title: Mouse embryos stressed by physiological levels of osmolarity become arrested in the late 2-cell stage before entry into M phase.
abstract: Preimplantation mouse embryos of many strains become arrested at the 2-cell stage if the osmolarity of culture medium that normally supports development to blastocysts is raised to approximately that of their normal physiological environment in the oviduct. Arrest can be prevented if molecules that serve as "organic osmolytes" are present in the medium, because organic osmolytes, principally glycine, are accumulated by embryos to provide intracellular osmotic support and regulate cell volume. Medium with an osmolarity of 310 mOsM induced arrest of approximately 80% of CF1 mouse embryos at the 2-cell stage, in contrast to the approximately 100% that progressed beyond the 2-cell stage at 250 or 301 mOsM with glycine. The nature of this arrest induced by physiological levels of osmolarity is unknown. Arrest was reversible by transfer to lower-osmolarity medium at any point during the 2-cell stage, but not after embryos would normally have progressed to the 4-cell stage. Cessation of development likely was not due to apoptosis, as shown by lack of external annexin V binding, detectable cytochrome c release from mitochondria, or nuclear DNA fragmentation. Two-cell embryos cultured at 310 mOsM progressed through the S phase, and zygotic genome activation markers were expressed. However, most embryos failed to initiate the M phase, as evidenced by intact nuclei with decondensed chromosomes, low M-phase promoting factor activity, and an inactive form of CDK1, although a few blastomeres were arrested in metaphase. Thus, embryos become arrested late in the G(2) stage of the second embryonic cell cycle when stressed by physiological osmolarity in the absence of organic osmolytes.
authors.affiliation: School of Life Sciences, Sun Yat-sen University, Guangzhou, China.
authors.forename: Fang
authors.initials: F
authors.lastname: Wang
authors.forename: Megan
authors.initials: M
authors.lastname: Kooistra
authors.forename: Martin
authors.initials: M
authors.lastname: Lee
authors.forename: Lin
authors.initials: L
authors.lastname: Liu
authors.forename: Jay M
authors.initials: JM
authors.lastname: Baltz
mesh_terms: D000818:Animals; D015415:Biomarkers / Q000378:metabolism; D001755:Blastocyst / Q000378:metabolism / Q000473:pathology*; D001757:Blastomeres / Q000648:ultrastructure; D016203:CDC2 Protein Kinase / Q000737:chemistry / Q000378:metabolism; D002455:Cell Division; D002467:Cell Nucleus / Q000378:metabolism / Q000648:ultrastructure; D048429:Cell Size; D042002:Chromatin Assembly and Disassembly; D003433:Crosses, Genetic; D004477:Ectogenesis; D015977:Eukaryotic Initiation Factor-1 / Q000235:genetics / Q000378:metabolism; D005998:Glycine / Q000378:metabolism; D016200:Maturation-Promoting Factor / Q000378:metabolism; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D008677:Metaphase; D051379:Mice; D015778:Minor Histocompatibility Antigens / Q000235:genetics / Q000378:metabolism; D008970:Molecular Weight; D009997:Osmotic Pressure; D012333:RNA, Messenger / Q000378:metabolism; D013312:Stress, Physiological; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism; D051140:Selenoproteins
chemical_list: D015415:Biomarkers; D016203:CDC2 Protein Kinase; D015977:Eukaryotic Initiation Factor-1; D005998:Glycine; D016200:Maturation-Promoting Factor; D008565:Membrane Proteins; D015778:Minor Histocompatibility Antigens; D012333:RNA, Messenger; D014157:Transcription Factors; C534888:Zscan4d protein, mouse; C026022:eukaryotic peptide initiation factor-1A; C532537:Selenos protein, mouse; D051140:Selenoproteins