Intracellular pH increase accompanies parthenogenetic activation of porcine, bovine and murine oocytes

2000 ◽  
Vol 12 (4) ◽  
pp. 201 ◽  
Author(s):  
Nancy T. Ruddock ◽  
Zoltán Macháty ◽  
Randall S. Prather
Keyword(s):  
Intracellular Ph ◽  
Prolonged Exposure ◽  
Calcium Ionophore ◽  
Calcium Ionophore A23187 ◽  
Ionophore A23187 ◽  
Oocyte Activation ◽  
Parthenogenetic Activation ◽  
Sea Urchin Egg ◽  
Porcine Oocyte ◽  
Bovine Oocytes

Although an intracellular pH (pH i ) increase at the time of fertilization is necessary for activation of the sea urchin egg, recent reports in the mouse and rat have indicated that there is not a pHi increase during fertilization or during 7% ethanol activation in the mouse. It has been suggested that mammals may have lost the need for a pH i increase at the time of fertilization and the present study reports significant pH i changes during parthenogenetic activation of porcine IVM oocytes, as well as pH i responses to activation in bovine and murine oocytes. Transient intracellular pH changes were found during porcine oocyte activation when using 7% ethanol and with 50 or 100 M calcium ionophore (A23187). Treatment with 200 M thimerosal resulted in an increase in pH i after a delay of approximately 12 min. Murine oocytes showed a significant increase during activation with 7% ethanol and A23187 as well as during prolonged exposure to thimerosal. Bovine oocytes exhibited an increase in pH i only when activated with 50 or 100 M A23187. The final set of experiments aimed to determine whether the porcine oocyte has mechanisms to alleviate induced acidic and alkaline challenges. Both acidic (~20 mM acetic acid) and alkaline (~30 mM ammonium chloride) challenges caused significant changes in pH i that porcine IVM oocytes were capable of recovering from within 35 min. Future studies will focus on determining which of the mechanisms is producing the pH i increase at the time of parthenogenetic activation in the porcine oocyte.

Protein synthesis is not required for male pronuclear formation in bovine zygotes

Zygote ◽  
1996 ◽  
Vol 4 (1) ◽  
pp. 41-48 ◽  
Author(s):  
R.C. Chian ◽  
M.A. Sirard
Keyword(s):  
Protein Synthesis ◽  
Calcium Ionophore ◽  
Sperm Nucleus ◽  
Calcium Ionophore A23187 ◽  
Protein Synthesis Inhibitor ◽  
Ionophore A23187 ◽  
Oocyte Activation ◽  
Synthesis Inhibitor ◽  
Parthenogenetic Activation ◽  
Pronuclear Formation

SummaryFollowing fertilisation, the sperm triggers a series of intracellular changes which initiate oocyte activation and pronuclear formation. Oocyte activation can also be induced artificially by several chemicals, such as the calcium ionophore A23187. The sperm nucleus is transformed into the male pronucleus through the interaction of oocyte cytoplasmic factors. The profile of protein synthesis is different in bovine oocytes following fertilisation and parthenogenetic activation. The formation of male and female pronuclei was not blocked by the presence of the protein synthesis inhibitor cycloheximide. These results suggest that bovine oocyte activation by sperm and parthenogenetic activation induce different cytoplasmic responses for protein synthesis and that new protein synthesis is not required for male pronuclear formation in bovine zygotes.

Effect of centrifugation on early embryonic development and parthenogenetic activation of bovine oocytes matured in vitro

2001 ◽  
Vol 13 (6) ◽  
pp. 383 ◽  
Author(s):  
Jin-Tae Chung ◽  
Bruce R. Downey ◽  
Robert F. Casper ◽  
Ri-Cheng Chian
Keyword(s):  
Embryonic Development ◽  
Calcium Ionophore ◽  
Calcium Ionophore A23187 ◽  
Early Embryonic Development ◽  
Ionophore A23187 ◽  
Vitro Fertilization ◽  
Parthenogenetic Activation ◽  
Treatment Groups ◽  
Bovine Oocytes

This study examined the fertilization, early developmental competence and capacity for parthenogenetic activation of bovine oocytes matured in vitro after centrifugation. Immature oocytes were cultured in tissue culture medium 199 supplemented with 10% fetal bovine serum and 75 mIU mL–1 FSH + LH at 5% CO2 to facilitate maturation. After culture for 24 or 30 h, the metaphase-II stage oocytes were centrifuged at 3000, 5000, 7000 or 10000g for 5 min before in vitro fertilization or parthenogenetic activation. Frozen–thawed bull semen was used for in vitro fertilization. For parthenogenetic activation, the oocytes were exposed to 20 M calcium ionophore A23187 for 5 min at room temperature. Fertilization rates were not different between control and treatment groups (87.7% v. 74.6%, 73.4%, 75.9% and 76.4% respectively). Also, there were no differences in early embryonic development between control and treatment groups (rates of blastocyst formation were 21.1% v. 20.2%, 28.8%, 31.2% and 24.1% respectively). When the oocytes were centrifuged at various speeds alone, the activation rate of oocytes was significantly higher (P<0.05) in the 10 000g treatment group compared with control (10.8% v. 0.0%). There were no differences in the activation rates of oocytes between control and treatment groups at speeds up to 7000g (70.9% v. 71.9%, 78.3% and 77.2% respectively) after centrifugation and stimulation with Ca2+-ionophore. However, the activation rate of oocytes was significantly higher (P<0.05) in the 10 000g treatment group compared with control (70.9% v. 83.1%). In addition, the percentage of activated oocytes with diploid formation was significantly higher in the oocytes after centrifugation at 10 000g and stimulation with calcium ionophore A23187 than in the control (18.4% v. 7.1%). These results indicate that centrifugation of oocytes matured in vitro has no detrimental effect on fertilization and subsequent early embryonic development. They also indicate that the oocytes might be parthenogenetically activated after centrifugation and that high-speed centrifugation may induce activation of some oocytes. The results suggest that the optimal speed for centrifugation of bovine oocytes might be ≤7000g to enhance the visibility of nuclear elements for further micromanipulation.

