Cholesterol-cyclodextrin complex as a replacement for egg yolk in bull semen extender: sperm characteristics post-thawing and in vivo fertility

Semen extenders protect sperm during initial cooling and cryopreservation processes. Lipoproteins, commonly of animal origin, such as egg yolk, are constituents of conventional semen extenders that protect sperm from initial cold shock during the transition from room- to refrigeration-temperature. Disadvantages of the use of animal products in semen extenders, however, are the undefined composition of these products and potential as a source for microbial contamination. There is a need to find an alternative to egg yolk in semen extenders with definable constituents that effectively prevent cold shock and minimize the potential of microbial contamination. Cholesterol has an essential function in maintaining membrane stability and resisting membrane phase transition when there is storage at lesser temperatures by preventing aggregation of membrane phospholipids and domain reorganizations during cooling, and by minimizing disruption of lipid-protein interactions. Cyclodextrins, cyclic oligosaccharides with a hydrophobic center and hydrophilic exterior, can be sources for cholesterol transfer into the cell membrane. Supplementation of sperm plasma membrane with cholesterol-cyclodextrin (CC), in addition to egg yolk extender, resulted in improved post-thaw quality in boar, ram, and bull sperm. The objectives of the present study, therefore, were to determine the motion and structural characteristics of sperm post-thawing, and in vivo fertility when bull semen pre-treated with different concentrations of cholesterol-cyclodextrin complex before dilution in Tris-glycerol extender was used for AI, and to assess the efficacy of imposing a letrozole-based treatment regimen for synchronization of timing of ovarian functions among cows. Bull semen was frozen in either egg yolk or cholesterol-cyclodextrin complex (0.5, 1 or 2 mg/mL semen) in Tris-glycerol (TG) extender. Sperm motion characteristics examined, and plasma membrane and acrosome integrity were similar for all extenders. Semen treated with cholesterol-cyclodextrin complex (2 mg/mL semen) followed by dilution in TG extender resulted in lesser pregnancy rates (P < 0.05). There was a pregnancy rate of as great as 56% when sperm cryopreserved in 0.5 mg/mL cholesterol-cyclodextrin Tris-glycerol extender. Results indicate cholesterol-cyclodextrin Tris-glycerol extender, with a chemically defined composition and without inclusion of egg yolk, may be used to cryopreserve bull sperm with acceptable pregnancy rates, using fixed-time artificial insemination.