254 DYNAMICS OF GOLGI APPARATUS DURING BOVINE OOCYTE IN VITRO MATURATION: EFFECTS OF INHIBITORS ON CDC2A AND CYTOPLASMIC-DYNEIN ATPase ACTIVITY

2009 ◽  
Vol 21 (1) ◽  
pp. 225
Author(s):  
S. E. Racedo ◽  
V. Y. Rawe ◽  
H. Niemann
Keyword(s):  
Golgi Apparatus ◽  
Atpase Activity ◽  
Oocyte Maturation ◽  
In Vitro Maturation ◽  
Germinal Vesicle ◽  
Cytoplasmic Dynein ◽  
Bovine Oocyte ◽  
Germinal Vesicle Breakdown

The process of maturation encompasses a complex series of molecular and structural events. Completion of the nuclear changes to produce a metaphase II (MII) oocyte does not reflect the molecular and structural maturity of an oocyte, which is sometimes termed cytoplasmic maturation. The Golgi apparatus phosphorylates, fragments, and changes the localization during oocyte maturation. GM130 and phospho-GM130 are used as markers for the Golgi apparatus and phosphorylated Golgi apparatus, respectively. The goal of this study was to analyze the dynamics of the Golgi apparatus and its association with microtubules in bovine oocytes at different stages of in vitro maturation [IVM; i.e. germinal vesicle (GV), germinal vesicle breakdown (GVBD), metaphase I (MI), and MII]. The roles of CDC2A kinase (also known as p34cdc2) and cytoplasmic-dynein ATPase on Golgi dynamics were studied by using specific inhibitors. The distribution of the markers was assessed by immunocytochemistry and laser confocal microscopy. To unravel the role of CDC2A and cytoplasmic dynein ATPase on the dynamics of the Golgi apparatus, the inhibitors roscovitine (ROS) and sodium-orthovanadate (SOV), respectively, were used. In the first experiment, the nuclear maturation rate was checked in the presence of the inhibitors at different times and for different incubation times to explore whether oocytes were able to reach the MII stage. At the GV and GVBD stages, the Golgi apparatus is observed as fragments named mini-Golgies and at the MI and MII stages as punctate foci throughout the cytoplasm. Our results showed 2 well-defined movements of the Golgi apparatus toward opposite directions, depending on the maturation stage. The first movement was observed between 5 and 9 h of IVM (i.e. the GVBD stage), when the Golgi apparatus relocalized from the ooplasm to the periphery. The second movement was observed between 9 and 15 h of IVM (i.e. the MI stage), when the Golgi apparatus moved from the cortex to throughout the cytoplasm and remained there up to the MII stage. The use of inhibitors on CDC2A and cytoplasmic-dynein ATPase at selected time points revealed that CDC2A played a crucial role on the distribution of this organelle during the first movement, whereas the final localization at the GVBD stage was dependent on cytoplasmic-dynein transport. The second movement of the Golgi apparatus was disturbed by the SOV treatment, but not by the use of ROS, suggesting a role of cytoplasmic-dynein-dependent transport during the distribution and organization of the punctate foci at the MI stage. The phosphorylation status of Golgi was not affected at the different incubation times with inhibitors, except in those oocytes incubated with ROS for 24 h, suggesting a role of CDC2A. In conclusion, we describe the involvement of CDC2A during the first movement of the Golgi apparatus and the importance of cytoplasmic-dynein ATPase activity in the first and second relocalization of Golgi during bovine oocyte maturation. DAAD.

250 CYTOPLASMIC DYNEIN INTERMEDIATE CHAIN AND DYNACTIN p150Glued EXHIBIT DISTINCT SPATIAL AND TEMPORAL MICROTUBULE ASSOCIATIONS DURING BOVINE IN VITRO MATURATION AND ARE AFFECTED BY FOLLICLE SIZE

2008 ◽  
Vol 20 (1) ◽  
pp. 204
Author(s):  
S. E. Racedo ◽  
M. C. Branzini ◽  
D. Salamone ◽  
V. Y. Rawe ◽  
H. Niemann
Keyword(s):  
Oocyte Maturation ◽  
In Vitro Maturation ◽  
Chromatin Condensation ◽  
Calf Serum ◽  
Germinal Vesicle ◽  
Cytoplasmic Dynein ◽  
Meiotic Spindle ◽  
Germinal Vesicle Breakdown ◽  
Intermediate Chain

