Dysfunction in gap junction intercellular communication induces aberrant behavior of the inner cell mass and frequent collapses of expanded blastocysts in mouse embryos

• Published in: Journal of assisted reproduction and genetics Vol. 32; no. 6; pp. 969 - 976
• Main Authors: Togashi, Kazue, Kumagai, Jin, Sato, Emiko, Shirasawa, Hiromitsu, Shimoda, Yuki, Makino, Kenichi, Sato, Wataru, Kumazawa, Yukiyo, Omori, Yasufumi, Terada, Yukihiro
• Format: Journal Article
• Language: English
• Published: New York Springer US 01-06-2015; Springer Nature B.V
• Subjects: Animals; Blastocyst Inner Cell Mass - drug effects; Blastocyst Inner Cell Mass - metabolism; Blastocyst Inner Cell Mass - ultrastructure; Cell Communication - drug effects; Cell Communication - physiology; Cell Differentiation - drug effects; Communication; Cytoplasm - ultrastructure; Embryo Biology; Embryo, Mammalian - cytology; Embryo, Mammalian - drug effects; Embryo, Mammalian - metabolism; Embryonic Development - drug effects; Embryos; Female; Females; Gap Junctions - drug effects; Gap Junctions - physiology; Gap Junctions - ultrastructure; Gynecology; Heptanol - pharmacology; Human Genetics; Medicine; Medicine & Public Health; Mice; Mice, Inbred ICR; Morphology; Oleic Acids - pharmacology; Proteins; Reproductive Medicine; Signal transduction; Time-Lapse Imaging; United States > US; Japan; Delayed blastocyst formation; Gap junction inhibitor; Time-lapse movie observation; Blastocoel collapse; ICM division
• ISSN: 1058-0468; 1573-7330
• DOI: 10.1007/s10815-015-0479-1

Purpose
We investigated the role of gap junctions (GJs) in embryological differentiation, and observed the morphological behavior of the inner cell mass (ICM) by time-lapse movie observation (TLM) with gap junction inhibitors (GJis).
Methods
ICR mouse embryos were exposed to two types of GJis in CZB medium: oleamide (0 to 50 μM) and 1-heptanol (0 to 10 mM). We compared the rate of blastocyst formation at embryonic day 4.5 (E4.5) with E5.5. We also observed and evaluated the times from the second cleavage to each embryonic developing stage by TLM. We investigated embryonic distribution of DNA, Nanog protein, and Connexin 43 protein with immunofluorescent staining.
Results
In the comparison of E4.5 with E5.5, inhibition of gap junction intercellular communication (GJIC) delayed embryonic blastocyst formation. The times from the second cleavage to blastocyst formation were significantly extended in the GJi-treated embryos (control vs with oleamide, 2224 ± 179 min vs 2354 ± 278 min,
p
 = 0.013). Morphological differences were traced in control versus GJi-treated embryos until the hatching stage. Oleamide induced frequent severe collapses of expanded blastocysts (77.4 % versus 26.3 %,
p
 = 0.0001) and aberrant ICM divisions connected to sticky strands (74.3 % versus 5.3 %,
p
 = 0.0001). Immunofluorescent staining indicated Nanog-positive cells were distributed in each divided ICM.
Conclusions
GJIC plays an important role in blastocyst formation, collapses of expanded blastocysts, and the ICM construction in mouse embryos.