Utilizing some known physiological preovulatory hormone levels and specific cytological end-points, experiments were carried out with two different media to obtain a clearer understanding of meiotic maturation in vitro of ewe, cow, monkey and human oocytes. In the first series of experiments, it was shown that, in McCoy's medium, ovine LH, prostaglandin E1 and ovine prolactin increased the percentage of divisions of ewe oocytes, without cAMP as a demonstrable intermediate. Monkey oocytes incubated in McCoy's medium with fetal calf serum were stimulated by ovine LH, prostaglandin E2 or cAMP. Human oocytes were stimulated to divide in Diploid medium by ovine LH, oestradiol-17β with LH added later, or prostaglandin E1. Diploid medium containing added sheep serum markedly inhibited the meiotic activity of ewe oocytes when ovine LH, HCG, cAMP, prostaglandin E1 and E2 or prolactin were added. This effect was studied by separating the contained fetal calf and sheep sera. It was noted that when LH, HCG or prostaglandins E1 and E2 were added to a system utilizing fetal calf serum, only spontaneous resumption of meiosis occurred: stimulated and spontaneous activity were reduced when cAMP or prolactin were added. The inhibition of ewe oocyte maturation in Diploid medium with sheep serum and additives was also seen with cow oocytes incubated in Diploid medium containing fetal calf serum and LH, HCG, cAMP or prostaglandin E1 and E2. Cow oocytes incubated in McCoy's medium with equivalent fetal calf serum and added LH were sharply inhibited but spontaneous maturation persisted in the presence of added HCG, cAMP or bovine prolactin.