Microtubule molecular motors are critically involved in transporting vesicles during interphase, in building and maintaining spindles during mitosis and meiosis, and also in the localization of various organelles. DYNC1I1 (cytoplasmic dynein 1 intermediate chain) and its cofactor DCTN1 (dynactin p150Glued) are crucial for oocyte maturation but their role during mammalian female meiosis is not yet known. The goal of this study was to analyze the dynamics of these proteins in oocytes collected from different-size follicles at different stages of in vitro maturation (IVM), i.e., germinal vesicle stage (GV), germinal vesicle breakdown (GVBD), metaphase I (MI), and metaphase II (MII), and their association with microtubules. Ovaries were collected at a local abattoir. Cumulus–oocyte complexes (COCs) were aspirated from follicles either <2 mm or 2–8 mm in size and matured in M199, supplemented with 1% fatty acid-free BSA, 10 UI pregnant mare serum gonadotropin (PMSG)/5 UI HCG, and 100 µm cysteamine, at 39�C and 5% CO2. Follicle sizes and time points for fixation were: GV-0 h; GVBD-8 h for oocytes <2 mm and 9 h for oocytes 2–8 mm; MI-15 h; MII-24 h (Racedo et al. 2007, pub. online: 10.1002/mrd.20770). The distribution of the proteins was assessed by immunocytochemistry and laser confocal microscopy. The attached cumulus cells and zona pellucida of oocytes were removed in TALP-HEPES medium containing 1 mg mL–1 hyaluronidase and 2 mg mL–1 pronase, respectively. The oocytes were then incubated in a fixation–permeabilization solution containing 2% formaldehyde and 0.1%Triton X-100 for 1 h. Samples were then blocked for 1 h in 10 mm PBS + 0.3% BSA + 1% fetal calf serum (ICC blocking solution). The primary antibody was applied over night at 4�C, followed by treatment with fluorochrome-conjugated secondary antibodies for 1 h at 37�C in the dark. After RNase treatment, oocytes were incubated with TOTO-3 (Invitrogen, Carlsbad, CA, USA) to visualize the DNA. The material was mounted in an anti-fade medium (Vectashield�, Vector Laboratories, Burlingame, CA, USA) and imaged with a Zeiss laser scanning microscope. Immediately after chromatin condensation (GVBD), dynactin was in close association with the DNA and interacting with the spindles in MI and MII oocytes recovered from large follicles. No clear association with the DNA was observed in GVBD oocytes obtained from small follicles; little dynactin was found in MI and MII spindles. Dynein localization did not differ from dynactin in GVs and was homogeneously distributed in the cytoplasm of both groups of follicles. Dynein was not associated with the DNA in the GVBD stage while at MI and MII it was associated with the meiotic spindle. The association of dynein with microtubules was weak at the MI stage in oocytes from small follicles. Results provide insight into the regulatory mechanisms of oocyte maturation and a possible relationship with oocyte competence.

scholarly journals Requirement of mosXe protein kinase for meiotic maturation of Xenopus oocytes induced by a cdc2 mutant lacking regulatory phosphorylation sites.

1992 ◽  
Vol 12 (7) ◽  
pp. 3192-3203 ◽  
Author(s):  
K M Pickham ◽  
A N Meyer ◽  
J Li ◽  
D J Donoghue
Keyword(s):  
Protein Kinase ◽  
Oocyte Maturation ◽  
Xenopus Oocytes ◽  
Germinal Vesicle ◽  
Cyclin B1 ◽  
Phosphorylation Sites ◽  
Germinal Vesicle Breakdown ◽  
Regulatory Phosphorylation

