Control of gap junction formation in early mouse embryos.

Intercellular communication via gap junctions begins in the eight-cell stage in early mouse embryos. We have studied the timing of this event in relation to compaction, and have begun to explore some of the possible control mechanisms underlying it. Gap junction formation was inferred by measuring ionic coupling as well as by observing the intercellular transfer of fluorescent dye. Embryos were obtained early on Day 3 of pregnancy by flushing the oviducts of HA/ICR mice that had been mated with CB6F1/J males. Gap junctions were detected only in those embryos which had achieved the fully compacted state. Inhibition of protein synthesis by cycloheximide treatment beginning as early as the late four-cell stage failed to block compaction or the acquisition of gap junctions, demonstrating that the necessary proteinaceous components are present in advance of these events. In order to test the possibility that gap junctions could be induced to form prematurely, fully compacted, communication-competent eight-cell embryos were aggregated with two- or four-cell embryos. Even after 10 hr of aggregation, no interembryonic gap junctions could be detected. Fully compacted eight-cell embryos when aggregated with each other, however, became ionically coupled within 3-5 hr. The number of interembryonic junctional channels was judged to be effectively small, since the aggregated embryos exhibited obvious ionic coupling but very weak dye coupling. In contrast to gap junction formation within embryos, junction formation between embryos was blocked by cycloheximide. These results demonstrate that gap junction formation in early mouse embryos is under precise temporal control, involving the assembly or mobilisation of preexisting components. This stockpile of components is either unavailable or insufficient to allow the formation of additional gap junctions between aggregated communication-competent embryos without new protein synthesis.