Inducible nitric oxide synthase-derived nitric oxide regulates germinal vesicle breakdown and first polar body emission in the mouse oocyte

Abstract

The present study investigated the subcellular localization of inducible nitric oxide synthase (iNOS) during mouse oocyte meiotic maturation and fertilization using confocal microscopy, and further studied the roles of iNOS-derived NO in oocyte maturation by using an iNOS-specific inhibitor aminoguanidine (AG) and iNOS antibody microinjection. In germinal vesiclestage oocytes, iNOS immunoreactivity was mainly localized in the germinal vesicle. Shortly after germinal vesicle breakdown, the iNOS immunoreactivity accumulated around the condensed chromosomes. At metaphase I and metaphase II, with the organization of chromosomes to the equatorial plate, iNOS immunoreactivity was concentrated around the aligned chromosomes, putatively the position of the metaphase spindle. The accumulation of iNOS immunoreactivity could not be detected at anaphase I and anaphase II. However, at telophase I and telophase II, the staining of iNOS was concentrated in the region between the separating chromosomes/chromatids. Furthermore, the staining of iNOS also accumulated in the male and female pronuclei in fertilized eggs. Germinal vesicle breakdown and the first polar body emission of the oocytes were significantly blocked by the iNOS-specific inhibitor AG in a dose-dependent manner. The germinal vesicle breakdown in oocytes injected with iNOS antibody was also inhibited. We found that the phosphorylation of mitogen-activated protein kinase in oocytes after germinal vesicle breakdown was inhibited by AG treatment. The control oocytes extruded a normal first polar body, while the AG-treated oocytes exhibited an elongated protrusion or no elongated protrusion. The results of confocal microscopy showed that the AG-treated oocytes were arrested at anaphase I -telophase I. Our results suggest that the iNOSderived NO pathway plays important roles in mouse oocyte meiotic maturation, especially in germinal vesicle breakdown and the anaphase -telophase transition.

Expression and localization of nitric oxide synthase isoforms during porcine oocyte growth and acquisition of meiotic competence

“…This peripheral localization is probably important for the propagation of NO-signalling in somatic cells (Webb et al, 2001), and we cannot exclude that it is also important for oocytes. Our observations are consistent with those made in oocytes from laboratory rodents (Van Voorhis et al, 1994Jablonka-Shariff and Olson, 1997;Nakamura et al, 2002;Huo et al, 2005). There are very limited and inconsistent data concerning the expression and localization of iNOS in porcine oocytes.…”

Reduction of nitric oxide level leads to spontaneous resumption of meiosis in diplotene-arrested rat oocytes culturedin vitro

“…Few studies suggest that high level of NO induces meiotic resumption from diplotene arrest in mouse, 7,15 rat, 16 porcine, 9,17,18 and murine oocytes. 13 On the other hand, NO inhibits meiotic resumption from diplotene arrest in rat .................................................................................................................. 41 and a reduced level of NO triggers meiotic resumption from diplotene arrest in rat oocytes cultured in vitro.…”

Developmental competence and expression pattern of bubaline (Bubalus bubalis) oocytes subjected to elevated temperatures during meiotic maturation in vitro

“…However, no changes (p>0.05) were observed in mRNA expression of Dna J in all treatments groups compared to control. MnSOD protects cells against oxidative damage [45] and iNOS has a role in germinal vesicle breakdown and the anaphase-telophase transition of oocytes [46], therefore, its upregulation may negatively impact buffalo oocytes during maturation and subsequent embryo development. Our results are in accordance with the study by Balasubramanium et al [47] where the expression of MnSOD decreased when embryos were cultured under high oxygen concentration (20 %) compared to the 5 % oxygen concentration normally occurring in vivo.…”

“…Our results are in accordance with the study by Balasubramanium et al [47] where the expression of MnSOD decreased when embryos were cultured under high oxygen concentration (20 %) compared to the 5 % oxygen concentration normally occurring in vivo. Since MnSOD is a mitochondrial enzyme [45][46][47], the decrease in its expression may be related to the altered mitochondrial activity after subjecting the oocytes to heat stress thereby, negatively impacting the developmental competence. However, the exact mechanism of down regulation remains unclear and warrants further validation.…”