RAPID INHIBITION OF MESSENGER-RNA SYNTHESIS DURING PREIMPLANTATION EMBRYO DEVELOPMENT - VITAL PERMEABILIZATION BY LYSOLECITHIN POTENTIATES THE ACTION OF ALPHA-AMANITIN

Lysolecithin was used to permeabilize embryonic cells to impermeant co mpounds without compromising embryo viability. Within 2 min of exposur e to 0.05% lysolecithin, mouse morulae became permeable to inositol 1, 4,5-trisphosphate leading to the release of calcium from intracellular stores. Although these morulae remained permeable for at least 2 h, t hey developed normally to the blastocyst stage during subsequent cultu re in vitro. Lysolecithin permeabilization was then used to potentiate the internalization of alpha-amanitin. Pretreatment with lysolecithin for 2 min markedly accelerated the onset of transcriptional inhibitio n from 3 h to 10 min after alpha-amanitin addition. The rapid inhibiti on by alpha-amanitin of mRNA synthesis in lysolecithin-treated embryos provided a precise method for delineating developmentally important t ranscriptional periods. Using this protocol, we found that mRNAs requi red for embryonic cavitation were synthesized between 87 and 91 h post -hCG, shortly before blastocoel formation commenced. (C) 1995 Academic Press, Inc.