Acquisition of meiotic competence in the rat: Role of gonadotropin and estrogen

Oocytes were removed from the follicles of rats at 15 to 31 days of age, and their ability to resume meiosis (“meiotic competence”) in vitro was correlated with their diameter and the stage of follicular development. The majority of oocytes explanted on day 15 did not resume meiosis when placed in culture, but the percentage of competent oocytes increased from 14.1% ± 3.0% on day 20 to 67.6% ± 3.3% on day 26 of age. This ability to resume maturation correlated well (r = 0.98) with the increase in diameter of oocytes and coincided with the development of antral follicles. Hypophysectomy on day 15 of age, but not on day 20, reduced the percentage (P < 0.001) and number (P < 0.001) of competent oocytes and was accompanied by a reduction in diameter of oocytes. Treatment with PMSG or E₂ increased the number (P < 0.001) and percentage (P < 0.001) of competent oocytes. These results suggest that the ability of oocytes to mature in vitro is dependent upon stimulation by gonadotropins and that this action of gonadotropin may be mediated by production of estrogen within the follicles.     

Superovulatory responses of cattle

Non-histological examination of superovulated ovaries of cows does not allow one to distinguish between corpora lutea and luteinized follicles. A better estimation of ovulation rate could, therefore, be made from the number of embryos recovered or from the levels of E₂-17β in the plasma 60 hours after PMSG.For comparison of different treatments, it is necessary to characterize activities of the stimulatory agents used. Administration of an FSH - LH preparation twice a day at decreasing doses gives the best mean responses, but no treatment has been found which can clearly decrease the large variation between individuals in their responses.Numerical, kinetic and endocrine ovarian factors can partly explain the variability of ovarian responses to PMSG in the heifer. Individual differences in follicular populations at the time of treatment, or in E₂-17β levels after stimulation, could be related to differences in responses in ovulation rate. Normal follicles >1.7 mm diameter before treatment would usually ovulate following PMSG injection, whereas early atretic follicles of the same size mostly luteinize.     

Excessive ovarian follicular development in pregnant pigtailed macaques (Macaca nemestrina)

To evaluate the status and possible control of ovarian follicular development during pregnancy, circulating levels of estrone (E₁), estradiol-17β (E₂), and follicle-stimulating hormone (FSH) were measured throughout gestation in both intact and ovariectomized pregnant pigtailed monkeys (Macaca nemestrina). From an additional group of pregnant monkeys, ovaries were obtained at late gestation (on day 150 or 159 of pregnancy) for histological studies. Circulating concentrations of E₁ and E₂ increased on day 13 and remained elevated for about 10 days; they then declined and reached low levels on day 32 of gestation. After day 60, there were gradual but smaller increases in estrogen levels to day 140, after which both E₁ and E₂ levels increased significantly, reaching maximum levels (E₁ = 832.2 ± 210.8 pg/ ml; E₂ = 1.66 ± 0.32 ng/ml) at the end of pregnancy. Removal of ovaries on day 35 of gestation did not affect pregnancy or the pattern of estrogen secretion. Serum concentrations of FSH demonstrated only minor fluctuations during pregnancy but were similar to those found during the early follicular phase of cycling pigtailed monkeys investigated in this study. Ovarian histology revealed extensive follicular growth; in addition to the corpus luteum of pregnancy, ovaries were packed with pre-antral, small antral, and medium-sized Graafian follicles. Some of these follicles appeared to be cystic and showed various degree of atresia; their general appearance was similar to the follicles of human females with polycystic ovary syndrome. Our data suggest that FSH may initiate ovarian follicular growth during gestation. High levels of estrogens were incapable of suppressing FSH secretion but may be responsible for the induction of atresia in a large number of follicles in pregnant pigtailed monkeys.     

The development of the mouse ovary and its response to exogenous gonadotrophins.

Immature mice aged 14 to 49 days were treated with a single injection of 4 i.u. HCG, or 3 i.u. PMSG followed 48 hr later by 2 i.u. HCG. After treatment with HCG alone the number of oocytes which were ovulated rose gradually from Day 21 to Day 28 and then remained constant, while the combined PMSG+HCG treatment induced a peak response between Days 24 and 28. The percentage of animals responding also varied with age and treatment. After the combined PMSG+HCG treatment, 90% of the animals ovulated on Day 21, while a similar proportion was not achieved in response to HCG alone until Day 32. The variation in response with age and treatment was related to follicular development within the ovary.     

