A total of 486 unfertilized, aged human oocytes were exposed to ethanol, calcium ionophore A23187, phorbol ester or puromycin and examined for evidence of activation. Five per cent of control oocytes (3/58) were spontaneously activated. Of the two agents which cause the release of intracellular Ca2+ ions, Ca2+ ionophore induced activation of only 16% of unfertilized oocytes, while ethanol did not have any effect. Phorbol ester, a stimulator of protein kinase C, also resulted in limited activation (14% of oocytes). In contrast, puromycin, an inhibitor of protein synthesis, resulted in activation of 91% of the exposed oocytes. It is speculated that puromycin probably inhibits a specific cytostatic factor or factors which are responsible for maintenance of the metaphase II block. Morphologically activated oocytes usually retained the second polar body and formed subnuclei. The developmental potential of activated oocytes appeared to be reduced, with only some oocytes capable of a single division.