Cholesterol addition aids the cryopreservation of dromedary camel (<I>Camelus dromedarius</I>) spermatozoa

Abstract

The cryopreservation of dromedary camel (Camelus dromedarius) sperm has proved challenging with little success reported. Yet the routine application of artificial insemination with frozen semen would assist the flow of valuable genetic material nationally and internationally. The current study sought to examine the effects of cholesterol (cholesterol-loaded cyclodextrin; CLC) pre-loading on camel sperm cryosurvival. Ejaculates (n = 3 males; 3 ejaculates/male) were collected using an artificial vagina during the breeding season and extended in Hepes- buffered TALP and allowed to liquefy in the presence of papain ( 0.1 mg/mL) prior to removal of the seminal plasma by centrifugation. Sperm pellets were re-suspended (120 million/mL) in fresh TALP and incubated (15 min; 37 °C) with 0, 1.5 or 4.5 mg CLC/mL. Sperm suspensions were then centrifuged and reconstituted in INRA-96® containing 20% (v:v) egg yolk and 2.5% (v:v) methylformamide, loaded in 0.5 mL plastic straws, sealed and cooled for 20 min at 4°C. Straws were frozen over liquid nitrogen (4 cm above liquid; 15 min), plunged and stored. Sperm motility, forward progressive status and acrosomal integrity were recorded at zero and 3 h post thaw and compared with these same parameters pre-freeze. Aliquots also were stained with chlortetracycline hydrochloride to assess spontaneous sperm capacitation status pre- and post-thaw. Pre-treatment with CLC (1.5 mg/mL and 4.5 mg/mL) enhanced cryosurvival (P 0.05) effect. In summary, dromedary camel sperm benefit from exposure to CLC prior to cryopreservation; this may facilitate the routine collection and storage of sperm from this species.