HVCN1 but Not Potassium Channels Are Related to Mammalian Sperm Cryotolerance

Abstract

Little data exist about the physiological role of ion channels during the freeze–thaw process in mammalian sperm. Herein, we determined the relevance of potassium channels, including SLO1, and of voltage-gated proton channels (HVCN1) during mammalian sperm cryopreservation, using the pig as a model and through the addition of specific blockers (TEA: tetraethyl ammonium chloride, PAX: paxilline or 2-GBI: 2-guanidino benzimidazole) to the cryoprotective media at either 15 °C or 5 °C. Sperm quality of the control and blocked samples was performed at 30- and 240-min post-thaw, by assessing sperm motility and kinematics, plasma and acrosome membrane integrity, membrane lipid disorder, intracellular calcium levels, mitochondrial membrane potential, and intracellular O2−⁻ and H2O2 levels. General blockade of K+ channels by TEA and specific blockade of SLO1 channels by PAX did not result in alterations in sperm quality after thawing as compared to control samples. In contrast, HVCN1-blocking with 2-GBI led to a significant decrease in post-thaw sperm quality as compared to the control, despite intracellular O2−⁻ and H2O2 levels in 2-GBI blocked samples being lower than in the control and in TEA- and PAX-blocked samples. We can thus conclude that HVCN1 channels are related to mammalian sperm cryotolerance and have an essential role during cryopreservation. In contrast, potassium channels do not seem to play such an instrumental role.

Ion Channels of Spermatozoa: Structure, Function, and Regulation Mechanisms

“…The plasma membrane of mature sperm contains a high diversity of ion transporters belonging to different protein families and showing different ion affinity, regulation mechanisms, and functional multiplicity [ 6 ]. The absence or dysfunction of even a single channel type may result in male subfertility [ 5 , 7 , 8 ] or affect sperm cryotolerance [ 9 ]; moreover, some ion channels can be used as contraception targets [ 8 , 10 ].…”

“…Most of the original research included in this special issue focuses on the physiological role of K + and H + transporters in mammal sperm physiology. In these species, the sperm plasmalemma contains different K + transporters, such as voltage-gated potassium channels (K v ), inward-rectifier potassium channels (K ir ), calcium-activated potassium channels (K Ca ), and tandem-pore-domain potassium channels (KCNK) [ 3 , 5 , 9 ]. Poli et al [ 5 ] performed an excellent study on the expression and function of Kir4.1 and Kir5.1 channels in mature sperm cells from mice.…”

“…The research by Rodríguez-Páez et al [ 4 ] evidenced that seminal polyamines regulate the activity of K + and Na + channels during in vitro capacitation of mice sperm, thus contributing to the sperm plasma membrane hyperpolarization; in contrast, these compounds did not seem to be implicated in Ca 2+ and Cl − import during in vitro capacitation [ 4 ]. Despite K + currents being relevant for sperm capacitation of mammalian sperm, the relevant research conducted by Delgado-Bermúdez et al [ 9 ] showed that the addition of K + channel blockers to the cryopreservation media did not affect the cryotolerance of pig sperm.…”

“…Interestingly, Rodríguez-Páez et al [ 4 ] proposed that upon ejaculation polyamines may interact with the soluble adenylate cyclase (sAC) and thus favour the activation of NHE and further pH i alkalinization during mouse-sperm capacitation. In pigs, the physiological relevance of HVCN1 channels during in vitro sperm capacitation and cryopreservation has been extensively described by Yeste et al [ 2 ] and Delgado-Bermúdez et al [ 9 ]. Both studies showed that the addition of a HVCN1 pharmacological blocker to either the in vitro capacitation medium [ 2 ] or the cryopreservation media [ 9 ] results in reduced sperm viability and motility, impaired mitochondrial potential, and premature acrosomal exocytosis as compared to control samples.…”

“…In pigs, the physiological relevance of HVCN1 channels during in vitro sperm capacitation and cryopreservation has been extensively described by Yeste et al [ 2 ] and Delgado-Bermúdez et al [ 9 ]. Both studies showed that the addition of a HVCN1 pharmacological blocker to either the in vitro capacitation medium [ 2 ] or the cryopreservation media [ 9 ] results in reduced sperm viability and motility, impaired mitochondrial potential, and premature acrosomal exocytosis as compared to control samples. It is worth noting that the results by Delgado-Bermúdez et al [ 9 ] and Rodríguez-Páez et al [ 4 ] must be taken into consideration in the formulation of specific media to dilute and/or preserve sperm cells.…”