scholarly journals The role of nitric oxide in parthenogenetic activation of pig oocytes: A review

2008 ◽  
Vol 52 (No. 11) ◽  
pp. 363-377 ◽  
Author(s):  
J. Petr ◽  
E. Chmelíková ◽  
L. Tůmová ◽  
M. Ješeta
Keyword(s):  
Nitric Oxide ◽  
Calcium Ionophore ◽  
Signalling Pathways ◽  
Oocyte Activation ◽  
Parthenogenetic Activation ◽  
Complex Processes ◽  
Parthenogenetic Development ◽  
Adequate Stimulus ◽  
Signalling Cascade

Parthenogenetic activation of mammalian oocytes with artificial stimuli is commonly applied in various reproductive biotechniques, e.g. cloning using nuclear transfer. For this reason, many studies focus on oocyte activation <I>in vitro</I>. Recently we have described the activation of pig oocytes using nitric oxide. This activating stimulus is very specific in many aspects. However, it does not provide an adequate stimulus for parthenogenetic development. It was shown that nitric oxide stimulated some signalling pathways which are inactive in conventional treatments for parthenogenetic activation, e.g. the cGMP-dependent signalling cascade. On the other hand, nitric oxide does not stimulate certain signalling pathways involved in oocyte activation after calcium ionophore, e.g. the PKC signalling cascade. The aim of this review is to characterize the complex processes induced in oocytes after treatment with nitric oxide. Perspectives for further research and the application of nitric oxide for parthenogenetic activation of oocytes are outlined.

253. Calcium ionophore induction of marmoset oocyte activation

2005 ◽  
Vol 17 (9) ◽  
pp. 101
Author(s):  
C. G. Grupen ◽  
R. B. Gilchrist ◽  
S. J. Schulz ◽  
L. J. Ritter ◽  
D. T. Armstrong
Keyword(s):  
Assisted Reproductive Technologies ◽  
Cumulus Cell ◽  
Calcium Ionophore ◽  
Reproductive Technologies ◽  
Primate Species ◽  
Oocyte Activation ◽  
Cell Stage ◽  
Developmental Potential ◽  
Parthenogenetic Activation

The marmoset monkey (Callithrix jacchus) is a valuable model for developing assisted reproductive technologies in humans and endangered primate species. Calcium ionophore treatments have been used to induce parthenogenetic activation in a number of species, but the effectiveness of this reagent in initiating marmoset embryo development has not yet been reported. The aim of this study was to determine the developmental potential of in vitro matured (IVM) marmoset oocytes, following treatment with calcium ionophore. Immature oocytes from large (LA; >1.5 mm) and small (SA; 0.67–1.5 mm) antral follicles were isolated from the ovaries of FSH-primed animals and cultured in modified G2 medium for 26–30 h at 37.0°C in 6% CO2 in air. Meiotically mature oocytes were sequentially incubated with 5 μM ionomycin for 5 min and 2 mM 6-dimethylaminopurine for 3 h and cultured in G1/G2 sequential medium at 37.0°C in 5% O2, 6% CO2, 89% N2 for 10 days. Cumulus cell expansion associated with LA oocytes (n=118) was greater than that of SA oocytes (n=212), as determined using well established classification criteria (2.7±0.1 v. 1.8±0.2; P<0.01). A greater proportion of LA oocytes completed meiosis to the metaphase-II stage compared with SA oocytes (85±7% v. 63±7%; P<0.05). Pronuclear formation was induced at similar rates in mature oocytes of both groups, but the rate of cleavage was higher for LA oocytes compared with SA oocytes (93±6% v. 66±5%; P<0.05). The number of cells per embryo was not different between the groups.This is the first study to demonstrate that calcium ionophore effectively induces parthenogenetic activation in IVM marmoset oocytes. However, the development of parthenotes was limited beyond the 8-cell stage. Further studies are needed to determine the cause of the developmental block.

scholarly journals Effects of protein kinase C on parthenogenetic activation of pig oocytes using calcium ionophore or nitric oxide-donor

2008 ◽  
Vol 52 (No. 12) ◽  
pp. 415-422 ◽  
Author(s):  
J. Petr ◽  
E. Chmelíková ◽  
A. Dörflerová ◽  
M. Ješeta ◽  
Z. Kuthanová
Keyword(s):  
Nitric Oxide ◽  
Protein Kinase C ◽  
Protein Kinase ◽  
Calcium Ionophore ◽  
Nitric Oxide Donor ◽  
Oocyte Activation ◽  
Parthenogenetic Activation ◽  
Kinase C ◽  
Activation Rate ◽  
Pkc Inhibitors

