title: Effects of milk proteins on sperm binding to the zona pellucida and intracellular Ca(2+) concentration in stallion sperm.
abstract: Objectives were to determine the effects of extracellular Ca(2+) and milk proteins on intracellular Ca(2+) concentrations in stallion sperm; and to determine the effects of single caseins on sperm binding to the zona pellucida (ZP). In Experiment I, sperm were incubated in media containing 2 or 4mM Ca(2+) and intracellular Ca(2+) concentration was determined after ionomycin treatment and long-term incubation (3h). Extracellular Ca(2+) concentrations (2 compared with 4mM) did not affect baseline intracellular Ca(2+) concentration of sperm. However, incubating sperm in a medium containing 4 compared with 2mM Ca(2+) resulted in greater (P<0.05) influx of Ca(2+) into sperm. In Experiment II, sperm incubated in media containing 1mg/mL of native phosphocaseinate (NP) or sodium caseinate (SC) showed similar baseline intracellular Ca(2+) and influx of Ca(2+) than control (TALP). In Experiment III, sperm-ZP binding assays were performed in TALP medium containing: no additions (TALP); 1mg/mL SC; 1 or 3mg/mL of α-casein; 1 or 3mg/mL of β-casein; and 1 or 3mg/mL of κ-casein. The number of stallion sperm bound to bovine ZP was greatest (P<0.05) when SC was used. Co-incubation in media containing single caseins (α-, β- or κ-casein) resulted in similar results to TALP; however, a dose effect (P<0.05) was observed for β- and κ-caseins. In conclusion, extracellular Ca(2+) concentration and milk proteins did not affect baseline intracellular calcium in stallion sperm. It appears that β- and κ-caseins may be responsible for enhancing sperm binding to ZP, but the mechanism remains unknown.
authors.affiliation: Department of Veterinary Clinical Sciences, The Ohio State University, Columbus, OH 43210, USA. Electronic address: marco.dasilva@cvm.osu.edu.
authors.forename: Marco A
authors.initials: MA
authors.lastname: Coutinho da Silva
authors.affiliation: Animal Reproduction and Biotechnology Laboratory, Colorado State University, Fort Collins, CO 80523, USA.
authors.forename: George E
authors.initials: GE
authors.lastname: Seidel
authors.affiliation: Gluck Equine Research Center, University of Kentucky, Lexington, KY 40546, USA.
authors.forename: Edward L
authors.initials: EL
authors.lastname: Squires
authors.affiliation: Animal Reproduction and Biotechnology Laboratory, Colorado State University, Fort Collins, CO 80523, USA.
authors.forename: James K
authors.initials: JK
authors.lastname: Graham
authors.affiliation: Animal Reproduction and Biotechnology Laboratory, Colorado State University, Fort Collins, CO 80523, USA.
authors.forename: Elaine M
authors.initials: EM
authors.lastname: Carnevale
mesh_terms: D000818:Animals; D002118:Calcium / Q000378:metabolism*; D061207:Calcium Ionophores / Q000494:pharmacology; D006736:Horses / Q000502:physiology*; D015759:Ionomycin / Q000494:pharmacology; D008297:Male; D008894:Milk Proteins / Q000494:pharmacology*; D010063:Ovum / Q000502:physiology; D013081:Sperm Motility; D013084:Sperm-Ovum Interactions / Q000187:drug effects*; D013094:Spermatozoa / Q000187:drug effects* / Q000378:metabolism; D013997:Time Factors; D015044:Zona Pellucida / Q000187:drug effects / Q000502:physiology*
chemical_list: D061207:Calcium Ionophores; D008894:Milk Proteins; D015759:Ionomycin; D002118:Calcium
keywords: Calcium; Casein; Equine; Sperm; Stallion; ZP-binding