AbstractPreimplantation mouse embryos (ICR strain) were removed from the oviducts of pregnant females at 55, 60, and 65 hours after ovulation and cultured in a medium containing inorganic salts, horse serum, and sodium pyruvate. Experimental embryos were exposed to a range of dosages of actinomycin D (10−1–10−5μg/ml) and were observed at 24 and 48 hours after the initiation of the experiment. The results obtained were independent of embryonic stage but demonstrated a distinct dose‐dependency. Exposure to 10−1μg/ml was incompatible with further development. Embryos exposed to 10−2μg/ml formed blastocysts (24 hours) at a rate lower than corresponding controls and all were degenerated at the termination of the experiment (48 hours). Blastocyst formation by embryos exposed to 10−3μg/ml occurred at a rate that was equal to or exceeded control values, but a significant percentage of these blastocysts were degenerated after 48 hours in vitro. Embryos exposed to the lower dosages (10−4and 10−5μg/ml) did not differ from corresponding control embryos with respect to the percentage of blastocysts observed at 24 and 48 hours.These results suggest that actinomycin D affects two developmental processes: blastocyst formation and blastocyst survival or differentiation. The latter of these is more sensitive to the developmental effects of the agent and is at least circumstantially related to the early appearance of the nucl