title: Effects of dead spermatozoa on motion characteristics and membrane integrity of live spermatozoa in fresh and cooled-stored equine semen.
abstract: The aim of this study was to determine if dead spermatozoa reduced motility or membrane integrity of live spermatozoa in fresh and cooled-stored equine semen. Three ejaculates from each of three stallions were centrifuged and virtually all seminal plasma was removed. Spermatozoa were resuspended to 25 x 10(6) spermatozoa/ml with EZ-Mixin CST extender and 10% autologous seminal plasma, then divided into aliquots to which 0 (control), 10, 25, 50, or 75% (v/v) dead spermatozoa were added. Dead spermatozoa preparations contained 25 x 10(6) spermatozoa/ml and 10% seminal plasma from pooled ejaculates of the three stallions, in EZ-Mixin CST extender. Spermatozoa were killed in the pooled ejaculates by repeated freezing and thawing, then stored at -20 degrees C until warmed to 37 degrees C and mixed with aliquots of fresh spermatozoa to be cooled and stored in an Equitainer for 24h. Motion characteristics (% total motility (MOT), % progressive motility (PMOT), and mean curvilinear velocity (VCL)) for fresh and 24h cooled samples were determined using a computerized spermatozoal motion analyzer. The presence of up to 75% dead spermatozoa did not adversely affect MOT or PMOT of live spermatozoa in either fresh or cooled-stored semen. However, VCL and the percentage of membrane-intact spermatozoa were reduced compared to control samples when 75% (v/v) dead spermatozoa were added. Membrane integrity, as assessed by staining with carboxyfluoresein diacetate-propidium iodide, was highly correlated (r>0.8; P<0.001) with MOT and PMOT in both fresh and cooled-stored semen samples. Results of this study have application to the processing of both cooled and frozen equine semen.
authors.affiliation: Department of Large Animal Medicine and Surgery, College of Veterinary Medicine, Texas A&M University, College Station, TX 77843-4475, USA.
authors.forename: S P
authors.initials: SP
authors.lastname: Brinsko
authors.forename: T L
authors.initials: TL
authors.lastname: Blanchard
authors.forename: S L
authors.initials: SL
authors.lastname: Rigby
authors.forename: C C
authors.initials: CC
authors.lastname: Love
authors.forename: D D
authors.initials: DD
authors.lastname: Varner
mesh_terms: D000818:Animals; D002462:Cell Membrane / Q000502:physiology; D015925:Cryopreservation / Q000662:veterinary; D006736:Horses / Q000502:physiology*; D008297:Male; D012661:Semen / Q000166:cytology; D012662:Semen Preservation / Q000662:veterinary*; D013076:Sperm Count / Q000662:veterinary; D013081:Sperm Motility / Q000502:physiology; D013094:Spermatozoa / Q000502:physiology*