The p34cdc2 protein kinase is a component of maturation-promoting factor, the master regulator of the cell cycle in all eukaryotes. The activity of p34cdc2 is itself tightly regulated by phosphorylation and dephosphorylation. Predicted regulatory phosphorylation sites of Xenopus p34cdc2 were mutated in vitro, and in vitro-transcribed RNAs were injected into Xenopus oocytes. The cdc2 single mutants Thr-14----Ala and Tyr-15----Phe did not induce germinal vesicle breakdown (BVBD) upon microinjection into oocytes. In contrast, the cdc2 double mutant Ala-14/Phe-15 did induce GVBD. Both the Ala-14 and Ala-14/Phe-15p34cdc2 mutants were shown to coimmunoprecipitate cyclin B1 and to phosphorylate histone H1 in immune complex kinase assays. Microinjection of antisense oligonucleotides to c-mosXe was used to demonstrate the role of mos protein synthesis in the induction of GVBD by the Ala-14/Phe-15 cdc2 mutant. Thr-161 was also mutated. p34cdc2 single mutants Ala-161 and Glu-161 and triple mutants Ala-14/Phe-15/Ala-161 and Ala-14/Phe-15/Glu-161 failed to induce GVBD in oocytes and showed a decreased binding to cyclin B1 in coimmunoprecipitations. Each of the cdc2 mutants was also assayed by coinjection with cyclin B1 or c-mosXe RNA into oocytes. Several of the cdc2 mutants were found to affect the kinetics of cyclin B1 and/or mos-induced GVBD upon coinjection, although none affected the rate of progesterone-induced maturation. We demonstrate here the significance of Thr-14, Tyr-15, and Thr-161 of p34cdc2 in Xenopus oocyte maturation. In addition, these results suggest a regulatory role for mosXe in induction of oocyte maturation by the cdc2 mutant Ala-14/Phe-15.

scholarly journals Dynamic changes of the Golgi apparatus during bovine in vitro oocyte maturation

Reproduction ◽  
2012 ◽  
Vol 143 (4) ◽  
pp. 439-447 ◽  
Author(s):  
S E Racedo ◽  
V Y Rawe ◽  
H Niemann
Keyword(s):  
Golgi Apparatus ◽  
Germinal Vesicle ◽  
Male Gamete ◽  
Active Role ◽  
Cytoplasmic Dynein ◽  
Dynamic Changes ◽  
Kinesin Eg5 ◽  
Specific Inhibitors

For successful fertilization by the male gamete, oocyte cytoplasmic organelles such as the Golgi apparatus have to undergo specific changes: the entire process is known as cytoplasmic maturation. The goal of this study was to unravel the dynamics of the Golgi apparatus in bovine oocytes at critical stages ofin vitromaturation, i.e. germinal vesicle (GV), GV breakdown (GVBD), metaphase I (MI) and metaphase II, and to investigate the role of various molecules critically involved therein. The cytoplasmic distribution of proteins was assessed by immunocytochemistry and laser confocal microscopy. We applied specific inhibitors, including nocodazole to unravel the functional role of the microtubular elements; sodium orthovanadate, which primarily inhibits cytoplasmic dynein ATPase activity; monastrol which inhibits the kinesin EG5; and roscovitine to inhibit the kinase cyclin-dependent kinase 2A (CDC2A). Prior to GVBD, the Golgi apparatus was translocated from the centre of the cytoplasm to the cortical area in the periphery, where it underwent fragmentation. A second translocation was observed between GVBD and MI stages, when the Golgi apparatus was moved from the cortex to the centre of the cytoplasm. Incubation with the specific inhibitors revealed that microtubules played an active role in the final localization at GVBD, while CDC2A was essential for Golgi fragmentation at GVBD stage. This partitioning was a precondition for the second movement. In conclusion, for the first time we show basic mechanisms critically involved in the regulation of the dynamic changes of Golgi apparatus during meiosis of the bovine oocyte.

scholarly journals 316EFFECTS OF ESTRADIOL-17² AND PROGESTERONE SUPPLEMENT ON THE RESUMPTION OF MEIOSIS OF CANINE OOCYTES MATURED IN VITRO

2004 ◽  
Vol 16 (2) ◽  
pp. 277
Author(s):  
M.K. Kim ◽  
Y.H. Fibrianto ◽  
H.J. Oh ◽  
G. Jang ◽  
K.S. Lee ◽  
...  
Keyword(s):  
Oocyte Maturation ◽  
In Vitro Maturation ◽  
Culture Medium ◽  
Linear Models ◽  
Uv Light ◽  
Germinal Vesicle ◽  
Domestic Animal ◽  
Estradiol 17Β