Porcine oocytes matured <I>in vitr</I>o were activated for parthenogenetic development using either calcium ionophore (50&mu;M for 10 min) or nitric oxide donor SNAP (2mM for 23.5 hours). Protein kinase C (PKC) inhibitors, bisindolylmaleimide I or rottlerin, are able to inhibit parthenogenetic activation induced by calcium ionophore. The rate of activated oocytes decreased from 69% to 2% (<I>P</I> < 0.05) under the effect of bisindolylmaleimide I at a concentration of 0 or 20nM, respectively. The activation rate decreased from 68% to 0% (<I>P</I> < 0.05) under the influence of 0 or 20&mu;M rottlerin, respectively. PKC inhibitors Go6976 or hispidin had no effect on the oocyte activation using calcium ionophore or on oocytes activated by a nitric oxide donor. The activation of oocytes by a nitric oxide donor is not significantly influenced even under the effects of bisindolylmaleimide I or rottlerin. Based on these data we can conclude that the oocyte activation induced by calcium ionophore depends on PKC, especially on PKC-δ. On the other hand, the oocyte activation induced by nitric oxide is independent of the tested isotypes of PKC.

scholarly journals Effects of cycloheximide or 6-dimethyl aminopurine on the parthenogenetic activation of pig oocytes using pulsatile treatment with nitric oxide donor

2009 ◽  
Vol 54 (No. 7) ◽  
pp. 293-306 ◽  
Author(s):  
T. Krejčová ◽  
J. Petr ◽  
M. Krejčová ◽  
K. Kheilová
Keyword(s):  
Nitric Oxide ◽  
Kinase Inhibitor ◽  
Calcium Ionophore ◽  
Continuous Treatment ◽  
Nitric Oxide Donor ◽  
Activation Rate ◽  
Morula Stage ◽  
Inhibitor Of Protein Synthesis ◽  
Parthenogenetic Embryos

Pig oocytes matured <I>in vitro</I> were parthenogenetically activated using nitric oxide donor SNAP (2mM). Continuous treatment successfully activated the oocytes only after more than 12 hours of exposure. Pulsatile treatments during which oocytes were repeatedly exposed to 2mM SNAP for a short time (10, 20 or 30 minutes) were more efficient with regard to the activation rate, even when the total exposure time did not exceed 4 hours. Parthenogenetic development was very limited after continuous treatment with 2mM SNAP. A significantly higher proportion of developing parthenogenetic embryos was observed after the pulsatile treatment (development to the morula stage 0 vs. 18%; development to the blastocyst 0 vs. 7%; <I>P</I> < 0.05). However, this developmental rate was significantly lower (<I>P</I> < 0.05) than the development induced by conventional activation treatment with calcium ionophore (development to the morula stage, 23%; development to the blastocyst stage, 18%). When we combined pulsatile SNAP-treatment with the effect of protein kinase inhibitor 6-dimethyl aminopurine (6-DMAP) (2mM 6-DMAP for 2 hours) or with the inhibitor of protein synthesis cycloheximide (CHX) (10 µM CHX for 2 hours), we observed a significant increase (<I>P</I> < 0.05) in the activation rate when compared to the respective pulsatile SNAP-treatment without 6-DMAP or CHX (63 vs. 78% of activated oocytes for 6-DMAP; 63 vs. 83% of activated oocytes for CHX). However, the development of parthenogenetic embryos was not enhanced when the pulsatile SNAP-treatment was combined with 6-DMAP or with CHX.

scholarly journals Improved development by Taxol pretreatment after vitrification of in vitro matured porcine oocytes

Reproduction ◽  
2006 ◽  
Vol 131 (4) ◽  
pp. 795-804 ◽  
Author(s):  
Wen-Qing Shi ◽  
Shi-En Zhu ◽  
Dong Zhang ◽  
Wei-Hua Wang ◽  
Guo-Liang Tang ◽  
...  
Keyword(s):  
Survival Rate ◽  
Embryo Development ◽  
Survival Rates ◽  
Developmental Competence ◽  
Fluorescein Diacetate ◽  
Microtubule Organization ◽  
Cell Stage ◽  
Parthenogenetic Activation ◽  
Parthenogenetic Development

