Following fertilization, the newly formed zygote faces several critical decisions regarding cell fate and lineage commitment. First, the parental genomes must be reprogrammed and reset for the zygotic genome to assume responsibility for gene expression. Second, blastomeres must be committed to form either the inner cell mass or trophectoderm before implantation. A variety of epigenetic mechanisms underlies each of these steps, allowing for proper activation of transcriptional circuits which function to specify a cell's identity and maintain or adjust that state as developmental and environmental conditions dictate. These epigenetic mechanisms encompass DNA methylation, post-translational histone modification, chromatin remodeling, and alterations in nuclear architecture. In recent years, stem cells derived from the inner cell mass have been used to examine the epigenetic pathways that regulate pluripotency, differentiation, and lineage commitment. From a technical standpoint, embryonic stem cells provide an easier system to work with compared to preimplantation embryos; however, it is currently unknown how closely the epigenetic mechanisms of cultured stem cells resemble their counterparts in the intact embryo. Furthermore, it remains unclear how similar the reprogramming pathways in artificially created systems, such as nuclear transfer-derived embryos and induced pluripotent stem cells, are to those in naturally created embryos. In this review, we summarize the current knowledge of epigenetic influences during preimplantation development and shed light on the extent to which these pathways are conserved in cultured pluripotent cells in vitro. In doing so, we demonstrate the critical role that epigenetic mechanisms play in the establishment of cell fate during the earliest stages of mammalian development. Birth Defects Research (Part C) 87:297–313, 2009. © 2009 Wiley-Liss, Inc.