Effects of egg yolk during the freezing step of cryopreservation on the viability of goat spermatozoa.
Four experiments were conducted to investigate the effects of egg yolk during the freezing step of cryopreservation (namely, the process except for the cooling step), on the viability of goat spermatozoa. The effects of egg yolk on sperm motility and acrosome integrity during the freezing step were investigated in Experiment 1. Spermatozoa diluted with Tris-citric acid-glucose (TCG) solution containing 20% (v/v) egg yolk were cooled to 5 degrees C, washed, and then frozen in TCG with egg yolk (TCG-Y), TCG without egg yolk (TGG-NY), 0.370 M trehalose with egg yolk (TH-Y), or trehalose without egg yolk (TH-NY). All extenders contained glycerol. In frozen-thawed spermatozoa, the inclusion of egg yolk in the freezing extenders increased (P<0.05) percentages of motile sperm, progressively motile sperm, and the recovery rate (ratio of post-thaw to pre-freeze values), but decreased (P<0.05) acrosomal integrity. Moreover, extenders with trehalose had better (P<0.05) post-thaw sperm viability. In Experiment 2, the effects of egg yolk on acrosome status before and after freezing were studied. Egg yolk significantly decreased the proportion of intact acrosomes before freezing, leading to fewer (P<0.05) intact acrosomes post-thaw and lower (P<0.05) recovery rates for intact acrosomes. In Experiment 3, including sodium dodecyl sulfate (SDS) in a diluent containing egg yolk tended to preserve the acrosome compared with the egg yolk containing diluent free of SDS, however, spermatozoa had a lower (P<0.05) proportion of intact acrosomes than those in a yolk-free diluent. However, after cooling, spermatozoa were diluted with a glycerolated extender containing egg yolk. Therefore, the objective of Experiment 4 was to explore whether the egg yolk or glycerol was responsible for the reduced intact acrosome percentage. In this experiment, after cooling and washing the spermatozoa were diluted in TCG with glycerol and/or egg yolk. The combination of glycerol and egg yolk in the extender reduced (P<0.05) the proportion of intact acrosomes compared with egg yolk or glycerol alone. In conclusion, the inclusion of egg yolk significantly improved sperm motility, indicating its beneficial effects during the freezing step of cryopreservation; trehalose appeared to synergistically increase its cryoprotective effects. Furthermore, although neither glycerol nor egg yolk per se affected the proportion of intact acrosomes, the combination of the two significantly reduced the proportion of acrosome-intact spermatozoa.
Laboratory of Animal Reproduction, Graduate School of Biosphere Sciences, Hiroshima University, Higashi Hiroshima, Hiroshima 739-8528, Japan.
Eiman M-E
EM
Aboagla
T
T
Terada
D000177:Acrosome / Q000648:ultrastructure; D000818:Animals; D002470:Cell Survival; D019343:Citric Acid; D015925:Cryopreservation / Q000379:methods / Q000662:veterinary*; D004530:Egg Yolk; D005947:Glucose; D005990:Glycerol; D006041:Goats; D008297:Male; D012662:Semen Preservation / Q000379:methods / Q000662:veterinary*; D012996:Solutions; D013081:Sperm Motility; D013094:Spermatozoa / Q000502:physiology* / Q000648:ultrastructure; D014199:Trehalose; D014325:Tromethamine
D012996:Solutions; D014325:Tromethamine; D019343:Citric Acid; D014199:Trehalose; D005947:Glucose; D005990:Glycerol