In the bitch, oocytes are ovulated at the germinal vesicle (GV) stage and mature in the isthmus of the oviduct around 3 days after ovulation, it is not known what elements trigger the release of this meiotic arrest. Canine IVM has shown limited success with maturation rates, usually around 20% (MII) (Farstad W, 2000 Anim. Reprod. Sci. 60–61, 375–387). Estrogen and progesterone are suggested to play a significant role in causing oocyte resumption of meiosis and progression to MII stage. The purpose of this study was to investigate the role of estradiol-17β (E2) and progesterone (P4) during in vitro maturation of canine oocytes in serum-free tissue culture medium (TCM)-199. Canine oocytes collected from bitches were categorized into three groups based on estrous stages, follicular, luteal, or anestrus, at routine ovariohystrectomy. Oocytes were cultured in vitro in TCM-199 supplemented with E2, P4 or E2+P4 according to experimental design at 39°C in 5% CO2 and O2. After 72h of maturation culture, oocytes were denuded, fixed in a 3.7% paraformaldehyde solution for 10min, stained with Hoechst 33342 in glycerol, and observed under the UV light. Three groups of oocytes were cultured in TCM-199 supplemented with different concentrations (0, 0.1, 1.0 or 2.0μgmL−1) of E2 (Experiment 1, n=898, replications: 5) or P4 (0, 0.5, 1.0 or 2.0μgmL−1, Experiment 2, n=734, replications: 5). Multiple comparisons were implemented using Generalized Linear Models in the SAS 8.12 program. The rates of oocyte maturation to MII stage were higher (P&lt;0.05) in follicular stage oocytes cultured with 2μgmL−1 E2 (17.9%) compared to other supplement groups (0 to 7.6%). No differences (P&lt;0.05) in rate of MII stage oocytes among P4 supplement groups were observed. In Experiment 3, to investigate the combined effects of E2 and P4 on in vitro maturation, three groups of oocytes were cultured in TCM-199 supplemented with 2μgmL−1 E2 and various concentration of P4 (0, 0.5, 1.0 or 2.0μgmL−1, Experiment 3, n=1613, replications: 5). The rate of oocyte maturation to MII stage (11.5%) was higher (P&lt;0.05) in follicular stage oocytes cultured with 2μgmL−1 E2+2.0μgmL−1 P4 supplement compared to other supplement groups (0 to 6.4%). In conclusion, the present study demonstrated that E2 supplement in the culture medium increased maturation of canine oocyte to MII stage and that supplement of P4 alone did not promote oocyte maturation. However, P4 supplemented with E2 further promoted oocyte maturation in the follicular stage compared to E2 supplement alone, indicating that P4 acts synergistically with E2 on canine oocyte maturation in the presence of E2. From our results, we conclude that canine oocytes are exposed to high levels of P4 during maturation due to the preovulatory luteinization of canine follicles which gives rise to high intrafollicular as well as intratubal P4 concentrations-this is very different from the situation in oocytes from other domestic animal species. This study was supported by Biogreen 21-1000520030100000.

Behavior of γ-tubulin during spindle formation in Xenopus oocytes: requirement of cytoplasmic dynein-dependent translocation

Zygote ◽  
2005 ◽  
Vol 13 (3) ◽  
pp. 219-226 ◽  
Author(s):  
Tomoya Kotani ◽  
Masakane Yamashita
Keyword(s):  
Oocyte Maturation ◽  
Late Stage ◽  
Xenopus Oocytes ◽  
Germinal Vesicle ◽  
Cytoplasmic Dynein ◽  
Microtubule Organization ◽  
Germinal Vesicle Breakdown ◽  
Spindle Formation ◽  
Bipolar Spindle

