To fertilize the egg, sperm cells must reside in the female reproductive tract for several hours during which they undergo chemical and motility changes collectively called capacitation. During capacitation, the sperm develop a unique type of motility known as hyperactivated motility (HAM). The semen contains Zn in millimolar concentrations, whereas in the female reproductive tract, the concentration is around 1 µM. In this study, we characterize the role of Zn in human sperm capacitation focusing on its effect on HAM. Western blot analysis revealed the presence of GPR39-type Zn-receptor localized mainly in the sperm tail. Zn at micromolar concentration stimulates HAM, which is mediated by a cascade involving GPR39-adenylyl cyclase (AC)-cyclic AMP (cAMP)-protein kinase A-tyrosine kinase Src (Src)-epidermal growth factor receptor and phospholipase C. Both the transmembrane AC and the soluble-AC are involved in the stimulation of HAM by Zn . The development of HAM is precisely regulated by cAMP, in which relatively low concentration (5-10 µM) stimulated HAM, whereas at 30 µM no stimulation occurred. A similar response was seen when different concentrations of Zn were added to the cells; low Zn stimulated HAM, whereas at relatively high Zn , no effect was seen. We further demonstrate that the Ca -channel CatSper involved in Zn -stimulated HAM. These data support a role for extracellular Zn acting via GPR39 to regulate signaling pathways in sperm capacitation, leading to HAM induction.

Several studies proposed the importance of zinc ion in male fertility. Here, we describe the properties, roles and cellular mechanisms of action of Zn2+ in spermatozoa, focusing on its involvement in sperm motility, capacitation and acrosomal exocytosis, three functions that are crucial for successful fertilization. The impact of zinc supplementation on assisted fertilization techniques is also described. The impact of zinc on sperm motility has been investigated in many vertebrate and invertebrate species. It has been reported that Zn2+ in human seminal plasma decreases sperm motility and that Zn2+ removal enhances motility. Reduction in the intracellular concentration of Zn2+ during epididymal transit allows the development of progressive motility and the subsequent hyper activated motility during sperm capacitation. Extracellular Zn2+ affects intracellular signaling pathways through its interaction with the Zn2+ sensing receptor (ZnR), also named GPR39. This receptor was found in the sperm tail and the acrosome, suggesting the possible involvement of Zn2+ in sperm motility and acrosomal exocytosis. Our studies showed that Zn2+ stimulates bovine sperm acrosomal exocytosis, as well as human sperm hyper-activated motility, were both mediated by GPR39. Zn2+ binds and activates GPR39, which activates the trans-membrane-adenylyl-cyclase (tmAC) to catalyze cAMP production. The NHE (Na+/H+-exchanger) is activated by cAMP, leading in increased pHi and activation of the sperm-specific Ca2+ channel CatSper, resulting in an increase in [Ca2+]i, which, together with HCO3−, activates the soluble adenylyl-cyclase (sAC). The increase in [cAMP]i activates protein kinase A (PKA), followed by activation of the Src-epidermal growth factor receptor-Pphospholipase C (Src-EGFR-PLC) cascade, resulting in inositol-triphosphate (IP3) production, which mobilizes Ca2+ from the acrosome, causing a further increase in [Ca2+]i and the development of hyper-activated motility. PKA also activates phospholipase D1 (PLD1), leading to F-actin formation during capacitation. Prior to the acrosomal exocytosis, PLC induces phosphadidylinositol-4,5-bisphosphate (PIP2) hydrolysis, leading to the release of the actin-severing protein gelsolin to the cytosol, which is activated by Ca2+, resulting in F-actin breakdown and the occurrence of acrosomal exocytosis.

To fertilize the egg, sperm cells must reside in the female reproductive tract for several hours during which they undergo chemical and motility changes collectively called capacitation. During capacitation, the sperm develop a unique type of motility known as hyperactivated motility (HAM). The semen contains Zn 2+ in millimolar concentrations, whereas in the female reproductive tract the concentration is around 1 µM. In this study, we characterize the role of Zn 2+ in human sperm capacitation focusing on its effect on HAM. Western blot analysis revealed the presence of G protein-coupled receptor 39 (GPR39) type Zn-receptor localized mainly in the sperm tail. Zn 2+ at micromolar concentration stimulates HAM, which is mediated by a cascade involving GPR39-AC-cAMP-PKA-Src-EGFR and phospholipase C. Both the transmembrane adenylyl cyclase (AC) and the soluble-AC are involved in the stimulation of HAM by Zn 2+ . The development of HAM is precisely regulated by cyclic adenosine monophosphate, in which relatively low concentration (5-10 µM) stimulated HAM, whereas at 30 µM no stimulation occurred. A similar responsewas seen when different concentrations of Zn 2+ were added to the cells; low Zn 2+ stimulated HAM, whereas at relatively high Zn 2+ , no effect was seen. We further demonstrate that the Ca 2+ -channel CatSper involved in Zn 2+ -stimulated HAM. These data support a role for extracellular Zn 2+ acting via GPR39 to regulate signaling pathways in sperm capacitation, leading to HAM induction.

Introduction: The aim of the study was to study the effect of Zn 2+ addition to human sperm prior to and after cryopreservation on post-thaw sperm motility. Material and methods: Human semen was liquefied, and the semen was loaded on a gradient and centrifuged for 10 min at 3000 g at room temperature. The lower layer containing the sperm was collected, resuspended in Ham's F-10 medium then centrifuged again, and the sperm were allowed to "swim up" after the last wash at 37°C. The motile cells were collected, resuspended in capacitation medium and supplemented with 0.1% human serum albumin. Samples with or without ZnCl 2 were kept at room temperature for 10 min, after which they were mixed with equal volumes of cryoprotective medium and aliquots were transferred to screw-top cryovials and stored in liquid nitrogen for several days. The samples were removed from liquid nitrogen, subjected to rapid thawing and washed in HamF-10 medium. Sperm motility was analyzed using computer-assisted sperm analysis (CASA). Results: Freezing of human sperm in the presence of Zn 2+ led to a significant increase in the number of motile sperm as well as preserving motility for a longer period of time. The percentage of sperm presenting progressive motility was also increased by Zn 2+ depending on the fluid volume used for freezing. The presence of Zn 2+ in the freezing medium also protects sperm motility after second freezing. Conclusions: In order to achieve good motility after freezing, human sperm should be frozen in 0.1 ml of medium containing 50 µM Zn 2+ .