This study was designed to examine the effect of Taxol pretreatment on vitrification of porcine oocytes matured in vitro by an open pulled straw (OPS) method. In the first experiment, the effect of Taxol pretreatment and fluorescein diacetate (FDA) staining on parthenogenetic development of oocytes was evaluated. In the second experiment, viability, microtubule organization and embryo development of oocytes were assessed after oocytes were exposed to vitrification/warming solutions or after vitrification with or without Taxol pretreatment. The results showed that Taxol pretreatment and/or FDA staining did not negatively influence the oocyte’s developmental competence after parthenogenetic activation. After being exposed to vitrification/warming solutions, the survival rate (83.3%) of the oocytes was significantly (P < 0.05) reduced as compared with that in the control (100%). Vitrification/warming procedures further reduced the survival rates of oocytes regardless of oocytes being treated with (62.1%) or without (53.8%) Taxol. The proportions of oocytes with normal spindle configuration were significantly reduced after the oocytes were exposed to vitrification/warming solutions (38.5%) or after vitrification with (10.3%) or without (4.1%) Taxol pretreatment as compared with that in control (76.8%). The rates of two-cell-stage (5.6–53.2%) embryos at 48 h and blastocysts (0–3.8%) at 144 h after activation were significantly reduced after exposure to vitrification/warming solutions or after vitrification as compared with control (90.9% and 26.6% respectively). However, the proportion of vitrified oocytes developed to two-cell stage was significantly higher when oocytes were pretreated with (24.3%) than without (5.6%) Taxol. These results indicate that pretreatment of oocytes with Taxol before vitrification helps to reduce the damage induced by vitrification and is a potential way to improve the development of vitrified porcine oocytes.

scholarly journals Reactive oxygen species production and redox state in parthenogenetic and sperm-mediated bovine oocyte activation

Reproduction ◽  
2013 ◽  
Vol 145 (5) ◽  
pp. 471-478 ◽  
Author(s):  
S Morado ◽  
P Cetica ◽  
M Beconi ◽  
J G Thompson ◽  
G Dalvit
Keyword(s):  
Reactive Oxygen Species ◽  
Embryo Development ◽  
Redox State ◽  
Reactive Oxygen ◽  
Developmental Competence ◽  
Ros Production ◽  
Oxygen Species ◽  
Oocyte Activation ◽  
Parthenogenetic Activation

The knowledge concerning redox and reactive oxygen species (ROS)-mediated regulation of early embryo development is scarce and remains controversial. The aim of this work was to determine ROS production and redox state during early in vitro embryo development in sperm-mediated and parthenogenetic activation of bovine oocytes. Sperm-mediated oocyte activation was carried out in IVF-modified synthetic oviductal fluid (mSOF) with frozen–thawed semen. Parthenogenetic activation was performed in TALP plus ionomycin and then in IVF-mSOF with 6-dimethylaminopurine plus cytochalasin B. Embryos were cultured in IVF-mSOF. ROS and redox state were determined at each 2-h interval (7–24 h from activation) by 2′,7′-dichlorodihydrofluorescein diacetate and RedoxSensor Red CC-1 fluorochromes respectively. ROS levels and redox state differed between activated and non-activated oocytes (P<0.05 by ANOVA). In sperm-activated oocytes, an increase was observed between 15 and 19 h (P<0.05). Conversely, in parthenogenetically activated oocytes, we observed a decrease at 9 h (P<0.05). In sperm-activated oocytes, ROS fluctuated throughout the 24 h, presenting peaks around 7, 19, and 24 h (P<0.05), while in parthenogenetic activation, peaks were detected at 7, 11, and 17 h (P<0.05). In the present work, we found clear distinctive metabolic patterns between normal and parthenogenetic zygotes. Oxidative activity and ROS production are an integral part of bovine zygote behavior, and defining a temporal pattern of change may be linked with developmental competence.

scholarly journals 273 OPTIMIZATION OF PROTOCOLS FOR ACTIVATION OF GOAT OOCYTES WITH IONOMYCIN IN COMBINATION WITH 6-DIMETHYLAMINOPURINE

2005 ◽  
Vol 17 (2) ◽  
pp. 286
Author(s):  
J.-H. Tan ◽  
G.-C. Lan ◽  
Z.-L. Chang ◽  
D. Han ◽  
Z.-B. Han ◽  
...  
Keyword(s):  
Laser Scanning ◽  
Cumulus Cell ◽  
Laser Scanning Confocal Microscopy ◽  
Oocyte Activation ◽  
Scanning Confocal Microscopy ◽  
Multiple Comparison Test ◽  
Parthenogenetic Development ◽  
Activation Rate ◽  
Activation Rates