Vertebrate oocytes do not contain centrosomes and therefore form an acentrosomal spindle during oocyte maturation. γ-Tubulin is known to be essential for nucleation of microtubules at centrosomes, but little is known about the behaviour and role of γ-tubulin during spindle formation in oocytes. We first observed sequential localization of γ-tubulin during spindle formation in Xenopus oocytes. γ-Tubulin assembled in the basal regions of the germinal vesicle (GV) at the onset of germinal vesicle breakdown (GVBD) and remained on the microtubule-organizing centre (MTOC) until a complex of the MTOC and transient-microtubule array (TMA) reached the oocyte surface. Prior to bipolar spindle formation, oocytes formed an aggregation of microtubules and γ-tubulin was concentrated at the centre of the aggregation. At the late stage of bipolar spindle formation, γ-tubulin accumulated at each pole. Anti-dynein antibody disrupted the localization of γ-tubulin, indicating that the translocation described above is dependent on dynein activity. We finally revealed that XMAP215, a microtubule-associated protein cooperating with γ-tubulin for the assembly of microtubules, but not γ-tubulin, was phosphorylated during oocyte maturation. These results suggest that γ-tubulin is translocated by dynein to regulate microtubule organization leading to spindle formation and that modification of the molecules that cooperate with γ-tubulin, but not γ-tubulin itself, is important for microtubule reorganization.

Requirement of mosXe protein kinase for meiotic maturation of Xenopus oocytes induced by a cdc2 mutant lacking regulatory phosphorylation sites

1992 ◽  
Vol 12 (7) ◽  
pp. 3192-3203
Author(s):  
K M Pickham ◽  
A N Meyer ◽  
J Li ◽  
D J Donoghue
Keyword(s):  
Protein Kinase ◽  
Oocyte Maturation ◽  
Xenopus Oocytes ◽  
Germinal Vesicle ◽  
Cyclin B1 ◽  
Phosphorylation Sites ◽  
Germinal Vesicle Breakdown ◽  
Regulatory Phosphorylation

The p34cdc2 protein kinase is a component of maturation-promoting factor, the master regulator of the cell cycle in all eukaryotes. The activity of p34cdc2 is itself tightly regulated by phosphorylation and dephosphorylation. Predicted regulatory phosphorylation sites of Xenopus p34cdc2 were mutated in vitro, and in vitro-transcribed RNAs were injected into Xenopus oocytes. The cdc2 single mutants Thr-14----Ala and Tyr-15----Phe did not induce germinal vesicle breakdown (BVBD) upon microinjection into oocytes. In contrast, the cdc2 double mutant Ala-14/Phe-15 did induce GVBD. Both the Ala-14 and Ala-14/Phe-15p34cdc2 mutants were shown to coimmunoprecipitate cyclin B1 and to phosphorylate histone H1 in immune complex kinase assays. Microinjection of antisense oligonucleotides to c-mosXe was used to demonstrate the role of mos protein synthesis in the induction of GVBD by the Ala-14/Phe-15 cdc2 mutant. Thr-161 was also mutated. p34cdc2 single mutants Ala-161 and Glu-161 and triple mutants Ala-14/Phe-15/Ala-161 and Ala-14/Phe-15/Glu-161 failed to induce GVBD in oocytes and showed a decreased binding to cyclin B1 in coimmunoprecipitations. Each of the cdc2 mutants was also assayed by coinjection with cyclin B1 or c-mosXe RNA into oocytes. Several of the cdc2 mutants were found to affect the kinetics of cyclin B1 and/or mos-induced GVBD upon coinjection, although none affected the rate of progesterone-induced maturation. We demonstrate here the significance of Thr-14, Tyr-15, and Thr-161 of p34cdc2 in Xenopus oocyte maturation. In addition, these results suggest a regulatory role for mosXe in induction of oocyte maturation by the cdc2 mutant Ala-14/Phe-15.

scholarly journals Nucleus, Cytoskeleton, and Mitogen-Activated Protein Kinase p38 Dynamics during In Vitro Maturation of Porcine Oocytes

Animals ◽  
2019 ◽  
Vol 9 (4) ◽  
pp. 163
Author(s):  
Payungsuk Intawicha ◽  
Li-Kuang Tsai ◽  
Shih-Ying Yen ◽  
Neng-Wen Lo ◽  
Jyh-Cherng Ju
Keyword(s):  
Oocyte Maturation ◽  
Mammalian Cells ◽  
In Vitro Maturation ◽  
Germinal Vesicle ◽  
Mitogen Activated Protein Kinase ◽  
Meiotic Maturation ◽  
Germinal Vesicle Breakdown ◽  
Mitogen Activated Protein ◽  
Porcine Oocyte

