Delayed implantation, considered a state of suspended animation, is widespread in mammals. Blastocysts under this condition remain dormant for an extended period but resume implantation competence upon favorable conditions. The underlying mechanism by which extended longevity of dormant blastocysts is maintained is not clearly understood. Using autophagy markers and the well-defined delayed implantation model in mice, we show that autophagy is important for the extended longevity of dormant blastocysts in utero during delayed implantation. However, prolonged dormancy leads to reduced developmental competency of blastocysts and cellular damage with compromised pregnancy outcome. Estrogen supplementation, which activates implantation of dormant blastocysts, induces the formation of multivesicular bodies in the trophectoderm in vivo. Collectively, our results suggest that autophagy is a critical cellular mechanism that is utilized for the prolonged survival of dormant blastocysts. (Endocrinology 152: 2067-2075, 2011)

Endocannabinoid signaling plays various roles in directing reproductive processes. Mouse embryos are shown to express high levels of CB1 receptor (CB1R). Low concentrations of anandamide stimulate embryo growth and implantation but at higher concentrations it adversely affects implantation. We tested the hypothesis that high levels of endocannabinoids cause autophagic activation and cell death in preimplantation mouse embryos. We used methanandamide (METH), a selective CB1R agonist, to examine the effect of heightened endocannabinoid signaling on autophagy in mouse embryos. Western blotting, immunofluorescence staining, transmission electron microscopy and TUNEL analysis were performed. We observed that METH treatment in vitro or in vivo up-regulated autophagic response in preimplantation mouse embryos. In blastocysts, apoptosis was also increased after METH injections. At 28 nM, which is considered a high physiological dose to embryonic cells, METH up-regulated autophagic activation in trophoblast stem cells. This work demonstrates for the first time that blastocysts respond to higher than normal levels of endocannabinoid by increasing autophagic activation and apoptosis.

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