Conditions to optimise the developmental competence of immature equine oocytes

2020 ◽  
Vol 32 (11) ◽  
pp. 1012
Author(s):  
Elizabeth S. Metcalf ◽  
Keith R. Masterson ◽  
David Battaglia ◽  
Jeremy G. Thompson ◽  
Robert Foss ◽  
...  
Keyword(s):  
Adenylate Cyclase ◽  
Culture Media ◽  
Phosphodiesterase Inhibitor ◽  
Developmental Competence ◽  
Sodium Ascorbate ◽  
Blastocyst Formation ◽  
Developmental Potential ◽  
Holding Temperature ◽  
Cyclase Activator ◽  
Adenylate Cyclase Activator

Optimising the developmental potential of immature equine oocytes and invitro-produced (IVP) embryos was explored through modifications of established media and holding temperature. In Experiment 1, delaying spontaneous resumption of meiosis through the process of simulated physiological oocyte maturation with the addition of the adenylate cyclase activator forskolin (50µM) and the phosphodiesterase inhibitor 3-isobutyl-1-methylxanthine (100µM) to overnight holding medium before maturation improved blastocyst production (P<0.05). In Experiment 2, the blastocyst production rate was increased significantly when cumulin (100ng mL−1) was added to the overnight holding or culture media (P<0.05). In Experiment 3, immature oocytes held overnight at 16°C before maturation had improved developmental competence than those held at 20°C and 5°C (P<0.05). There was no difference between maturation rates, but blastocyst formation per cleaved oocyte was significantly greater in oocytes held overnight at 16°C than at 20°C or 5°C. Furthermore, blastocyst formation per recovered oocyte and per fertilised oocyte was greater when oocytes were held before maturation at 16°C than at 5°C (P<0.05). In Experiment 4, the addition of sodium ascorbate (AC; 50µg mL−1) to the maturation and/or culture media of oocytes and IVP embryos did not improve blastocyst production, but did appear to lower cleavage rates compared with oocytes and embryos cultured without AC.

176. INHIBITION OF PORCINE OOCYTE NUCLEAR MATURATION IN VITRO USING A PHOSPHODIESTERASE INHIBITOR AND AN ADENYLATE CYCLASE ACTIVATOR

2010 ◽  
Vol 22 (9) ◽  
pp. 94
Author(s):  
M. Bertoldo ◽  
T. Sellens ◽  
C. G. Grupen
Keyword(s):  
Adenylate Cyclase ◽  
Phosphodiesterase Inhibitor ◽  
Phosphodiesterase Inhibitors ◽  
Extended Period ◽  
Nuclear Maturation ◽  
Porcine Oocyte ◽  
Defined Culture ◽  
Cyclase Activator ◽  
Adenylate Cyclase Activator

Asynchronous nuclear and cytoplasmic maturation is thought to contribute to poor embryo production in vitro. Nuclear arrest is mediated by cAMP and can be maintained within the oocyte using non-specific phosphodiesterase inhibitors (3-isobutyl-1-methylxanthine ; IBMX) and the adenylate cyclase activator forskolin (FSK) (1). The aim of this study was to investigate the effect of IBMX and FSK supplementation on porcine oocyte nuclear maturation during COC recovery and IVM using a defined culture system. In all experiments, cAMP modulators were added to Hepes-buffered media held in collection tubes. COCs recovered from 3–5 mm diameter follicles of prepubertal ovaries were cultured in basic maturation media in the absence of FSH. Nuclear maturation was assessed using orcein dye. In Experiment 1, IVM media was supplemented with 0, 50 or 500 µM IBMX. In Experiment 2, IVM media was supplemented with 0, 5, 10, 50 and 100µM FSK. In Experiment 3, IVM medium was supplemented with combinations of IBMX and FSK to give the treatments; control, 50IBMX/50FSK, 50IBMX/100FSK, 500IBMX/50FSK and 500IBMX/100FSK. Nuclear maturation was assessed at 0, 2, 4 and 18 h after the onset of IVM. At 18 h of culture, there were no differences in the proportion of oocytes supplemented with 0, 50 or 500 µM IBMX reaching MII. Incubation with 10, 50 or 100 µM FSK resulted in 8-16% of oocytes at MII at 18 h compared to the other groups (25–29%; P < 0.001). The combinations of IBMX and FSK resulted in greater proportions (86–98%) of oocytes remaining at the GV stage at 18 h compared to the control (16%; P < 0.001). There were no differences in the proportion of oocytes remaining at the GV stage at the earlier time points (P > 0.05). The results demonstrate that these cAMP modulators, in combination, are highly effective in maintaining porcine oocyte meiotic arrest in vitro for an extended period. (1) Albuz FK et al., Proceedings of the 25th Annual Meeting of ESHRE, Amsterdam, The Netherlands, 2009.