The mitogen-activated kinase (MAPK) p38, a member of the MAPK subfamily, is conserved in all mammalian cells and plays important roles in response to various physiologic cues, including mitogens and heat shock. In the present study, MAPK p38 protein expression in porcine oocytes was analyzed during in vitro maturation (IVM) by Western blotting and immunocytochemistry. The levels of p-p38 or activated p38 and p38 expression were at the lowest in the germinal vesicle (GV) stage oocyte, gradually rising at the germinal vesicle breakdown (GVBD) and then reaching a plateau throughout the IVM culture (p < 0.05). Similarly, the expression level of total p38 was also lower in the GV oocyte than in the oocyte of other meiotic stages and uprising after GVBD and remained high until the metaphase III (MII) stage (p < 0.05). In the GV stage, phosphorylated p38 (p-p38) was initially detectable in the ooplasm and subsequently became clear around the nucleus and localized in the ooplasm at GVBD (18 h post-culture). During the metaphase I (MI) and metaphase II (MII) stages, p-p38 was evenly distributed throughout the ooplasm after IVM for 30 or 42 h. We found that the subcellular localization increased in p-p38 expression throughout oocyte maturation (p < 0.05) and that dynamic reorganization of the cytoskeleton, including microfilaments and microtubules, was progressively changed during the course of meiotic maturation which was likely to be associated with the activation or networking of p38 with other proteins in supporting oocyte development. In conclusion, the alteration of p38 activation is essential for the regulation of porcine oocyte maturation, accompanied by the progressive reorganization and redistribution of the cytoskeleton and MAPK p38, respectively, in the ooplasm.

scholarly journals 290 MODULATION OF ARYLHYDROCARBON RECEPTOR ACTIVITY DURING IN VITRO MATURATION OF BOVINE OOCYTES

2005 ◽  
Vol 17 (2) ◽  
pp. 295
Author(s):  
D. Nestler ◽  
M. Risch ◽  
B. Fischer ◽  
P. Pocar
Keyword(s):  
Oocyte Maturation ◽  
Target Genes ◽  
Critical Role ◽  
Germinal Vesicle ◽  
Protein Synthesis Inhibitor ◽  
Bovine Oocyte ◽  
Meiotic Arrest ◽  
Germinal Vesicle Breakdown ◽  
Arylhydrocarbon Receptor

The arylhydrocarbon receptor (AhR), a ligand activated transcription factor, has been extensively characterized from a toxicological point of view due to its ability to mediate the adverse effects of a variety of halogenated aromatic hydrocarbons. Recent reports on AhR knockout mice suggest that the AhR may play a role in ovarian physiology. We have previously demonstrated that AhR activity (as indicated by the up-regulation of the target gene cytochrome p450 1A1: CYP1A1) is stimulated during bovine oocyte IVM in the absence of exogenous ligands. Furthermore, exposure to specific AhR antagonists, besides down-regulating the expression of CYP1A1, significantly impairs the ability of the oocyte to complete maturation until the metaphase II stage (Pocar et al. 2004 Endocrinology 145, 1594–1601). The aim of the present study was to further investigate the mechanisms underlying the AhR activation during IVM. Several reports point to a critical role of phosphorylation in the regulation of the AhR-complex. Furthermore, the mitogen-activated protein kinase (MAPK, extracellular regulated kinase (ERK 1 and 2)) cascade has been shown to play a crucial role in regulating meiotic cell cycles during bovine oocyte maturation. A total of 572 bovine cumulus-oocyte complexes were used to investigate the potential role of the MAPK in modulating the activity of the AhR during IVM. The effect of the broad-spectrum serine/threonine kinase inhibitor, 6-dimethylaminopurine (6-DMAP), on the induction of CYP1A1 during oocyte maturation was investigated. As expected, exposure to 6-DMAP induced meiotic arrest (at the stage of germinal vesicle/germinal vesicle breakdown) and down-regulated the expression level of phosphorylated ERK 1 and 2. Interestingly, a significant down-regulation of the target genes CYP1A1 and CYP1B1 (9.5% and 26.8% of control, respectively) and an up-regulation of the AhR (199.4% of control) were observed at the mRNA level. This phenomenon was partially reversible after a period of further 24 h of culture in the absence of 6-DMAP. In this condition, besides a recovery of oocyte maturation and phosphorylation status of ERK 1 and 2 to levels comparable to control, a significant up-regulation of CYP1A1 mRNA was observed (68.3% of control). Finally, to confirm the role of serine/threonine kinases in modulating the activity of AhR during resumption of meiosis, we exposed the oocytes to cycloheximide, a protein synthesis inhibitor, also known to arrest oocyte maturation. Furthermore, although cycloheximide exposure induced meiotic arrest, no significant differences in the expression levels of AhR or its target genes compared to control were observed. Each experiment was replicated at least three times. Data were assessed using ANOVA followed by Duncans multiple rance test. The criterion for significance was set at P < 0.05. In conclusion, our results strongly suggest that 6-DMAP-sensitive kinase(s) is (are) involved in the regulation of AhR during bovine oocyte maturation. Further analyses are necessary to understand the biological significance of these observations.