Ovarian toxicity of hexachlorobenzene (HCB) in the superovulated female rat

Hexachlorobenzene (HCB) is a persistent environmental contaminant which has been measured in human serum, fat, semen, and follicular fluid. In animal testing HCB has been shown to be a reproductive toxin. Discrepant results were obtained from prior studies concerning the effect of HCB treatment on ovarian steroidogenesis. The current study was designed to assess the impact of HCB on the ovary and gonadal steroid levels in the superovulated rat. Female Sprague-Dawley rats (n = 24) were dosed with HCB (0.0, 1.0, 10.0, or 100.0 mg/kg BW/day) for 21 days. All rats received 10 IU pregnant mare serum gonadotropin (PMSG) s.c. on day 18 of treatment and 15 IU of human chorionic gonadotropin (hCG) on day 20. A terminal blood sample was collected and circulating levels of estradiol (Emr₂) and progesterone. (P₄) were determined. Serum concentrations of P₄ were significantly (p < 0.0034) elevated by HCB treatment at all dose levels. Ovarian weights were significantly increased (p < 0.05) in the lowest dose group only compared to the control group. Serum concentrations of E_v uterine weight, weight gain, and general animal health were not affected by HCB treatment. We conclude that during HCB treatment the rat ovary remains responsive to gonadotropin stimulation. Moreover, it is suggested that HCB effects on ovarian steroidogenesis are indirect.     

Analysis of aneuploidy rate in antral and ovulated mouse oocytes during female aging

Two forms of oocytes termed SN (surrounded nucleolus) and NSN (nonsurrounded nucleolus) differing for the spatial distribution of nuclear and nucleolar-associated chromatin have been described within the antral compartment of the ovary of a number of mammals. The biological significance of these two kind of oocytes is as yet not completely clear. In previous studies we have shown that prior to ovulation, mouse SN oocytes isolated from the antral compartment, matured and fertilized in vitro have a far better meiotic and developmental competence than NSN oocytes. Immediately after ovulation SN and NSN oocytes remaining in the antral compartment do not develop beyond the 2-cell stage. To further examine the correlation between chromatin distribution and meiotic competence of mouse antral oocytes, in the present study we have analyzed chromosome segregation at the first meiotic division in antral (SN and NSN) and in ovulated oocytes. SN and NSN oocytes were isolated before (48 h post PMSG injection) or after (15 h post–hCG injection) ovulation from ovaries of females of increasing age, they were cultured in vitro to metaphase II, and their aneuploidy rate was examined. Comparison of data obtained before and after ovulation highlights two main points: 1. Following ovulation a statistically significant increase of aneuploidy is observed in antral oocytes in most age groups and it is attributable to SN oocytes. 2. The aneuploidy rate of ovulated oocytes does not increase during female aging. We have found a correlation between chromatin distribution, hormonal status, and the incidence of aneuploidy during the oocyte first meiotic division. Mol. Reprod. Dev. 50 :305–312, 1998. © 1998 Wiley-Liss, Inc.     

Cellular localization of ovarian proopiomelanocortin messenger RNA during follicular and luteal development in the rat

Opioid peptides are expressed in the reproductive system and have been reported to regulate reproductive function. The present study used in situ hybridization to selectively localize ovarian cells containing high levels of proopiomelanocortin (POMC) mRNA, an opioid precursor, during different stages of ovarian development. Prepubertal rats were primed with PMSG to stimulate follicular development, followed by hCG to induce ovulation. Treatment groups consisted of control (no treatment), PMSG (2 days post-PMSG), 1 day corpus luteum (CL; 1 day post-hCG), and 8 day CL (8 days post-hCG). POMC mRNA-containing cells were present in antral follicles, CL, and the interstitial compartment. With gonadotropin treatment, the percentage of follicles containing heavily labeled cells increased in the PMSG and 1 day CL groups. The number of POMC mRNA-containing cells per follicle also increased in the 1 day CL group. In the CL, no difference was observed in the percentage of CL exhibiting labeled cells between the 1 day CL and 8 day CL groups; however, more labeled luteal cells per CL were present in the 1 day CL group. A marked increase in POMC mRNA-containing cells was observed in the interstitial compartment of the 1 day CL group. These results indicate that the number of POMC mRNA-containing cells increases with follicular development and CL formation; however, the ovarian distribution suggests that the labeled cells could be nonendocrine cells, possibly white blood cells. The in situ hybridization findings are indicative of low total concentrations of ovarian POMC mRNA, suggesting mainly an autocrine or paracrine role for POMC or POMC-derived peptides.(ABSTRACT TRUNCATED AT 250 WORDS)     

