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Updated: Jul 11, 2026
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Published on: September 13, 2022
Overcoming MIII arrest from spontaneous activation in cultured rat oocytes
Vasiliy Galat1, Yue Zhou, Greg Taborn
1Department of Pediatrics, Northwestern University Feinberg School of Medicine and the Developmental Biology Program of Children's Memorial Research Center, Chicago, Illinois 60614, USA.
Abstract:
The rat oocyte spontaneously activates under a wide variety of conditions. This process progresses to MIII arrest that is not responsive to parthenogenetic activation and development. Insofar as activation involves extrusion of the second polar body (PBII), we set out to determine if preventing this step by inhibiting microfilaments would change the course of spontaneous activation (SA). In particular, how long does the effect of SA persist while retaining reversibility of PBII extrusion once inhibitors are removed? We wanted to determine if the eggs would be responsive to parthenogenetic activation and capable of resuming development once a permanent inhibition is achieved. We set out to determine whether SA would depend on the ovular age of oocytes. Inhibiting of PBII extrusion was achieved by affecting microtubules with demecolcine or nocodazole or actin filaments with cytochalasin B (CB) and cytochalasin D (CD). We found that all oocytes undergo SA and progression to MIII; however, the rapidity of spontaneous activation is a function of the ovular age of the oocyte. The resumption of the meiosis period changes dramatically from 20 to 180 min with decreasing ovular age. We established that suppression of PB formation can be effectively achieved in oocytes of younger ovular age, and that inhibition of PB extrusion became irreversible after 3.5 h of treatment. We established that drug-treated oocytes could undergo subsequent reactivation and in vitro development to blastocysts. The rate of in vitro development of cytochalasin-treated group was comparable to parthenogenetic controls, while nocodazole and demecolcine produced oocytes that developed at lower frequencies. Thus, the application of the microfilament inhibiting drugs helps to overcome the negative effect of SA that results in MIII arrest. Here we also show optimized parthenogenetic stimulation that resulted in development to the blastocyst stage at frequency comparable to development of fertilized embryos.
Insights
Inhibiting the second polar body extrusion in rat oocytes prevents spontaneous activation arrest. This method allows for subsequent parthenogenetic activation and in vitro development to blastocysts.
Area of Science:
- Reproductive Biology
- Developmental Biology
- Cell Biology
Background:
- Rat oocytes spontaneously activate (SA), arresting at MIII, unresponsive to further activation.
- Second polar body extrusion (PBII) is a key event in oocyte activation.
Purpose of the Study:
- To investigate if inhibiting PBII extrusion affects spontaneous activation (SA) progression and reversibility.
- To determine if oocytes with inhibited PBII extrusion can resume development.
- To assess the influence of oocyte ovular age on SA and PBII extrusion.
Main Methods:
- Oocytes were treated with microtubule inhibitors (demecolcine, nocodazole) or actin filament inhibitors (cytochalasin B/D).
- PBII extrusion was monitored, and inhibition reversibility was assessed over time.
- Treated oocytes were subjected to parthenogenetic activation and in vitro development to the blastocyst stage.
Main Results:
- All oocytes underwent SA and MIII arrest; SA rapidity correlated with ovular age.
- PBII extrusion suppression was effective, becoming irreversible after 3.5 hours.
- Cytochalasin-treated oocytes developed to blastocysts at rates comparable to controls; microtubule inhibitor groups showed lower rates.
Conclusions:
- Inhibiting microfilaments overcomes SA-induced MIII arrest, enabling subsequent development.
- Oocyte ovular age influences the timing of SA and PBII extrusion.
- Optimized parthenogenetic stimulation achieved blastocyst development rates comparable to fertilized embryos.