Role of prolactin in nuclear maturation and ovulation in amphibian oocytes

Zygote ◽  
2005 ◽  
Vol 13 (3) ◽  
pp. 265-268
Author(s):  
Inés Ramos ◽  
Susana Cisint ◽  
Marcela F. Medina ◽  
Claudia A. Crespo ◽  
Silvia N. Fernández
Keyword(s):  
Oocyte Maturation ◽  
Incubation Medium ◽  
Germinal Vesicle ◽  
Sexual Cycle ◽  
Breeding Period ◽  
Germinal Vesicle Breakdown ◽  
Nuclear Maturation ◽  
Bufo Arenarum

The present study was undertaken to determine the effect of prolactin (Prl) on Bufo arenarum oocyte maturation and ovulation, two characteristic events of the breeding period, the stage of the sexual cycle in which gamete growth is complete. We observed that Prl, at the doses assayed, did not affect nuclear maturation per se. In addition, when follicles were pretreated with Prl and progesterone was later added to the medium as a physiological nuclear maturation inducer, the percentage of germinal vesicle breakdown obtained with the steroid was unaffected by Prl. The analysis of the in vitro ovulation process demonstrated that pituitary homologous homogenate (PHH) produced a dose- and month-dependent stimulating effect. The maximum percentage of ovulated oocytes was obtained from the end of July to October, the period in which oviposition naturally occurs. Prl by itself did not affect the ovulation process, but when both the hormone and PHH were present in the incubation medium, a significant increase in ovulated oocytes was observed. The results suggest that Prl does not participate in oocyte maturation; however, its presence in the incubation medium would increase oocyte sensitivity to the action of the physiological ovulation inducers.

Antioxidants Stimulate Germinal Vesicle Breakdown, But Inhibit Chromosome Formation And Spindle Assembly In Porcine Oocytes

2000 ◽  
Vol 6 (S2) ◽  
pp. 964-965
Author(s):  
Qing-Yuan Sun ◽  
Randall S. Prather ◽  
Heide Schatten
Keyword(s):  
Oocyte Maturation ◽  
Germinal Vesicle ◽  
Spindle Assembly ◽  
Mouse Oocyte ◽  
Germinal Vesicle Breakdown ◽  
Oocyte Meiosis ◽  
First Polar Body ◽  
Porcine Oocyte ◽  
Chromosome Formation

Mammalian oocytes are arrested at the diplotene stage of the first meiotic division. Release of oocytes from their follicles induces meiotic resumption characterized by germinal vesicle breakdown (GVBD), followed by the chromosome formation and metaphase I spindle organization and finally the extrusion the first polar body. Recently it was shown that cellpermeant antioxidants significantly inhibit spontaneous resumption of meiosis in mouse oocytes, which may indicate a role of oxygen radicals in oocyte maturation. The regulation of mouse oocyte meiosis resumption is different from that of large domestic animals in that GVBD is independent of Ca2+ and protein synthesis. The present study investigated the influence of two cell-permeant antioxidants, 2(3)-ter-butyl-4-hydroxyanisole (BHA) and nordihydroguaiaretic acid (NDGA), on porcine oocyte meiosis resumption, chromatin behavior and spindle assembly. Our findings revealed a different role of antioxidants in porcine oocyte meiosis resumption than in mouse oocyte maturation.

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