Normal oxygen atmosphere is essential for the solitary long-term culture of early preantral mouse follicles

This study compares the effects of reduced (5%) or normal (5% CO₂ in air; 20% O₂) oxygen tension on the in vitro maturation of early preantral ovarian follicles isolated from 14-day-old (C57BI/6J × CBAca) F1 mice. Intact follicles (100–130 μm) are singly cultured in 20 μl droplets α-MEM enriched with FCS and rFSH under mineral oil at 37°C and 100% humidity. In this culture system the follicles are allowed to attach to the bottom of the petri dishes. Follicle in vitro growth, hormone secretory capacity, and in vitro ovulation were studied under the two oxygen tensions. Spontaneous oocyte release from the follicle during a 16-day culture period was observed significantly more under 5% oxygen. Antrallike cavity formation was not observed under 5% O₂. The follicles in the 5% O₂ cultures reaching day 16 were stripped of their granulosa cell layers, and 83% of the retrieved oocytes had already undergone spontaneous germinal-vesicle breakdown (GVBD). Under 20% O₂, the GV stage was maintained until day 16 in 77% of the oocytes. Under 5% O₂, intact follicle survival up to day 12 was significantly reduced as compared to the 5% CO₂ in air conditioning. The hCG stimulus on day 12 induced mucification in a significantly larger proportion of follicles cultured under 20% O₂ (79% vs. 47%). Germinal-vesicle breakdown (20% O₂:95%, 5%, O₂:42%) and first polar body extrusion (20% O₂:40%, 5% O₂:15%) were significantly more prevalent under normal oxygen tension. A reduced secretory capacity of E₂ and inhibin was demonstrated for follicles cultured under 5% O₂. The histological study of serially sectioned follicles showed increased areas of centrally located granulosa cell necrosis and pyknosis in the cumulus cells. Gassing follicle cultures using 5% CO₂ in air provided appropriate conditions for normal growth, enhanced whole-follicle survival, differentiation, and hormone production, and improved the yield of meiotic competent oocytes. © 1996 Wiley-Liss, Inc.     

Effects of oxygen tension and follicle cells on maturation and fertilization of porcine oocytes during in vitro culture in follicular fluid

The objective was to investigate the effects of oxygen tension and follicle cells (FCs) during in vitro maturation of porcine oocytes in only porcine (Sus scrofa domesticus) follicular fluid (pFF), using static and non-static (rotating) culture systems, on the nuclear maturation and subsequent in vitro fertilization of the oocytes. In the first experiment, cumulus-oocyte complexes (COCs) were matured for 48 h in pFF supplemented with (+) or without (−) FCs (5.2 × 10⁶ cells/mL), using the static (S) and rotating (R) culture systems (+FC/S, −FC/S, +FC/R, and −FC/R) under 5% or 20% O₂. Co-culture with FCs in the static culture system (+FC/S) had a detrimental effect on the meiotic competence of oocytes, whereas co-culture with FCs in the rotating culture system (+FC/R) increased maturation rates. In both culture systems, oxygen tension had no apparent effects on meiotic competence of oocytes, irrespective of culture system and FC addition. In the second experiment, COCs were matured under 5% or 20% O₂ using the −FC/S or +FC/R culture systems and then fertilized. Oxygen tension had no significant effects on fertilization parameters, irrespective of the culture system. The rotating culture system increased rates of sperm penetration and male pronuclear formation and decreased polyspermic fertilization compared with the static culture system (P < 0.05). In conclusion, both −FC/S and +FC/R culture systems supported meiotic competence, irrespective of oxygen tension. However, the +FC/R culture system may be superior to the −FC/S culture system for promoting fertilization.     

