Abnormal development and dye coupling produced by antisense RNA to gap junction protein in mouse preimplantation...

Antisense RNA to the 27/32-kDa rat liver gap junction (GJ) protein was used to explore the role of GJs in preimplantation embryos. When all blastomeres of two- and four-cell embryos were injected with GJ antisense RNA, the percentage of embryos compacted at 60 hr of development was reduced to less than 20%, while 90% of uninjected embryos and 75% of embryos injected with an unrelated RNA were compacted. When most cells of compacted eight-cell embryos were injected with the GJ antisense RNA, 20% of the embryos were decompacted and only 5% had developed to the blastocyst stage at 90 hr, when blastulation had occurred in 90% of the control embryos. When antisense RNA was injected in one blastomere of four-cell embryos, 40% of the embryos presented a large cell that was not included in the compacted embryo at the time of compaction, and an additional 30% of the embryos had two smaller, excluded blastomeres. These excluded cells were identified as the injected cell with a rhodamine-conjugated dextran marker. To assess effects on junctional communication, one blastomere of some embryos was injected with Lucifer yellow, a GJ-penetrating dye, at various times after a blastomere was injected with antisense RNA. The dye was visible in the whole cell mass of control embryos, but it was excluded from a portion of experimental embryos when the delay between the RNA and the Lucifer yellow injections was 1 hr or longer.