The Voltage-Gated Sodium Channel Nav1.8 Is Expressed in Human Sperm
et al. (2013) The Voltage-Gated Sodium Channel Nav1.8 Is
Expressed in Human Sperm. PLoS ONE 8(9): e76084. doi:10.1371/journal.pone.0076084
The Voltage-Gated Sodium Channel Na 1.8 Is Expressed v in Human Sperm
Antonio Cejudo-Roman 0
Francisco M. Pinto 0
Nerea Subirn 0
Cristina G. Ravina 0
Manuel Fernndez- 0
Snchez 0
Natalia Prez-Hernndez 0
Ricardo Prez 0
Alberto Pacheco 0
Jon Irazusta 0
Luz Candenas 0
Agustin Guerrero-Hernandez, Cinvestav-IPN, Mexico
0 1 Instituto de Investigaciones Quimicas (IIQ), CSIC - Universidad de Sevilla , Sevilla , Spain , 2 Departamento de Fisiologia, Facultad de Medicina y Odontologia, Universidad del Pais Vasco , Leioa, Bizkaia , Spain , 3 Instituto Valenciano de Infertilidad , Sevilla , Spain , 4 Instituto Madrileno de Infertilidad , Madrid , Spain
The role of Na+ fluxes through voltage-gated sodium channels in the regulation of sperm cell function remains poorly understood. Previously, we reported that several genes encoding voltage-gated Na+ channels were expressed in human testis and mature spermatozoa. In this study, we analyzed the presence and function of the TTX-resistant VGSC subunit Nav1.8 in human capacitated sperm cells. Using an RT-PCR assay, we found that the mRNA of the gene SCN10A, that encode Na v1.8, was abundantly and specifically expressed in human testis and ejaculated spermatozoa. The Na v1.8 protein was detected in capacitated sperm cells using three different specific antibodies against this channel. Positive immunoreactivity was mainly located in the neck and the principal piece of the flagellum. The presence of Na v1.8 in sperm cells was confirmed by Western blot. Functional studies demonstrated that the increases in progressive motility produced by veratridine, a voltage-gated sodium channel activator, were reduced in sperm cells preincubated with TTX (10 M), the Na v1.8 antagonist A-803467, or a specific Na v1.8 antibody. Veratridine elicited similar percentage increases in progressive motility in sperm cells maintained in Ca2+containing or Ca2+-free solution and did not induce hyperactivation or the acrosome reaction. Veratridine caused a rise in sperm intracellular Na+, [Na+]i, and the sustained phase of the response was inhibited in the presence of A-803467. These results verify that the Na+ channel Na v1.8 is present in human sperm cells and demonstrate that this channel participates in the regulation of sperm function.
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Funding: This work was financially supported by grants from Junta de Andaluca (P08-CVI-04185) and Ministerio de Economa y Competitividad
(CTQ2011-25564), Spain, with joint financing by FEDER and FSE funds from the European Union. The funders had no role in study design, data collection
and analysis, decision to publish, or preparation of the manuscript.
Competing interests: The authors have declared that no competing interests exist.
Ion channels play a central role in the regulation of sperm
intra- and inter-cellular signaling [1-9]. The rapid ion fluxes
through these membrane proteins permit a quick transfer of
information between sperm and its surrounding [1,10-12]. This
communication is essential for correct sperm guidance
throughout the female reproductive tract as well as for
acquisition of fertilization competence and interaction with the
oocyte [10-13]. Many different ion channels have been
identified in the sperm cell membrane. Among them, Ca2+, K+,
H+ and anion channels are widely distributed in the head and
flagellum and play an important role in regulating sperm
function including motility, capacitation and acrosome reaction
[1,2,5,6,11,12]. Recently, different works have pointed to the
importance of the sperm-specific Ca2+ channels CatSper in the
control of intracellular Ca2+ concentration, [Ca2+]i, in sperm cells
[3,7,8,14,15]. These channels mediate the
progesteroneinduced Ca2+ influx in human sperm and are essential for
sperm function and male fertility [7,8,15]. Na+ channels should
also play an important role in sperm, as the gradient of this ion
across the plasma membrane plays a central role in the
regulation of membrane potential (Em), a parameter that govern
the rate and direction of ion-flow through channels and
exchangers and modulates the intracellular pH (pHi) [16,17]. In
this context, the presence of epithelial Na+ channels of the
ENaC family [17] and of voltage-gated Na+ channels (VGSC)
[18] has been demonstrated in sperm cells, although their role
remain poorly understood.
VGSCs are complex proteins composed by a and one or
more auxiliary subunits [19]. Nine different VGSCs subunits
have been cloned in mammals, each one encoded by a
different gene [19,20]. They can be further characterized by
their sensitivity to the highly selective blocker tetrodotoxin
(TTX). The TTX-sensitive subunits are inhibited in the
nanomolar range by TTX and include Na v1.1, Na v1.2, Na v1.3,
Na v1.4, Na v1.6 and Na v1.7. The TTX-resistant subunits are
inhibited in the micromolar range by TTX and include Na v1.5,
(...truncated)