Hypoxanthine (HX) inhibition of in vitro meiotic resumption in goat oocytes

To improve in vitro maturation and to understand the mechanism for meiotic resumption of oocytes, meiotic progression, and its control by hypoxanthine (HX) were studied in goat oocytes. Ovaries were obtained from a local abattoir, and cumulus-oocyte complexes (COCs) and follicular fluid were collected from follicles of different surface diameters (SDs). The meiotic competence and progression of oocytes were observed, and the concentration of HX in the follicular fluid and culture media was measured by high-performance liquid chromatography (HPLC). Full meiotic competence of goat oocytes was acquired in follicles of >/=1.5 mm in SD with 90% of the oocytes developing to metaphase II (MII) stage after 24 hr in culture. The HX concentration in follicular fluid decreased with follicle development, from the highest level of 1.16 mM in /=5 mm follicles. HX inhibited meiotic resumption of goat oocytes in a concentration-related manner but this inhibitory effect declined gradually. When we renewed the medium at 4 hr of HX-199 (TCM-199 supplemented with 4 mM HX) culture, the percentage of oocytes with intact germinal vesicle (GV) did not increase but decreased significantly instead. HPLC measurement of HX in the HX-199 culture drops indicated that the HX concentration declined from 0 hr to 4 hr of culture and after medium renewal at 4 hr of culture. By adding dibutyryl cAMP (db-cAMP) at medium renewal, we found that db-cAMP held up the decline of GV percentages. Together, these results were consistent with the possibility that the decline of HX inhibitory effect was not due to HX depletion but rather due to the negative feedback of the metabolites on its further uptake by oocytes. Goat oocytes were capable of normal nuclear maturation and activation after temporal arrest by HX, but prolonged exposure to HX induced spontaneous activation.     

Pyrethroids cypermethrin,deltamethrin and fenvalerate have different effects on in vitro maturation of pig oocytes at different stages of growth

Pesticides can significantly harm reproduction in animals and people. Pyrethroids are often used as insecticides, and their toxicity for mammals is considered to be low. However, cypermethrin, deltamethrin and fenvalerate – as potent specific inhibitors of protein phosphatase calcineurin – can influence the meiosis of mammalian oocytes. The objective of this study was to evaluate the effects of these pyrethroids on the in vitro maturation of pig oocytes at different levels of meiotic competence. Under the tested concentrations, cypermethrin, deltamethrin and fenvalerate neither had a significant effect on the viability of oocytes nor did they induce significant degeneration of oocytes. However, these pyrethroids significantly affected meiotic maturation. The effects depended on the stage of meiotic competence of the oocytes. Maturation of growing pig oocytes with partial meiotic competence was induced. On the other hand, in fully grown pig oocytes with full meiotic competence, maturation in vitro was delayed. The specificity of these effects was further supported by the same effect of non-pyrethroidal inhibitors of calcineurin – cyclosporin A or hymenistatin I – on the maturation of oocytes with different levels of meiotic competence. However, pyrethroids, which do not inhibit calcineurin – allethrin or permethrin – had no effect on pig oocyte maturation. We demonstrated a significant effect of pyrethroids on the maturation of mammalian oocytes under in vitro conditions. This indicates that exposure to these substances could affect the fertility of people or animals.     

Size of the donor follicle, but not stage of reproductive cycle or seasonality, influences meiotic competency of selected domestic dog oocytes

Ability of ovarian oocytes from the domestic dog to complete nuclear maturation in vitro (IVM) varies markedly among donors and generally is 20% or less of all oocytes cultured. To identify the cause(s) underlying these significant variations in meiotic maturation (to metaphase II; MII), we retrospectively analyzed data from 1,643 oocytes recovered from 90 bitches for which stage of reproduction and season of year were known. Neither stage of reproduction (proestrus/estrus, diestrus, anestrus, or prepuberty) nor season (P > 0.05) influenced the ability of oocytes to achieve nuclear maturation in vitro. A second study was conducted to examine the impact of follicular size on meiotic maturation. Populations of large oocytes were recovered from four categories of follicles (ranging from <0.5 to > 2 mm in diameter) and cultured in TCM 199 for 48 hr. Follicular size influenced (P < 0.05) meiotic competence. Mean percentages of MII oocytes were 16.9 +/- 9.2, 26.1 +/- 7.6, 38.4 +/- 9.2, and 79.5 +/- 10.9 for oocytes recovered from < 0.5 mm, > or = 0.5-< 1 mm, 1-2 mm, and > 2 mm diameter follicles, respectively. In summary, stage of reproduction and season have no impact on the ability of dog oocytes to achieve nuclear maturation in vitro. However, we demonstrated for the first time that dog oocytes acquire meiotic competency during follicular development. IVM success of selected oocytes from large size follicles (almost 80%) is about 60% higher than measured in most previous studies involving randomly collected oocytes.     

