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Maturation conditions, post-ovulatory age, medium pH, and ER stress affect [Ca2+]i oscillation patterns in mouse oocytes

  • Gamete Biology
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Abstract

Insufficiency of oocyte activation impairs the subsequent embryo development in assisted reproductive technology (ART). Intracellular Ca2+ concentration ([Ca2+]i) oscillations switch the oocytes to resume the second meiosis and initiate embryonic development. However, the [Ca2+]i oscillation patterns in oocytes are poorly characterized. In this study, we investigated the effects of various factors, such as the oocytes age, pH, cumulus cells, in vitro or in vivo maturation, and ER stress on [Ca2+]i oscillation patterns and pronuclear formation after parthenogenetic activation of mouse oocytes. Our results showed that the oocytes released to the oviduct at 17 h post-human chorionic gonadotrophin (hCG) displayed a significantly stronger [Ca2+]i oscillation, including higher frequency, shorter cycle, and higher peak, compared with oocytes collected at earlier or later time points. [Ca2+]i oscillations in acidic conditions (pH 6.4 and 6.6) were significantly weaker than those in neutral and mildly alkaline conditions (pH from 6.8 to 7.6). In vitro-matured oocytes showed reduced frequency and peak of [Ca2+]i oscillations compared with those matured in vivo. In vitro-matured oocytes from the cumulus-oocyte complexes (COCs) showed a significantly higher frequency, shorter cycle, and higher peak compared with the denuded oocytes (DOs). Finally, endoplasmic reticulum stress (ER stress) severely affected the parameters of [Ca2+]i oscillations, including elongated cycles and lower frequency. The pronuclear (PN) rate of oocytes after parthenogenetic activation was correlated with [Ca2+]i oscillation pattern, decreasing with oocyte aging, cumulus removal, acidic pH, and increasing ER stress. These results provide fundamental but critical information for the mechanism of how these factors affect oocyte activation.

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Data availability

The datasets used and/or analyzed during the current study are available from the corresponding author on reasonable request.

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This work was sponsored by the National Natural Science Foundation of China (No 81671425 and No 81901477).

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Authors and Affiliations

  1. Reproductive Medicine Center, Guangdong Second Provincial General Hospital, Guangzhou, 510000, China

    Rui-Ying Yuan, Feng Wang, Sen Li, Jun-Yu Ma, Lei Guo, Xiao-Long Li, Hai-Jing Zhu, Xie Feng, Zi-Bin Lin & Xiang-Hong Ou

  2. State Key Laboratory of Stem Cell and Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing, 100101, China

    Feng Wang, Qian-Nan Li & Qian Zhou

  3. Department of Veterinary Pathobiology, University of Missouri, Columbia, MO, 65211, USA

    Heide Schatten

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  1. Rui-Ying Yuan
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  2. Feng Wang
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Supplemental Figure S1

Intracellular free calcium ([Ca2+]i) oscillations and parameters. A to E indicate different stages of two continuous [Ca2+]i oscillations. “Ca2+ move speed" was the changing fluorescence value every 20 s. We defined the length from the beginning of first [Ca2+]i rise to the beginning of next [Ca2+]i rise as one "cycle"; time from beginning to end of an [Ca2+]i rise as “maintain”; the maximum fluorescence value and maximum changing fluorescence value as “peak” and “move”; and “△peak" was defined as maximum [Ca2+]i fluorescence value minus baseline values. The baseline was defined as the stable minimum [Ca2+]i fluorescence value. The bright field and fluorescent embryo image specifications were as follows: magnification: 200X, time-point: 8008.867s - 9732.370s, wavelength: 191s. The embryos were derived from the 17 h group, and Fluo-4AM fluorescent dye was used. (JPG 160 kb)

Supplemental Figure S2

Relative gene expression was determined using the 2ΔΔCt method normalized to GAPDH. Expression of GRP78 and Chop in oocytes with endoplasmic reticulum stress-induced by tunicamycin (TM) in a dose-dependent manner. RNA was extracted from 100 mouse oocytes for each group. Data represent mean±SD, and Chi-square test and one-way ANOVA are used for statistical analysis. *P<0.05 versus the control group. (JPG 137 kb)

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Yuan, RY., Wang, F., Li, S. et al. Maturation conditions, post-ovulatory age, medium pH, and ER stress affect [Ca2+]i oscillation patterns in mouse oocytes. J Assist Reprod Genet 38, 1373–1385 (2021). https://doi.org/10.1007/s10815-021-02100-9

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