Comparison of method for activating mouse oocytes for spermatid nucleus tranfer

Zygote ◽  
1996 ◽  
Vol 4 (04) ◽  
pp. 269-274 ◽  
Author(s):  
Isoji Sasagawa ◽  
R. Yanagimachi
Keyword(s):  
Electrical Stimulation ◽  
Embryo Development ◽  
Electrical Pulse ◽  
Oocyte Activation ◽  
Cell Nuclei ◽  
Mouse Oocytes ◽  
Development Rates ◽  
Spermatid Nucleus ◽  
Additional Stimulation

SummaryIn the mouse, mature oocytes injected with prespermatozoal cell nuclei remain unactivated. Additional stimulation is needed to trigger oocyte activation leading to embryo development. We compared various electrical stimulations, treatment with cycloheximide alone or in combination with electrical stimulation, and injection of sperm-borne oocyte-activating factor (oscillogen) in terms of their oocyte activation and embryo development rates. Of all the treatments tested, a single electrical pulse (1.0 kV / cm, 128 μs) was the simplest, yet very effective, in allowing the development of the oocytes injected with spermatid nuclei.

Nuclear envelope removal/maintenance determines the structural and functional remodelling of embryonic red blood cell nuclei in activated mouse oocytes

Zygote ◽  
1998 ◽  
Vol 6 (1) ◽  
pp. 65-73 ◽  
Author(s):  
Daniel Szöllösi ◽  
Renata Czołowska ◽  
Ewa Borsuk ◽  
Maria S. Szöllösi ◽  
Pascale Debey
Keyword(s):  
Nuclear Envelope ◽  
Transcriptional Activity ◽  
Chromatin Condensation ◽  
Oocyte Activation ◽  
Cell Nuclei ◽  
Nuclear Envelope Breakdown ◽  
Mouse Oocytes ◽  
Female Pronucleus ◽  
Mouse Fetuses

SummaryNuclei of embryonic red blood cells (e-RBC) from 12-day mouse fetuses are arrested in Go phase of the cell cycle and have low transcriptional activity. These nuclei were transferred with help of polyethylene glycol (PEG)-mediated fusion to parthenogenetically activated mouse oocytes and heterokaryons were analysed for nuclear structure and transcriptional activity. If fusion proceeded 25–45 min after oocyte activation, e-RBC nuclei were induced to nuclear envelope breakdown and partial chromatin condensation, followed by formation of nuclei structurally identical with pronuclei. These ‘pronuclei’, similar to egg (female) pronuclei, remained transcriptionally silent over several hours of in vitro culture. If fusion was performed 1 h or later (up to 7 h) after activation, the nuclear envelope of e-RBC nuclei remained intact and nuclear remodelling was less spectacular (slight chromatin decondensation, formation of nucleolus precursor bodies). These nuclei, however, reinforced polymerase-II-dependent transcription within a few hours of in vitro culture. Our present experiments, together with our previous work, demonstrate that nuclear envelope breakdown/maintenance are critical events for nuclear remodelling in activated mouse oocytes and that somatic dormant nuclei can be stimulated to renew transcription at a time when the female pronucleus remains transcriptionally silent.

scholarly journals Prolonged exposure to hyaluronidase decreases the fertilization and development rates of fresh and cryopreserved mouse oocytes

2014 ◽  
Vol 60 (6) ◽  
pp. 454-459 ◽  
Author(s):  
Yuta ISHIZUKA ◽  
Toru TAKEO ◽  
Satohiro NAKAO ◽  
Hidetaka YOSHIMOTO ◽  
Yumiko HIROSE ◽  
...  
Keyword(s):  
Prolonged Exposure ◽  
Mouse Oocytes ◽  
Development Rates

scholarly journals Effects of Porcine Immature Oocyte Vitrification on Actin Microfilament Distribution and Chromatin Integrity During Early Embryo Development in vitro