The protocol of ionomycin in combination with 6-dimethylaminopurine (6-DMAP) is commonly used for activation of oocytes and reconstructed embryos of different species. Since numerous abnormalities and impaired development have been observed when oocytes are activated with 6-DMAP, this protocol needs optimization. Effects of concentration and treatment duration of both drugs on activation kinetics and parthenogenetic development of goat oocytes were examined in this study. When goat oocytes matured in vitro in TCM-199 were treated with 5 M ionomycin in PBS for different periods before exposure to 6-DMAP in CR1aa, the activation rate obtained with ionomycin treatment for 1 min (95.2%) was significantly (P < 0.05, Duncan multiple comparison test) higher than with ionomycin treatments for 3, 5, 7, or 9 min. When oocytes were treated with different concentrations of ionomycin for 1 min before exposure to 6-DMAP, activation rates obtained with 0.625, 1.25, 2.5, 5, 10, and 20 M ionomycin (87–95%) did not differ significantly but were significantly higher than that achieved with 0.3125 M ionomycin. Progressive reduction of time for 6-DMAP exposure showed that the duration of 6-DMAP treatment can be reduced to 1 h from the 2nd up to the 4th hour after ionomycin treatment, to produce activation rates greater than 85%. When oocytes were treated with different concentrations of 6-DMAP for the 3rd hour (atotal of 1 h, 3 h after the exposure to ionomycin), activation rates with 4 and 2 mM 6-DMAP (>90%) were significantly higher than those with 1and 0.5 mM. Therefore, the best protocol for goat oocyte activation would be a 1-min exposure to 2.5 M ionomycin followed by 2 mM 6-DMAP treatment for the 3rd hour. When oocytes matured in vitro for different times were stimulated with the best protocol, activation rates of the 27-, 30-, and 33-h oocytes (85, 85, and 91%) were significantly higher than those of the 24-, 26-, and 39-h oocytes. When activated oocytes were co-cultured in CR1aa with cumulus cell monolayers, the highest rates of cleavage (92%) and morulae/blastocysts (23%) were obtained with oocytes activated by the best protocol, and any increase in the intensity of ionomycin treatment and in the duration of 6-DMAP exposure impaired the development of the parthenotes. During anaphase II, chromosomes (the dyads) did not separate into two units in oocytes that were activated by long exposure to 6-DMAP, but they did in oocytes that were activated by short or no exposure to 6-DMAP; as a result, only one pronucleus developed in most of the former but two pronuclei were formed in most of the latter cases. Laser scanning confocal microscopy showed that microtubules also behaved differently in these two groups of activated oocytes. It is therefore concluded that to obtain better activation and development, goat oocytes matured in vitro for 27–33 h should be used, and these should be activated by a 1-min exposure to 2.5 M ionomycin followed by 2 mM 6-DMAP treatment for the 3rd hour. This study was supported by the “973” Project of China Sci. Technol. Ministry (No. G200016108).

Increased superoxide anion generation and altered vasoreactivity in rabbits on low-potassium diet

1998 ◽  
Vol 274 (6) ◽  
pp. H1955-H1961 ◽  
Author(s):  
B. C. Yang ◽  
D. Y. Li ◽  
Y. F. Weng ◽  
J. Lynch ◽  
C. S. Wingo ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Carotid Artery ◽  
Superoxide Anion ◽  
Control Diet ◽  
Calcium Ionophore ◽  
Potassium Depletion ◽  
Anion Formation ◽  
Urinary Potassium ◽  
Low Potassium

Potassium reduces blood pressure in populations at high risk of developing hypertension, which suggests that potassium depletion may increase vascular resistance. This study was designed to examine the effect of potassium depletion on thel-arginine-nitric oxide pathway in arterial tissues. New Zealand White rabbits were fed either a control diet, containing a normal amount of potassium, or a low-potassium diet for 1–3 wk. As expected, the low-potassium diet resulted in reduced serum and urinary potassium levels. Carotid arteries were excised, and their contractile and relaxant responses were determined in vitro. Carotid arterial ring contractile response to norepinephrine was enhanced, and relaxation in response to the endothelium-dependent vasodilators acetylcholine and calcium ionophore A-23187 was attenuated, in rabbits fed low-potassium diet (all P < 0.01 compared with responses in rabbits fed control diet). The vasomotor responses were similarly altered in rabbits fed low-potassium diet for 1 or 3 wk. Both the enhanced contraction and attenuated relaxation were abolished by treatment of arterial rings with superoxide dismutase but not by treatment with l-arginine or indomethacin. Carotid artery rings from rabbits fed the low-potassium diet showed ∼100% greater superoxide anion formation than those from rabbits fed control diet ( P < 0.01), whereas plasma and urinary nitrite levels were similar in both groups of rabbits. These observations indicate that low-potassium diet enhances the sensitivity of the carotid artery to vasoconstrictor stimuli and reduces the sensitivity to endothelium-dependent stimuli. Attenuation of endothelium-dependent relaxation appears to be secondary to increased free radical generation, which may degrade nitric oxide. Altered vasoreactivity may underlie the genesis of hypertension in populations consuming diets low in potassium.

