Gap junctional intercellular communication is assumed to play an important role during pre- and peri-implantation development. In this study, we eliminated connexin43 (Cx43) and connexin45 (Cx45), major gap junctional proteins in the pre- and peri-implantation embryo. We generated Cx43−/−Cx45−/− embryos by Cx43+/−Cx45+/− intercrossing, because mice deficient in Cx43 (Cx43−/−) exhibit perinatal lethality and those deficient in Cx45 (Cx45−/−) exhibit early embryonic lethality. Wild-type, Cx43−/−, Cx45−/−, and Cx43−/−Cx45−/− blastocysts all showed similar outgrowths in in vitro culture. Moreover, Cx43−/−Cx45−/− embryos were obtained at the expected Mendelian ratio up to embryonic day 9.5, when the Cx45−/− mutation proved lethal. The Cx43−/−Cx45−/− embryos seemed to have no additional developmental abnormalities in comparison with the single knockout strains. Thus, pre- and peri-implantation development does not require Cx43 and Cx45. Other gap junctional proteins are expressed around these stages and these may compensate for the lack of Cx43 and Cx45.

Many studies of the main gap junction protein, Cx43, have been conducted in porcine oocyte research, but they have been limited to investigations of cumulus–oocyte complexes (COCs). In this study, we verified Cx43 not in COCs, but in porcine oocytes during maturation, and conducted a quantitative time course analysis. The location and dynamics of Cx43 were examined by immunocytochemistry and western blotting, respectively. COCs were cultured in NCSU23 medium and processed for immunocytochemistry and western blotting at 0, 14, 28, and 42 h after denuding. A Cx43 signal was detected on oolemmas, transzonal projections and the surface of zona pellucidae. Western blotting showed that Cx43 band density increased from 0 to 14 h, and gradually decreased thereafter. Our results clarified that Cx43 is localized in the ooplasmic membrane through zona pellucidae and its level changes over time during culture in porcine oocytes.