Phosphorylation of protein tyrosine residues in fresh and cryopreserved stallion spermatozoa under capacitating conditions.
Phosphorylation of tyrosine residues on sperm proteins is one important intracellular mechanism regulating sperm function that may be a meaningful indicator of capacitation. There is substantial evidence that cryopreservation promotes the capacitation of sperm and this cryocapacitation is frequently cited as one factor associated with the reduced longevity of cryopreserved sperm in the female reproductive tract. This study was designed to determine whether stallion sperm express different levels of tyrosine phosphorylation after in vitro capacitation and whether thawed sperm display similar phosphorylation characteristics in comparison with freshly ejaculated sperm. Experiments were performed to facilitate comparisons of tyrosine phosphorylation, motility, and viability of sperm prior to and following in vitro capacitation in fresh and frozen-thawed sperm. We hypothesized that equine spermatozoa undergo tyrosine phosphorylation during capacitation and that this phosphorylation is modified when sperm have been cryopreserved. We also hypothesized that tyrosine phosphorylation could be enhanced by the use of the activators dibutyryl cAMP (db cAMP) and caffeine, as well as methyl beta-cyclodextrin-which causes cholesterol efflux from the spermatozoa-and inhibited by the protein kinase A (PK-A) inhibitor H-89. Our results indicate that equine sperm capacitation is mediated by a signaling pathway that involves cAMP-dependent PK-A and tyrosine kinases and that cryopreserved sperm may be more sensitive to inducers of capacitation, which could explain their limited life span when compared with fresh sperm.
Pommer
Angela C
AC
School of Veterinary Medicine, Department of Anatomy, Physiology, and Cell Biology, University of California, Davis, California 95616, USA.
Rutllant
Josep
J
Meyers
Stuart A
SA
D000818:Animals; D003994:Bucladesine / Q000494:pharmacology; D002110:Caffeine / Q000494:pharmacology; D015925:Cryopreservation; D003505:Cyclodextrins / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005455:Fluorescent Antibody Technique; D006736:Horses / Q000502:physiology*; D007546:Isoquinolines / Q000494:pharmacology; D008297:Male; D010766:Phosphorylation / Q000187:drug effects; D013075:Sperm Capacitation / Q000502:physiology*; D013094:Spermatozoa / Q000378:metabolism*; D013449:Sulfonamides; D014443:Tyrosine / Q000378:metabolism*; D047392:beta-Cyclodextrins
D003505:Cyclodextrins; D004791:Enzyme Inhibitors; D007546:Isoquinolines; D013449:Sulfonamides; D047392:beta-Cyclodextrins; C108732:methyl-beta-cyclodextrin; D002110:Caffeine; D014443:Tyrosine; D003994:Bucladesine; C063509:N-(2-(4-bromocinnamylamino)ethyl)-5-isoquinolinesulfonamide