Sheep oocyte activation after intracytoplasmic sperm injection (ICSI)

1998 ◽  
Vol 10 (2) ◽  
pp. 197 ◽  
Author(s):  
M. C. Gomez ◽  
J. W. Catt ◽  
G. Evans ◽  
W. M. C. Maxwell
Keyword(s):  
Calcium Chloride ◽  
Calcium Ionophore ◽  
Fertilization Rate ◽  
Oocyte Activation ◽  
Sham Injection ◽  
Injection Process ◽  
Oviduct Fluid ◽  
Free Media ◽  
Mechanical Injection

The effect of calcium concentration on fertilization and activation was examined in oocytes injected in vitro with sperm. Oocytes were subjected to sperm injection, to sham injection or remained uninjected, and were then cultured for 19 h in bicarbonate-buffered synthetic oviduct fluid (BSOF) without calcium, or containing either calcium chloride or calcium ionophore. There was no difference in fertilization rates after ICSI when oocytes were cultured in vitro in media containing calcium chloride or calcium ionophore but the rate was lower in calcium-free media. There was also no difference in the fertilization rate after ICSI when oocytes were culturedin vivocompared with that observedin vitro in media containing calcium chloride or calcium ionophore. In calcium chloride-treated oocytes, activation was induced by mechanical injection, and in calcium ionophore-treated oocytes, by the ionophore. In uninjected oocytes, calcium itself did not cause oocyte activation. It is concluded that it is possible to induce activation by the injection process, but that manipulation alone is inadequate to cause proper oocyte activation unless calcium is also present. No difference in oocyte activation between ICSI and sham injection was found, indicating that the sperm may play no role in the early events of oocyte activation.

236 PARTHENOGENETIC ACTIVATION OF IN VITRO-MATURED OVINE OOCYTES WITH STRONTIUM

2008 ◽  
Vol 20 (1) ◽  
pp. 197
Author(s):  
J. Zhu ◽  
K. H. S. Campbell
Keyword(s):  
Nuclear Transfer ◽  
Cytochalasin B ◽  
Blastocyst Stage ◽  
Oocyte Activation ◽  
Parthenogenetic Development ◽  
Activation Rate ◽  
Electrical Pulses ◽  
Blastocyst Rate ◽  
Activation Rates

The objective of the present experiments was to examine whether strontium could activate in vitro-matured ovine oocytes. Oocytes were collected and matured as previously described (Lee and Campbell 2006 Biol. Reprod. 74, 691–698). Briefly, selected cumulus–oocyte complexes were cultured in modified TCM-199 medium supplemented with 20% sheep serum and hormones for 22–23 h, at 39°C, 5% CO2 in air. Matured oocytes were randomly divided into four groups and treated as follows: (1) cultured in 10 mm strontium + 5 μg mL–1 cytochalasin B in Ca2+-free CZB medium for 4–5 h; (2) electrically activated in Ca2+-containing medium, then cultured in 10 mm strontium + 5 μg mL–1 cytochalasin B in Ca2+-free CZB medium for 4–5 h; (3) electrically activated in Ca2+-containing medium and then cultured in SOF medium containing 5 μg mL–1 cytochalasin B for 4–5 h; and (4) electrically activated in Ca2+-free medium and then transferred into SOF medium + 5 μg mL–1 cytochalasin B for 4–5 h. This experiment was repeated three times. Activation rates based on the number of pronuclear formations/the number of oocytes cultured were 96.7% (147/152), 95.9% (116/121), 75.9% (101/133), and 43.0% (56/107) in Groups 1–4, respectively. After 7 days of culture in SOF medium, 26.8%, 33.3%, 19.6%, and 0% of oocytes in Groups 1, 2, 3, and 4 developed to the blastocyst stage, respectively. Significant differences in blastocyst rate were observed across these groups except between groups 1 and 2 (P < 0.01). However, there were no significant differences in mean number of nuclei/blastocyst across Groups 1, 2, and 3 (P > 0.05). Our results demonstrated that in vitro-matured ovine oocytes can be effectively activated with strontium alone, resulting in an activation rate of 96.7% and a blastocyst rate of 26.8% (blastocysts/oocytes). Also, a combination of strontium and electrical pulses could benefit sheep oocyte activation and embryo development to the blastocyst stage (95.9% and 33.3%, respectively). We conclude that strontium is an effective activator for sheep oocyte activation and it could be used for sheep nuclear transfer. Table 1. Parthenogenetic development of oocytes activated by SrCl2+ and electrical pulses