Cell cycle progression of parthenogenetically activated mouse oocytes to interphase is dependent on the level of internal calcium

1992 ◽  
Vol 103 (2) ◽  
pp. 389-396 ◽  
Author(s):  
C. Vincent ◽  
T.R. Cheek ◽  
M.H. Johnson
Keyword(s):  
Cell Cycle Progression ◽  
Polar Body ◽  
Calcium Ionophore ◽  
Cytosolic Calcium ◽  
Calcium Ionophore A23187 ◽  
Mouse Oocyte ◽  
Free Calcium ◽  
Ionophore A23187 ◽  
Parthenogenetic Activation ◽  
Mouse Oocytes

Nuclear maturation of the mouse oocyte becomes arrested in metaphase of the second meiotic division (MII). Fertilization or parthenogenetic activation induces meiotic completion, chromosomal decondensation and formation of a pronucleus. This completion of meiosis is probably triggered by a transient increase in cytosolic calcium ions. When activated just after ovulation by a low concentration of the calcium ionophore A23187, the majority of the mouse oocytes go through a metaphase to anaphase transition and extrude their second polar body but they do not proceed into interphase; instead their chromatids remain condensed and a microtubular metaphase spindle reforms (metaphase III). However, a high percentage of these oocytes will undergo a true parthenogenetic activation assessed by the formation of a pronucleus, when exposed to a higher concentration of the calcium ionophore. The capacity of the mouse oocyte to pass into metaphase III is lost with increasing time post-ovulation. Direct measurement of intracellular calcium with Fura-2 reveals higher levels of cytosolic calcium in aged oocytes and/or using higher concentrations of calcium ionophore for activation. It is concluded that the internal free calcium level determines the transition to interphase.

A combination of calcium ionophore and puromycin effectively produces human parthenogenones with one haploid pronucleus

Zygote ◽  
2001 ◽  
Vol 9 (1) ◽  
pp. 83-88 ◽  
Author(s):  
Koji Nakagawa ◽  
Shuji Yamano ◽  
Hisayo Nakasaka ◽  
Kenji Hinokio ◽  
Midori Yoshizawa ◽  
...  
Keyword(s):  
Polar Body ◽  
Calcium Ionophore ◽  
Calcium Ionophore A23187 ◽  
Control Group ◽  
Ionophore A23187 ◽  
Cleavage Rate ◽  
Parthenogenetic Activation ◽  
Second Polar Body ◽  
Activation Rates ◽  
Pronuclear Formation

Parthenogenetic activation with various combinations of the calcium ionophore A23187 and protein synthesis or phosphorylation inhibitors was investigated as a means of producing human parthenogenones with one haploid pronucleus. Unfertilised human aged oocytes exposed to 5 μM A23187 for 5 min were treated with 10 μg/ml puromycin (puromycin group, 46 oocytes) or 2 mM 6-dimethylaminopurine (DMAP group, 42 oocytes) for 5 h. Oocytes treated only with A23187 served as a control (control group, 40 oocytes). After washing the oocytes, they were incubated for up to 37 h. Evidence of activation (pronuclear formation) and cleavage was observed 18 h and 42 h after A23187 treatment, respectively. Activation rates in the puromycin and DMAP groups were significantly higher than in the control group (91% (42/46) and 77% (34/44) vs 20% (8/40), p < 0.05, respectively). In the puromycin group, 81% (34/42) of the activated oocytes showed one pronucleus with the second polar body (2ndPB), whereas none (0/34) of the activated oocytes in the DMAP group extruded the 2ndPB. The cleavage rate in the puromycin group was significantly lower than in the DMAP group (38% vs 68%, p < 0.05). The activated oocytes which had one pronucleus with the 2ndPB in the puromycin group showed a haploid set of chromosomes (10/13). In conclusion, the combination of A23187 and puromycin is effective for producing human parthenogenones with one haploid pronucleus.

Artificial oocyte activation with calcium ionophore A23187 in intracytoplasmic sperm injection cycles using surgically retrieved spermatozoa

2009 ◽  
Vol 92 (1) ◽  
pp. 131-136 ◽  
Author(s):  
Edson Borges Jr. ◽  
Daniela Paes de Almeida Ferreira Braga ◽  
Tatiana Carvalho de Sousa Bonetti ◽  
Assumpto Iaconelli Jr. ◽  
José Gonçalves Franco Jr.
Keyword(s):  
Intracytoplasmic Sperm Injection ◽  
Calcium Ionophore ◽  
Calcium Ionophore A23187 ◽  
Ionophore A23187 ◽  
Oocyte Activation ◽  
Artificial Oocyte Activation

scholarly journals Successful delivery following intracytoplasmic sperm injection with calcium ionophore A23187 oocyte activation in a partially globozoospermic patient

2012 ◽  
Vol 11 (3) ◽  
pp. 159-164 ◽  
Author(s):  
Hiroshi Kamiyama ◽  
Taketoshi Shimizu ◽  
Tomomi Oki ◽  
Terumi Asada ◽  
Yasuyuki Araki ◽  
...  
Keyword(s):  
Intracytoplasmic Sperm Injection ◽  
Calcium Ionophore ◽  
Calcium Ionophore A23187 ◽  
Ionophore A23187 ◽  
Oocyte Activation
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