Effect of cariporide on ram sperm pH regulation and motility: possible role of NHE1

Abstract

Sperm motility, a feature essential for fertilization, is influenced by intracellular pH (pH) homeostasis. Several mechanisms are involved in pH regulation, among which sodium-hydrogen exchangers (NHEs), a family of integral transmembrane proteins that catalyze the exchange of Na for H across lipid bilayers. A preliminary characterization of NHE activity and kinetic parameters, followed by analysis of the expression and localization of the protein in ram spermatozoa was performed. NHE activity showed an apparent for external Na of 17.61 mM. Immunoblotting revealed a molecular mass of 85 kDa. Immunolocalization pattern showed some species-specific aspects, such as positive labeling at the equatorial region of the sperm head. Cariporide, a selective NHE1 inhibitor, significantly reduced pH recovery (85%). Similarly, exposure to cariporide significantly inhibited different motility parameters, including those related to sperm capacitation. fertilization (IVF) was not affected by cariporide, possibly due to the non-dramatic, although significant, drop in motility and velocity parameters or due to prolonged exposure during IVF, which may have caused progressive loss of its inhibitory effect. In conclusion, this is the first study documenting, in a large animal model (sheep) of well-known translational relevance, a direct functional role of NHE on sperm pH and motility. The postulated specificity of cariporide toward isoform 1 of the Na/H exchanger seems to suggest that NHE1 may contribute to the observed effects on sperm cell functionality.

Na+/H+ Exchangers Involve in Regulating the pH-Sensitive Ion Channels in Mouse Sperm

“…As the main transporters for eliminating the cytosolic protons, six isoforms of NHEs: NHE1 [31][32][33], NHE5 [34], NHE8 [35], sNHE [36,37], NHA1 [38], and NHA2 [39] had been reported to be expressed in sperm. However, which NHE is the determinant modulator for the activation of KSper and CatSper in murine sperm remained ambiguous.…”

The Role of Cariporide in Protecting Saphenous Vein among Different Aldehyde Dehydrogenase Genotypes

“…Previous studies showed that caspase 3 plays an important role in apoptosis in the endoplasmic reticulum [8]. In this study, the GSVs discarded during CABG were collected, and the vein was preserved according to the condition of each group.…”

Blocking NHE Channels Reduces the Ability of In Vitro Capacitated Mammalian Sperm to Respond to Progesterone Stimulus

“…Interestingly, previous research using patch-clamping in mouse sperm demonstrated that NHE channels are functionally coupled to the CatSper ones [ 4 , 11 ]. Furthermore, recent studies reported that sNHE in humans [ 17 ] and NHE1 in sheep [ 18 ] are implicated in the regulation of sperm motility. Since not only does the process through which a spermatozoon acquires their capacity to fertilize an oocyte entail changes in sperm kinematics but also in plasma membrane permeability, intracellular calcium levels, mitochondrial activity, and acrosome integrity, the objective of the present study was to determine the physiological relevance of NHE channels in the sperm ability to elicit in vitro capacitation and progesterone-induced acrosome exocytosis.…”

“…Since not only does the process through which a spermatozoon acquires their capacity to fertilize an oocyte entail changes in sperm kinematics but also in plasma membrane permeability, intracellular calcium levels, mitochondrial activity, and acrosome integrity, the objective of the present study was to determine the physiological relevance of NHE channels in the sperm ability to elicit in vitro capacitation and progesterone-induced acrosome exocytosis. These assays were based on the pharmacological blockage of NHEs channels using a specific inhibitor (5-(N,N-dimethyl)-amiloride; DMA) at different concentrations [ 4 , 18 ]. DMA induces the total blockage of NHE1 and NHE2 isoforms and the partial blockage of NHE5; in contrast, NHE3 and NHE4 are amiloride-resistant isoforms (reviewed in [ 14 ]).…”

“…To the best of our knowledge, no data about the inhibitory effect of this drug on sNHE, NHA1 or NHA2 exist. Considering that only NHE1, NHE5, and sNHE isoforms have been identified in the plasma membrane of mammalian sperm [ 11 , 17 , 18 ], our pharmacological assay was predicted to lead to the total blockage of NHE1 channels, and either a lack or partial blockage of the other NHE isoforms. At each relevant time point, sperm motility parameters were evaluated using a computer-assisted sperm analysis (CASA) system, whereas plasma membrane and acrosome integrity, mitochondrial activity, and intracellular levels of calcium, superoxide, and overall ROS were measured by flow cytometry.…”