2021 ◽  
Vol 9 ◽  
Author(s):  
Alma López ◽  
Yvonne Ducolomb ◽  
Eduardo Casas ◽  
Socorro Retana-Márquez ◽  
Miguel Betancourt ◽  
...  
Keyword(s):  
Embryo Development ◽  
Early Embryo ◽  
Actin Microfilaments ◽  
Cell Protection ◽  
Developmental Potential ◽  
Early Embryo Development ◽  
Immature Oocytes ◽  
Actin Microfilament ◽  
Development Rates ◽  
Chromatin Integrity

Vitrification is mainly used to cryopreserve female gametes. This technique allows maintaining cell viability, functionality, and developmental potential at low temperatures into liquid nitrogen at −196°C. For this, the addition of cryoprotectant agents, which are substances that provide cell protection during cooling and warming, is required. However, they have been reported to be toxic, reducing oocyte viability, maturation, fertilization, and embryo development, possibly by altering cell cytoskeleton structure and chromatin. Previous studies have evaluated the effects of vitrification in the germinal vesicle, metaphase II oocytes, zygotes, and blastocysts, but the knowledge of its impact on their further embryo development is limited. Other studies have evaluated the role of actin microfilaments and chromatin, based on the fertilization and embryo development rates obtained, but not the direct evaluation of these structures in embryos produced from vitrified immature oocytes. Therefore, this study was designed to evaluate how the vitrification of porcine immature oocytes affects early embryo development by the evaluation of actin microfilament distribution and chromatin integrity. Results demonstrate that the damage generated by the vitrification of immature oocytes affects viability, maturation, and the distribution of actin microfilaments and chromatin integrity, observed in early embryos. Therefore, it is suggested that vitrification could affect oocyte repair mechanisms in those structures, being one of the mechanisms that explain the low embryo development rates after vitrification.

Inhibition of protein kinases by 6-dimethylaminopurine accelerates the transition to interphase in activated mouse oocytes

1993 ◽  
Vol 104 (3) ◽  
pp. 861-872 ◽  
Author(s):  
M.S. Szollosi ◽  
J.Z. Kubiak ◽  
P. Debey ◽  
H. de Pennart ◽  
D. Szollosi ◽  
...  
Keyword(s):  
Protein Phosphorylation ◽  
Kinase Inhibitor ◽  
Polar Body ◽  
Histone H1 ◽  
Sperm Chromatin ◽  
Oocyte Activation ◽  
Mouse Oocytes ◽  
Spindle Rotation ◽  
Polar Body Extrusion ◽  
Pronuclear Formation

Mouse oocyte activation is followed by a peculiar period during which the interphase network of microtubules does not form and the chromosomes remain condensed despite the inactivation of MPF. To evaluate the role of protein phosphorylation during this period, we studied the effects of the protein kinase inhibitor 6-dimethylaminopurine (6-DMAP) on fertilization and/or parthenogenetic activation of metaphase II-arrested mouse oocytes. 6-DMAP by itself does not induce the inactivation of histone H1 kinase in metaphase II-arrested oocytes, and does not influence the dynamics of histone H1 kinase inactivation during oocyte activation. However, 6-DMAP inhibits protein phosphorylation after oocyte activation. In addition, the phosphorylated form of some proteins disappear earlier in oocytes activated in the presence of 6-DMAP than in the activated control oocytes. This is correlated with the acceleration of some post-fertilization morphological events, such as sperm chromatin decondensation and its transient recondensation, formation of the interphase network of microtubules and pronuclear formation. In addition, numerous abnormalities could be observed: (1) the spindle rotation and polar body extrusion are inhibited; (2) the exchange of protamines into histones seems to be impaired, as judged by the morphology of DNA fibrils by electron microscopy; (3) the formation of a new nuclear envelope around the sperm chromatin proceeds prematurely, while recondensation is not yet completed. These observations suggest that the 6-DMAP-sensitive kinase(s) is (are) involved in the control of post-fertilization events such as the formation of the interphase network of microtubules, the remodelling of sperm chromatin and pronucleus formation.