284 INTRACELLULAR PATHWAYS MEDIATING DIRECT EFFECTS OF PROLACTIN AND GROWTH HORMONE ON BOVINE METAPHASE-II OOCYTES AGING IN VITRO

2015 ◽  
Vol 27 (1) ◽  
pp. 231 ◽  
Author(s):  
I. Lebedeva ◽  
G. Singina ◽  
A. Lopukhov ◽  
E. Shedova ◽  
N. Zinovieva
Keyword(s):  
Tyrosine Kinases ◽  
Mek Inhibitor ◽  
Control Medium ◽  
Oocyte Activation ◽  
Direct Effects ◽  
Meiotic Arrest ◽  
Parthenogenetic Activation ◽  
Intracellular Pathways ◽  
Src Family Tyrosine Kinases

Maturation of mammalian oocytes coincides with various aging processes, which negatively affect the oocyte quality. Cellular and molecular changes in matured oocytes aging in vivo and in vitro are very similar, suggesting similarities in the underlying mechanisms. The goal of the present research was to study direct effects of prolactin (PRL) and growth hormone (GH) on bovine metaphase-II (M-II) oocytes aging in vitro and intracellular pathways mediating these effects. Cumulus-enclosed oocytes were matured for 20 h in TCM 199 containing 10% fetal calf serum (FCS), 10 µg mL–1 of porcine FSH, and 10 µg mL–1 of ovine LH. After IVM, oocytes were set free from cumulus and denuded oocytes (DO) were cultured for 44 h in the aging medium consisting of TCM 199 supplemented with 10% FCS in the absence of PRL and GH (Control), then in the presence of either 50 ng mL–1 of bovine PRL or 10 ng mL–1 of bovine GH or protein kinase inhibitors. The following inhibitors were added: (1) PP2 (an inhibitor of Src-family tyrosine kinases, 20 µM), (2) U0126 (a MEK inhibitor, 20 µM), and (3) triciribine (an inhibitor of Akt kinase, 50 µM). At the end of culture, the state of the nuclear material of oocytes and embryos was evaluated by the method of Tarkowski. The number of oocytes undergone spontaneous parthenogenetic activation was determined by summarising the numbers of embryos cleaved and oocytes reached anaphase-II to pronucleus stages. The data from 4 replicates were analysed by ANOVA. During aging in the control medium, the rate of M-II oocytes with destructive changes of chromosomes (decondensation, clumping, fragmentation) increased from 20.3 ± 1.7 to 65.1 ± 3.5% (P < 0.001) and was unaffected by PRL and GH. At the same time, the frequency of oocyte parthenogenetic activation (18.7 ± 4.9%) was reduced (P < 0.001) by both PRL and GH (up to 6.0 ± 2.9 and 5.9 ± 2.4%, respectively). The inhibitor of Src-family tyrosine kinases PP2 eliminated (at least P < 0.05) the supporting effects of PRL and GH on the meiotic arrest at M-II. The MEK inhibitor U0126 also abolished the effect of PRL (but not that of GH), increasing the frequency of the oocyte activation from 5.9 ± 2.3 to 21.6 ± 1.9% (P < 0.01). Concurrently, both inhibitors did not affect the meiotic arrest in the control group. The inhibitor of Akt kinase triciribine did not influence the frequency of the oocyte activation in the PRL- and GH-treated groups. Meanwhile, triciribine decreased the rate of M-II oocytes with chromosome abnormalities in the control medium (from 71.2 ± 1.7 to 47.0 ± 2.7%; P < 0.001). When added to the aging medium, PRL enhanced the triciribine effect, while GH suppressed it (P < 0.01). Thus, PRL and GH can directly support the meiotic arrest at M-II in aging oocytes by activating MAP kinases or Src-family tyrosine kinases. Besides, GH may enhance the chromosome destruction in DOs through activation of Akt kinase.The research was supported by RFBR (No. 13-04-01888).

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