Increase in protein and tubulin mRNA synthesis in frog sensory neurons treated with the adenylate cyclase activator, forskolin

1990 ◽  
Vol 1 (3,4) ◽  
pp. 225-232
Author(s):  
Richard C. Carlsen ◽  
Marino De Leon ◽  
Wolfram Tetzlaff ◽  
Irma M. Parhad ◽  
Mark A. Bisby
Keyword(s):  
Adenylate Cyclase ◽  
Sensory Neurons ◽  
Mrna Synthesis ◽  
Tubulin Mrna ◽  
Cyclase Activator ◽  
Adenylate Cyclase Activator

scholarly journals MicroRNAs from Extracellular Vesicles Secreted by Bovine Embryos as Early Biomarkers of Developmental Competence

2020 ◽  
Vol 21 (23) ◽  
pp. 8888
Author(s):  
Bárbara Melo-Baez ◽  
Yat S. Wong ◽  
Constanza J. Aguilera ◽  
Joel Cabezas ◽  
Ana C. F. Mançanares ◽  
...  
Keyword(s):  
Extracellular Vesicles ◽  
Fatty Acid Biosynthesis ◽  
Culture Media ◽  
Blastocyst Stage ◽  
Developmental Competence ◽  
Target Cells ◽  
Bovine Embryos ◽  
Developmental Potential ◽  
Maternal Communication

During early development, embryos secrete extracellular vesicles (EVs) that participate in embryo–maternal communication. Among other molecules, EVs carry microRNAs (miRNAs) that interfere with gene expression in target cells; miRNAs participate in embryo–maternal communication. Embryo selection based on secreted miRNAs may have an impact on bovine breeding programs. This research aimed to evaluate the size, concentration, and miRNA content of EVs secreted by bovine embryos with different developmental potential, during the compaction period (days 3.5–5). Individual culture media from in vitro–produced embryos were collected at day 5, while embryos were further cultured and classified at day 7, as G1 (conditioned-culture media by embryos arrested in the 8–16-cells stage) and G2 (conditioned-culture media by embryos that reached blastocyst stages at day 7). Collected nanoparticles from embryo conditioned culture media were cataloged as EVs by their morphology and the presence of classical molecular markers. Size and concentration of EVs from G1 were higher than EVs secreted by G2. We identified 95 miRNAs; bta-miR-103, bta-miR-502a, bta-miR-100, and bta-miR-1 were upregulated in G1, whereas bta-miR-92a, bta-miR-140, bta-miR-2285a, and bta-miR-222 were downregulated. The most significant upregulated pathways were fatty acid biosynthesis and metabolism, lysine degradation, gap junction, and signaling pathways regulating pluripotency of stem cells. The characteristics of EVs secreted by bovine embryos during the compaction period vary according to embryo competence. Embryos that reach the blastocyst stage secrete fewer and smaller vesicles. Furthermore, the loading of specific miRNAs into the EVs depends on embryo developmental competence.

Immunomodulation by an Adenylate Cyclase Activator, NKH477,in Vivoandin Vitro

1996 ◽  
Vol 79 (1) ◽  
pp. 25-35 ◽  
Author(s):  
Yutaka Furukawa ◽  
Akira Matsumori ◽  
Toshiro Hirozane ◽  
Shigeo Matsui ◽  
Yukihito Sato ◽  
...  
Keyword(s):  
Adenylate Cyclase ◽  
Cyclase Activator ◽  
Adenylate Cyclase Activator

β-Adrenergic induced K + current in Xenopus oocytes: role of cAMP, inhibition by muscarinic agents

1985 ◽  
Vol 223 (1232) ◽  
pp. 389-402 ◽  
Keyword(s):  
Adenylate Cyclase ◽  
Adrenergic Receptors ◽  
Xenopus Oocytes ◽  
Follicular Cells ◽  
K Current ◽  
Β Adrenergic Receptors ◽  
Cyclase Activator ◽  
Adenylate Cyclase Activator ◽  
Muscarinic Agents

The K + current induced by isoprenaline acting on (β-adrenergic receptors in Xenopus laevis has been studied in oocytes still surrounded by their follicular cells and inner ovarian epithelium. Forskolin, an adenylate cyclase activator, induced a similar K + current and when used at subliminal concentration it potentiated the current induced by isoprenaline. Inhibition of phosphodiesterase by methylisobutylxanthine also en­hanced the response to isoprenaline. 8-Br-cAMP, a permeant analogue of cAMP also produced a K + current. Acetylcholine produced a long lasting inhibition of the isoprenaline current. This inhibition was not seen in the presence of atropine. It is concluded that the K + current induced by the activation of β-adrenergic receptors in the oocyte is mediated by an intracellular rise of cAMP.

scholarly journals cAMP Modulators before In Vitro Maturation Decrease DNA Damage and Boost Developmental Potential of Sheep Oocytes

