title: The effect of cooling to different subzero temperatures on dog sperm cryosurvival.
abstract: The objective was to assess the effect of cooling to different subzero temperatures around ice formation (-5°C) on dog sperm cryosurvival and plasma membrane fluidity. Semen was centrifuged, and sperm were resuspended in a Tris-egg yolk medium (3% glycerol). Diluted sperm were cooled from 22 to 5°C, and then, a Tris-egg yolk medium containing 7% glycerol was added (final concentration of 5% glycerol and 200 × 106cells/ml). Sperm were packaged in 0.5-ml plastic straws, and equilibration was done 16 hr at 5°C before freezing. I. Straws (n = 47) at 5°C were exposed to nitrogen vapours to determine the freezing point. II. Other straws (from different ejaculates) processed as mentioned, were further cooled to -3, -5 or -7°C and immediately rewarmed in a water bath at 37°C. Motility, plasma membrane functionality and acrosome integrity were assessed. III. Other straws (from different ejaculates) processed as mentioned were further cooled to -3 or -5°C, frozen over nitrogen vapours and stored in liquid nitrogen for one month. Straws were thawed in a water bath at 38°C for 30 s. Motility, plasma membrane functionality, plasma membrane integrity, acrosome integrity, capacitation status and plasma membrane fluidity were assessed. Ice nucleation temperature was -14.3 ± 2.05°C (mean ± SD); cooling to +5, -3, -5 and -7°C, without freezing, produces no differences on sperm quality between target temperatures; cooling to +5, -3, and -5°C produced no differences on sperm survival and plasma membrane fluidity after freeze-thawing. In conclusion, cooling of dog spermatozoa to different subzero temperatures did not improve sperm cryosurvival and had no effect on plasma membrane fluidity after thawing.
authors.affiliation: Departamento de Ciencias Pecuarias, Facultad de Estudios Superiores - Cuautitlan, Universidad Nacional Autonoma de Mexico, Cuautitlan Izcalli, Mexico.
authors.forename: A
authors.initials: A
authors.lastname: Alcantar-Rodriguez
authors.affiliation: Departamento de Ciencias Pecuarias, Facultad de Estudios Superiores - Cuautitlan, Universidad Nacional Autonoma de Mexico, Cuautitlan Izcalli, Mexico.
authors.forename: A
authors.initials: A
authors.lastname: Medrano
mesh_terms: D000177:Acrosome; D000818:Animals; D002462:Cell Membrane / Q000502:physiology; D015925:Cryopreservation / Q000379:methods / Q000662:veterinary*; D004285:Dogs / Q000502:physiology*; D008297:Male; D012662:Semen Preservation / Q000379:methods / Q000662:veterinary*; D013081:Sperm Motility; D013094:Spermatozoa / Q000502:physiology
keywords: cooling; cryopreservation; male dog; semen; sperm membrane fluidity; subzero temperature