Mouse blastocyst outgrowth and implantation rates following exposure to ethanol or A23187 during culture in vitro

Abstract

This study focused on the effects of ethanol on blastocyst outgrowth and implantation in mice. Blastocysts were exposed to ethanol in Ham's F10 medium and then cultured free of ethanol on fibronectin-coated Petri dishes to assess trophoblast cell adhesion and migration. The time necessary for half of the embryos to outgrow was significantly less (P < 0.05) following treatment with 0.1%, 0.2%, 0.4% or 1.0% (w/v) ethanol for either 5 min or 24 h compared with controls. The rate of trophoblast cell migration was determined by measuring the mean area of outgrowing embryos using an image analysis system. Blastocysts exposed to ethanol for 5 min produced a greater (P < 0.05) average outgrowth area than did stage-matched controls. Acceleration of blastocyst cavitation by ethanol is known to be associated with an increase in the intracellular concentration of calcium. Here, treatment with the calcium ionophore A23187 stimulated (P < 0.05) trophoblast outgrowth and accelerated (P < 0.05) the rate of cell migration. In an attempt to correlate the effect of ethanol on outgrowth in vitro with implantation in utero, cultured blastocysts were either not exposed to ethanol or exposed to 0.1% ethanol for 5 min and transferred 24 h later to the uteri of pseudopregnant dams. The implantation rate (39.4%, n = 376) and the rate of development to term (2.45 pups per mouse, n = 20) were higher (P < 0.05) in mice receiving ethanol-treated embryos compared with those receiving control embryos (20.8%, n = 331; 1.16 pups per mouse, n = 18, respectively). The fetal survival rate of implanted embryos was not significantly different between the ethanol-treated (33.1%) and the control (30.4%) groups. These data suggest that treatment of cultured mouse blastocysts with agents that increase the intracellular calcium concentration accelerate trophoblast outgrowth in vitro and facilitate implantation in utero.