The expression of atrial natriuretic peptide in the oviduct and its functions in pig spermatozoa

Abstract

Locally synthesized atrial natriuretic peptide (ANP) and its receptors have been found in reproductive tissues of various mammals, and play an important role in the acrosome reaction of human sperm. The objective of the present study was to examine the expression of ANP and its receptors in pig spermatozoa and oviduct, and the effect of ANP on pig spermatozoa function. The expression of ANP and its receptors was analyzed by RT-PCR. Only natriuretic peptide receptors-A (NPRA) mRNA was detected in fresh sperm. While the levels of natriuretic peptide receptors-C (NPRC) mRNA were low with no obvious change among different oviductal phases, the levels of ANP mRNA were high in oviduct(OT)1 , OT3 and OT5, but were very low in OT2. On the other hand, the levels of NPRA mRNA were low in OT1 and OT2, increased in OT3 and reached a maximum in OT4 and OT5. Western blot analysis revealed that the level of ANP was high in OT1, decreased in OT2 and OT3, and arrived at the nadir in OT4 and OT5. The effect of ANP on spermatozoa function was studied by the acrosome reaction and IVF. Incubation with ANP for 1 h significantly induced acrosome reaction of preincubated spermatozoa, and maximal response of acrosome reaction (34·1 2·3%) was achieved at 1 nM ANP treatment. Both C-ANP-(4-23), a selective ligand of NPRC, and caffeine had no effect on the acrosome reaction. The stimulatory effect of ANP on acrosome reaction could be mimicked by the permeable cGMP analog, 8-Br-cGMP. ANP and caffeine had a similar effect on improving the oocytes penetration rate, polyspermy rate and the average number of sperm per penetrated oocyte. Also, ANP treatment had a similar effect on cleavage rate, blastocyst formation rate and the number of cells per blastocyst as that of caffeine treatment. The effects of ANP on the acrosome reaction and the parameters of oocyte penetration could be blocked by cGMP-dependent protein kinase (PKG) inhibitors KT5823 and/or Rp-8-pCPT-cGMPS. These results suggest that the expression of ANP in the oviduct may be involved in the regulation of the acrosome reaction and the fertilising ability of pig spermatozoa, and the PKG pathway possibly participates in the process.

Oviductal secretion and gamete interaction

“…Its receptor was detected in spermatozoa (Zhang et al 2006). The preincubation of boar spermatozoa with atrial natriuretic peptide has been shown to induce the acrosome reaction (Zhang et al 2006).…”

“…It has been shown that pre-incubation of spermatozoa with this peptide increased the oocyte penetration rate and decreased the polyspermy rate and the average number of sperm per penetrated oocyte in pigs (Zhang et al 2006). …”

Oviduct: roles in fertilization and early embryo development

“…The merging of follicular fluid and the oviductal fluid after ovulation may provide sufficient chemoattractants to ensure the arrival of sperm at the ampulla. Atrial natriuretic peptide (ANP) and its precursor A (NPPA) are found in pig, mouse, rat and rabbit oviducts (Kim et al 1997, Zhang et al 2006, Bian et al 2012. ANP receptor (NPR1), however, is expressed on the sperm.…”

“…ANP receptor (NPR1), however, is expressed on the sperm. Upon ANP and NPR1 binding, ANP activates the cyclic GMPdependent protein kinase pathway (PKG) and induces acrosome reactions in pig, cow and human sperm (Zamir et al 1995, Rotem et al 1998, Zhang et al 2006. This reaction might provide additional evidence regarding the role of ANP as a human sperm chemoattractant (Zamir et al 1993, Anderson et al 1995.…”

Deleted in malignant brain tumor 1 is secreted in the oviduct and involved in the mechanism of fertilization in equine and porcine species

“…Moreover, some oviductal proteins have been shown to interact with gametes and to improve efficiency of IVF in porcine (McCauley et al 2003, bovine (Martus et al 1998), human (O' Day-Bowman et al 1996), and equine (Mugnier et al 2009) species. Some of these proteins, such as osteopontin, have been identified (Hao et al 2006, Goncalves et al 2008, Monaco et al 2009) in cattle and pigs and atrial natriuretic peptide A (Anderson et al 1994, Zamir et al 1995, Zhang et al 2006 and oviductin (oviduct-specific glycoprotein; Martus et al 1998, Buhi 2002, McCauley et al 2003 in cattle, pigs, and humans.…”