Effects of glucose, methanol concentration, and time of equilibration on post-thaw sperm motility of rainbow trout semen

Abstract

The objective of this work was to develop a complex standardisation of cryopreservation procedure for rainbow trout semen. The specific objectives were to determine the effects of the final glucose concentration (0.11–0.23 M), final methanol concentration in the extender (0–20%), equilibration time (5–180 min), and post-thaw storage time of semen (0–180 min) on sperm motility parameters. The final glucose and methanol concentrations significantly affected sperm motility after thawing. The highest sperm motility (63–75%) was found for glucose concentrations ranging from 0.13 to 0.17 M and for 5.0 to 10.0% methanol, with the highest motility (70%) at 7.5% methanol. Concerning post-thaw storage equilibration times, sperm motility of the equilibrated semen was not affected by 120 min, while a significant decrease was recorded after 180 min of equilibration. The highest values of sperm motility immediately after thawing were observed in samples equilibrated for 5 to 120 min. The sperm motility after thawing was not significantly affected up to 60 min for samples equilibrated for 5, 15, and 30 min. However, for samples equilibrated for 60, 120, and 180 min, significant decreases were observed at 60 min after thawing. Our results indicate the importance of the standardisation of concentrations of cryoprotectants as well as cryopreservation equilibration time. The recommended cryopreservation procedure includes final sperm concentration (1.0 × 109 spermatozoa ml−1), as we determined previously, 0.15 M glucose, 7.5% methanol, 15 min equilibration time, and post-thaw storage up to 60 min. The developed standardised procedure has potential for the cryopreservation of rainbow trout semen for practical purposes.