Exposure to bisphenol A disrupts meiotic progression during spermatogenesis in adult rats through estrogen-like activity

The effect of bisphenol A (BPA) on the reproductive system is highly debated but has been associated with meiotic abnormalities. However, evidence is lacking with regard to the mechanisms involved. In order to explore the underlying mechanisms of BPA-induced meiotic abnormalities in adult male rats, we exposed 9-week-old male Wistar rats to BPA by gavage at 0, 2, 20 or 200 μg/kg body weight (bw)/day for 60 consecutive days. 17β-Estradiol (E₂) was administered at 10 μg/kg bw/day as the estrogenic positive control. Treatments with 200 μg/kg bw/day of BPA and E₂ significantly decreased sperm counts and inhibited spermiation, characterized by an increase in stage VII and decrease in stage VIII in the seminiferous epithelium. This was concomitant with a disruption in the progression of meiosis I and the persistence of meiotic DNA strand breaks in pachytene spermatocytes,and the ataxia–telangiectasia-mutated and checkpoint kinase 2 signal pathway was also activated; Eventually, germ cell apoptosis was triggered as evaluated by terminal dUTP nick-end labeling assay and western blot for caspase 3. Using the estrogen receptor (ER) antagonist ICI 182780, we determined that ER signaling mediated BPA-induced meiotic disruption and reproductive impairment. Our results suggest that ER signaling-mediated meiotic disruption may be a major contributor to the molecular events leading to BPA-related male reproductive disorders. These rodent data support the growing association between BPA exposure and the rapid increase in the incidence of male reproductive disorders.     

Reversal of indomethacin blockade of ovulation in gilts by prostaglandins

Prepubertal gilts given 750 IU pregnant mares′ serum gonadotropin (PMSG) followed 72 h later by 500 IU human chorionic gonadotropin (hCG) to induce follicular growth and ovulation fail to ovulate when 10 mg/kg indomethacin (INDO) is injected 24 h after hCG administration. This study examines the effects of administration of exogenous prostaglandins F_2α and E₂ (PGF_2α and PGE₂) alone or in combination, and at various times prior to the expected time of ovulation, on the INDO blockade of ovulation in PMSG/hCG-treated gilts. Occurrence of ovulation was determined by visual observation at laparotomy 48 h after hCG. When 5 mg or 10 mg PGF_2α was injected at each of 38, 40 and 42 h after hCG injection, 63% and 79%, respectively, of preovulatory follicles ovulated. In contrast, injection of 5 mg PGE₂ or 5 mg PGE₂ plus 5 mg PGF_2α induced ovulation in 0% and 24% of preovulatory follicles, respectively. In control groups, 100% of folicles in PMSG/hCG-treated gilts ovulated whereas none did so in PMSG/hCG/INDO-treated animals. These results indicate that administration of PGF_2α can induce ovulation in the PMSG/hCG/INDO-treated prepubertal gilt and suggest that PGE₂ is ineffective and may be antagonistic to PGF_2α in overcoming the ovulation blocking effect of INDO.     

Influence of equine growth hormone,insulin-like growth factor-I and its interaction with gonadotropins on in vitro maturation and cytoskeleton morphology in equine oocytes

In horses, successful in vitro fertilization procedures are limited by our inability to consistently mature equine oocytes by in vitro methods. Growth hormone (GH) is an important regulator of female reproduction in mammals, playing an important role in ovarian function, follicular growth and steroidogenesis. The objectives of this research were to investigate: the effects of equine growth hormone (eGH) and insulin-like growth factor-I (IGF-I) on the in vitro maturation (IVM) of equine oocytes, and the effects of eGH in addition to estradiol (E₂), gonadotropins (FSH and LH) and fetal calf serum (FCS) on IVM. We also evaluated the cytoskeleton organization of equine oocytes after IVM with eGH. Equine oocytes were aspirated from follicles <30 mm in diameter and matured for 30 h at 38.5°C in air with 5% CO₂. In experiment 1, selected cumulus–oocyte complexes (COCs) were randomly allocated as follows: (a) control (no additives); (b) 400 ng/ml eGH; (c) 200 ng/ml IGF-I; (d) eGH + IGF-I; and (e) eGH + IGF-I + 200 ng/ml anti-IGF-I. In addition to these treatment groups, we also added 1 μg/ml E₂, 5 IU/ml FSH, 10 IU/ml LH and 10% FCS in vitro (experiment 2). Oocytes were stained with markers for microtubules (anti-α-tubulin antibody), microfilaments (AlexaFluor 488 Phalloidin) and chromatin (TO-PRO₃-iodide) and assessed via confocal microscopy. No difference was observed when eGH and IGF-I was added into our IVM system. However, following incubation with eGH alone (40%) and eGH, E₂, gonadotropins and FCS (36.6%) oocytes were classified as mature v. 17.6% of oocytes in the control group (P < 0.05). Matured equine oocytes showed that a thin network of filaments concentrated within the oocyte cortex and microtubules at the metaphase spindle showed a symmetrical barrel-shaped structure, with chromosomes aligned along its midline. We conclude that the use of E₂, gonadotropins and FCS in the presence of eGH increases the number of oocytes reaching oocyte competence.     

