Abstract
This study sought to identify and localize SLO1 channels in boar spermatozoa by immunoblotting and immunofluorescence, and to determine their physiological role during in vitro sperm capacitation. Sperm samples from 14 boars were incubated in a capacitation medium for 300 min in the presence of paxilline (PAX), a specific SLO1-channel blocker, added either at 0 min or after 240 min of incubation. Negative controls were incubated in capacitation medium, and positive controls in capacitation medium plus tetraethyl ammonium (TEA), a general K+-channel blocker, also added at 0 min or after 240 min of incubation. In all samples, acrosome exocytosis was triggered with progesterone after 240 min of incubation. Sperm motility and kinematics, integrity of plasma and acrosome membranes, membrane lipid disorder, intracellular calcium levels and acrosin activity were evaluated after 0, 60, 120, 180, 240, 250, 270 and 300 min of incubation. In boar spermatozoa, SLO1 channels were found to have 80 kDa and be localized in the anterior postacrosomal region and the mid and principal piece of the tail; their specific blockage through PAX resulted in altered calcium levels and acrosome exocytosis. As expected, TEA blocker impaired in vitro sperm capacitation, by altering sperm motility and kinematics and calcium levels. In conclusion, SLO1 channels are crucial for the acrosome exocytosis induced by progesterone in in vitro capacitated boar spermatozoa.
Highlights
Capacitation includes a set of physiological changes that are required for a spermatozoon to be able to fertilize the oocyte
To understand the physiological role of SLO1 channels during in vitro capacitation, boar spermatozoa were incubated in a capacitation medium for 300 min in the presence of PAX or tetraethyl ammonium (TEA) blockers
The lack of differences between TEA and PAX blockers strongly suggests, for the first time, that SLO1 channels have an essential role in triggering calcium conductance in the sperm head during in vitro capacitation of boar spermatozoa and are involved in the acrosome exocytosis induced by progesterone
Summary
Capacitation includes a set of physiological changes that are required for a spermatozoon to be able to fertilize the oocyte These changes include augmented beat frequency and curvilinear motility, increase in intracellular pH, rise in intracellular calcium levels, plasma membrane hyperpolarization and tyrosine phosphorylation of certain sperm proteins (reviewed in [1,2,3]). Previous studies have reported that SLO channels are the main K+ channels in human and mouse spermatozoa These channels, which belong to the SLO gene family [2,3,9,11,12], have been reported to play a vital role in the regulation of sperm volume, and have been found to be involved in the sequence of changes that take place during capacitation [2,9,13]. SLO1 channels or big potassium (BK) or maxi K+ channels, exhibit high sensitivity to changes of both voltage and intracellular Ca2+ levels [15,16], whereas SLO3 channels are highly sensitive to intracellular alkalinization [9]
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