title: Studies on the intracellular Ca2+ concentration of thawed dog spermatozoa: influence of Equex from different sources, two thawing diluents and post-thaw incubation in capacitating conditions.
abstract: The addition of 0.5% (v/v) of Equex STM Paste (Nova Chemical Sales, Scituate Inc., MA, USA), whose active ingredient is sodium dodecyl sulphate (SDS), to a Tris-egg yolk extender was demonstrated to improve the longevity of frozen-thawed dog spermatozoa during in vitro incubation at 38 degrees C. The aim of the first experiment was to compare the effects of two SDS-containing compounds, Equex STM Paste and Equex Pasta (Minitüb, Tiefenbach, Germany), when added to a Tris-egg yolk based extender, on the post-thaw longevity of dog spermatozoa, as well as on the intracellular Ca2+ concentration of spermatozoa, during post-thaw incubation at 38 degrees C. The post-thaw sperm survival and longevity, as well as the quality of the sperm movement, were significantly better when using Equex STM Paste. Such prolonged sperm longevity, however, was associated to a higher intracellular Ca2+ concentration in a large subpopulation of the live spermatozoa. A second experiment was aimed to evaluate the effects of sperm dilution immediately post-thaw with a Tris buffer containing glucose or fructose. The two Tris buffers were no different for any of the sperm parameters studied. The aim of a third experiment was to evaluate the sperm longevity, motility patterns and intracellular Ca2+ concentration of cryopreserved dog spermatozoa during post-thaw incubation in capacitating conditions [canine capacitating medium (CCM) with or without 5 microg/ml of heparin]. Heparin had no significant effects on any of the sperm parameters evaluated. During the first 8 h of incubation, the majority of the live spermatozoa had a high intracellular Ca2+ content. However, after 8-10 h of incubation, it had significantly declined. The highest proportion of fast motile sperm, and the highest curvilinear velocity, average path velocity and amplitude of lateral head displacement for the total motile sperm were observed during the 2-4-h incubation period. It was concluded that: (a) the addition of 0.5% (v/v) of Equex STM Paste to a Tris-egg yolk based extender significantly improved the post-thaw longevity of dog spermatozoa, but the same concentration of Equex Pasta had no significant beneficial effects; (b) sperm dilution after thawing with a Tris buffer containing glucose or fructose made no difference in post-thaw sperm longevity; (c) the addition of 5 microg/ml of heparin to CCM had no significant capacitating effects on frozen-thawed dog spermatozoa.
authors.affiliation: Unit of Reproduction and Obstetrics, Department of Animal Pathology, Faculty of Veterinary Medicine, University of Santiago de Compostela, Lugo, Spain. anaipena@correo.lugo.usc.es
authors.forename: A I
authors.initials: AI
authors.lastname: Peña
authors.forename: L
authors.initials: L
authors.lastname: López-Lugilde
authors.forename: M
authors.initials: M
authors.lastname: Barrio
authors.forename: J J
authors.initials: JJ
authors.lastname: Becerra
authors.forename: L A
authors.initials: LA
authors.lastname: Quintela
authors.forename: P G
authors.initials: PG
authors.lastname: Herradón
mesh_terms: D000818:Animals; D002118:Calcium / Q000032:analysis*; D002470:Cell Survival / Q000187:drug effects; D015925:Cryopreservation / Q000662:veterinary; D004285:Dogs / Q000502:physiology*; D005434:Flow Cytometry / Q000662:veterinary; D006493:Heparin / Q000494:pharmacology; D008297:Male; D012662:Semen Preservation / Q000379:methods / Q000662:veterinary*; D012967:Sodium Dodecyl Sulfate / Q000494:pharmacology*; D013075:Sperm Capacitation / Q000187:drug effects* / Q000502:physiology; D013076:Sperm Count / Q000662:veterinary; D013081:Sperm Motility / Q000187:drug effects / Q000502:physiology; D013094:Spermatozoa / Q000737:chemistry* / Q000187:drug effects / Q000502:physiology; D013501:Surface-Active Agents / Q000494:pharmacology*; D013997:Time Factors
chemical_list: D013501:Surface-Active Agents; D012967:Sodium Dodecyl Sulfate; D006493:Heparin; D002118:Calcium