Oestradiol-17β plasma concentrations after intramuscular injection of oestradiol benzoate or oestradiol cypionate in llamas (<Emphasis Type="Italic">Lama glama</Emphasis>)

Background  

Llamas (Lama glama) are induced ovulators and the process of ovulation depends on dominant follicular size. In addition, a close relationship between behavioural estrus and ovulation is not registered in llamas. Therefore, the exogenous control of follicular development with hormones aims to predict the optimal time to mate. Oestradiol-17β (E₂) and its esters are currently used in domestic species, including camelids, in synchronization treatments. But, in llamas, there is no reports regarding the appropriate dosages to be used and most protocols have been designed by extrapolation from those recommended for other ruminants. The aim of the present study was to characterize plasma E₂ concentrations in intact female llamas following a single intramuscular (i.m.) injection of two oestradiol esters: oestradiol benzoate (EB) and oestradiol cypionate (ECP).     

Periodic 17β-Estradiol Pretreatment Protects Rat Brain from Cerebral Ischemic Damage via Estrogen Receptor-β

Although chronic 17β-estradiol (E₂) has been shown to be a cognition-preserving and neuroprotective agent in animal brain injury models, concern regarding its safety was raised by the failed translation of this phenomenon to the clinic. Previously, we demonstrated that a single bolus of E₂ 48 hr prior to ischemia protected the hippocampus from damage in ovariectomized rats via phosphorylation of cyclic-AMP response element binding protein, which requires activation of estrogen receptor subtype beta (ER-β). The current study tests the hypothesis that long-term periodic E₂-treatment improves cognition and reduces post-ischemic hippocampal injury by means of ER-β activation. Ovariectomized rats were given ten injections of E₂ at 48 hr intervals for 21 days. Hippocampal-dependent learning, memory and ischemic neuronal loss were monitored. Results demonstrated that periodic E₂ treatments improved spatial learning, memory and ischemic neuronal survival in ovariectomized rats. Additionally, periodic ER-β agonist treatments every 48 hr improved post-ischemic cognition. Silencing of hippocampal ER-β attenuated E₂-mediated ischemic protection suggesting that ER-β plays a key role in mediating the beneficial effects of periodic E₂ treatments. This study emphasizes the need to investigate a periodic estrogen replacement regimen to reduce cognitive decline and cerebral ischemia incidents/impact in post-menopausal women.     

Reactive oxygen and nitrogen species during meiotic resumption from diplotene arrest in mammalian oocytes

Mammalian ovary is metabolically active organ and generates by‐products such as reactive oxygen species (ROS) and reactive nitrogen species (RNS) on an extraordinary scale. Both follicular somatic cells as well as oocyte generate ROS and RNS synchronously and their effects are neutralized by intricate array of antioxidants. ROS such as hydrogen peroxide (H₂O₂) and RNS such as nitric oxide (NO) act as signaling molecules and modulate various aspects of oocyte physiology including meiotic cell cycle arrest and resumption. Generation of intraoocyte H₂O₂ can induce meiotic resumption from diplotene arrest probably by the activation of adenosine monophosphate (AMP)‐activated protein kinase A (PRKA)—or Ca²⁺‐mediated pathway. However, reduced intraoocyte NO level may inactivate guanylyl cyclase‐mediated pathway that results in the reduced production of cyclic 3′,5′‐guanosine monophosphate (cGMP). The reduced level of cGMP results in the activation of cyclic 3′,5′‐adenosine monophosphate (cAMP)‐phosphodiesterase 3A (PDE3A), which hydrolyses cAMP. The reduced intraoocyte cAMP results in the activation of maturation promoting factor (MPF) that finally induces meiotic resumption. Thus, a transient increase of intraoocyte H₂O₂ level and decrease of NO level may signal meiotic resumption from diplotene arrest in mammalian oocytes. J. Cell. Biochem. 111: 521–528, 2010. © 2010 Wiley‐Liss, Inc.