In vitro transcription and micro-injection of Kdm4a mRNA into mouse oocytes

2020 ◽  
Author(s):  
ADITYA SANKAR
Keyword(s):  
Survival Rate ◽  
Embryo Development ◽  
In Vitro Transcription ◽  
Mouse Oocytes ◽  
The University ◽  
The Impact

Abstract This experiment describes the in vitro transcription of Kdm4a wildtype and H188A catalytic dead mRNA. This details also its subsequent injection into mouse oocytes followed my IVF to track the impact on embryo development. The procedure is technically challenging and performed by the Transgenic Core Facility at the University of Copenhagen. Oocytes have a poorer survival rate following mRNA inject as against zygotes. However the objective was to demonstrate the earliest stage of intervention to rescue developmental failure of KDM4A maternal zygotic mutant embryos

238 CAFFEINE PREVENTS A LOSS OF CALCIUM OSCILLATORY RESPONSE ASSOCIATED WITH POSTOVULATORY AGING OF MOUSE OOCYTES

2009 ◽  
Vol 21 (1) ◽  
pp. 217
Author(s):  
T. Wakai ◽  
N. Zhang ◽  
R. A. Fissore
Keyword(s):  
Subsequent Development ◽  
Oocyte Quality ◽  
Developmental Competence ◽  
Oscillatory Activity ◽  
Strontium Chloride ◽  
Oocyte Activation ◽  
Mouse Oocytes ◽  
Oocyte Aging ◽  
Postovulatory Aging

Numerous studies have demonstrated that postovulatory aging of oocytes prior to fertilization has detrimental effects on oocyte quality and developmental competence. Oocyte aging is accompanied by abnormal oocyte activation and subsequent development, suggesting a disruption of Ca2+ oscillations after fertilization. The inositol 1,4,5-trisphosphate receptor type 1 (IP3R1) in mammals is responsible for the majority of Ca2+ release during fertilization (Miyazaki S et al. 1993 Dev. Biol.). Previously, we reported that phosphorylation of IP3R1 at an MPM-2 epitope may play an important role in facilitating the induction of Ca2+ oscillations at the MII stage (Lee B et al. 2006 Development), indicating that IP3R1 phosphorylation may be a good indicator of the health of the oocyte. However, few studies have investigated the alteration of the Ca2+ signaling and IP3R1 function associated with oocyte aging. On the other hand, a previous report showed that caffeine increased MPF activity and suppressed fragmentation after parthenogenetic activation of aged oocytes (Kikuchi K et al. 2000 Biol. Reprod.). Therefore, the purpose of the present study was to examine whether and how Ca2+ oscillatory activity changes during oocyte aging and to test if caffeine prevents the negative effects of oocyte aging. MII mouse oocytes were collected 14 h after hCG injection and cultured in vitro for 8, 24 or 48 h with or without caffeine (5 or 10 mm). Oocyte quality was assessed by the occurrence of spontaneous fragmentation, monitoring of Ca2+ oscillations after exposure to 10 mm strontium chloride, Western blot analysis of IP3R1 phosphorylation and immunostaining of IP3R1. In oocytes in vitro aged for 8 h, the duration of the first Ca2+ rise was significantly decreased compared with fresh MII oocytes, although this reduction was not observed in MII oocytes treated with 5 mm caffeine. The phosphorylation of IP3R1 at the MPM-2 epitope was slightly decreased during oocyte aging in both caffeine and noncaffeine treatment. Importantly, whereas IP3R1 in MII oocytes treated for 8 h with 5 mm caffeine displayed the typical cortical cluster organization, IP3R1 in aged oocytes without caffeine became dispersed in the cytoplasm. In addition, caffeine significantly suppressed the spontaneous fragmentation that is normally observed by 48 h of in vitro culture. These results suggest that the Ca2+ oscillatory activity is compromised during oocyte aging and caffeine prevents the loss of integrity of Ca2+ signaling possibly by keeping the cortical distribution of IP3R1.

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