Animals ◽  
2021 ◽  
Vol 11 (9) ◽  
pp. 2512
Author(s):  
Daniela-Alejandra Medina-Chávez ◽  
Irene Sánchez-Ajofrín ◽  
Patricia Peris-Frau ◽  
Carolina Maside ◽  
Vidal Montoro ◽  
...  
Keyword(s):  
Dna Damage ◽  
In Vitro Maturation ◽  
Phosphodiesterase Inhibitor ◽  
Cumulus Cells ◽  
Culture Conditions ◽  
Developmental Competence ◽  
Developmental Potential ◽  
Assisted Reproduction Technologies ◽  
Underlying Mechanisms

To date, the underlying mechanisms by which cAMP modulators act during in vitro maturation to improve oocyte developmental competence are poorly understood. Here, we sought to fill this knowledge gap by evaluating the use of phosphodiesterase inhibitor 3-isobutyl-1-methylxanthine (IBMX) and adenylyl cyclase activator forskolin during a culture period of 2 h before in vitro maturation (pre-IVM) on the nuclear and cytoplasmic maturation features in essential organelles, cumulus cells activity, and in vitro developmental potential of sheep oocytes. Results showed that pre-IVM treatment significantly decreased (p < 0.05) the DNA damage of mature oocytes (pre-IVM = 2.08% ± 3.51% vs. control = 20.58% ± 3.51%) and increased (p ≤ 0.05) expanded blastocyst rates compared to the control (from the total of oocytes: pre-IVM = 23.89% ± 1.47% vs. control = 18.22% ± 1.47%, and from the cleaved embryos: pre-IVM = 45.16% ± 1.73% vs. control = 32.88% ± 1.73%). Considering that oocytes are highly vulnerable to the accumulation of DNA damage because of exposure to in vitro culture conditions, our results suggest that the modulation of intra-oocyte cAMP levels with forskolin and IBMX before IVM might afford oocytes a more effective DNA repair mechanism to overcome damage obstacles and ultimately improve developmental competence. This previously unappreciated action of cAMP modulators could help to develop improved methods for assisted reproduction technologies in animal and clinical research.

210 CLEAVAGE ASSESSMENT AT DAY 2 TO PREDICT BLASTOCYST DEVELOPMENTAL POTENTIAL IN PORCINE IN VITRO-FERTILIZED EMBRYOS

2009 ◽  
Vol 21 (1) ◽  
pp. 203
Author(s):  
Y. J. Kim ◽  
Y. P. Jeon ◽  
S. H. Hyun
Keyword(s):  
Cumulus Cells ◽  
Preimplantation Development ◽  
Developmental Competence ◽  
Preimplantation Embryos ◽  
Embryo Morphology ◽  
Blastocyst Formation ◽  
Developmental Potential ◽  
Vitro Fertilization ◽  
The Relationship

Porcine embryos could be a valuable tool to study preimplantation development, implantation, and pregnancy, but to do this it is necessary to establish an efficient in vitro embryo production system. Because the cause of high mortality in embryos during preimplantation development is not clear, a noninvasive method of determining the developmental potential of cleavage-stage embryos is needed. The objective was to evaluate the developmental potential of Day 2 embryos in a porcine in vitro fertilization (IVF) system. Specifically, this study was conducted to examine the relationship between embryo morphology 48 h after IVF on rates of blastocyst formation 5 days later. To prepare in vitro maturation (IVM) of porcine oocytes, cumulus–oocyte complexes were obtained from slaughterhouse-derived ovaries and matured in M-199 medium supplemented with 10% pig follicular fluid and 0.57 mm cysteine for 44 h and then freed from cumulus cells. After IVM, cumulus-free oocytes were coincubated with frozen–thawed sperm (2 × 106 cells mL–1) and 2 mm caffeine for 6 h. Inseminated embryos were cultured in NCSU-23 medium that was supplemented with 0.5 mm pyruvate and 0.5 mm lactate. Data were analyzed by ANOVA and Duncan’s test (P < 0.05). Morphology data on a total of 919 embryos were analyzed retrospectively. Forty-eight hours after insemination, embryos were classified into the following 5 groups based on the cleavage state: 1 cell, 2 cells, 4 cells, 5 to 8 cells, and fragmentation. These groups were cultured another 120 h and then evaluated for blastocyst formation. Blastocyst formation rates were significantly higher in the 4-cell (38.07%) and 5- to 8-cell (40.65%) cleaving groups than in the other groups (P < 0.05). In contrast, the 2-cell and fragmentation groups produced 7.5 and 2.9% blastocysts, respectively. Data suggest that embryos reaching 4 cells and 5 to 8 cells by 48 h after insemination have high developmental competence, and this parameter may be useful to predict the development of preimplantation embryos and their ability to establish pregnancy. This work was supported by a grant (No. 20070301034040) from the BioGreen 21 program, Rural Development Administration, Republic of Korea.

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