1.5 or 2.0mM GSH. Extended semen was cooled to 4°C, equilibrated and filled in French straws. The straws were kept on liquid nitrogen vapors (5cm above the LN2 level) for 10min and plunged in liquid nitrogen for storage. Sperm motility (%), plasma membrane integrity (%), viability (%) and DNA integrity (%) were assessed at 0, 2 and 4h post-thawing (37°C). Extender supplementation with GSH (0.5, 1.0, 1.5 and 2.0mM) increased sperm motility, plasma membrane integrity and viability in a dose dependent manner. Sperm DNA integrity was higher (p<0.05) in all experimental extenders containing GSH when compared to the control extender (0mM GSH). The in vivo fertility rate of cryopreserved buffalo bull (n=2) spermatozoa was higher (p<0.05) in extender containing 2.0mM GSH compared to that of control. In summary, tris-citric acid extender supplemented with glutathione improved the freezability of buffalo bull spermatozoa in a dose dependant manner. Moreover, the addition of 2.0mM GSH to the extender enhanced the in vivo fertility of buffalo (Bubalus bubalis) bull spermatozoa.