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Rats were killed by decapitation at 3-h intervals during a 48-h delay of ovulation induced by sodium pentobarbital, as well as during the ensuing delayed proestrus, estrus and the first 4 days of gestation. Control animals were killed at the same intervals following injections of vehicle. Blood was collected and analyzed for LH, FSH, prolactin, progesterone and estradiol-17beta to determine if alterations in hormonal levels could account for the abnormal embryonic development which follows delayed ovulation. Hormonal concentrations in plasma were measured by radioimmunoassay except for progesterone, which was determined by competitive protein binding. Embryos were examined to verify the occurrence of abnormal development. Rate of oocyte maturation was studied in serial sections of ovaries from all animals killed at 30-min intervals on the afternoon of proestrus. Oocytes remained in meiotic arrest during the 48-h delay of ovulation, but resumed maturation at the expected time on the afternoon of the preovulatory surge of gonadotropins. Following delayed ovulation there was a significant increase in fertilized ova that were undergoing degeneration (6.5 vs 1.9%), abnormal development (3.7 vs 0.7%), and retarded development (3.1 vs 0.2%). A number of alterations in hormonal levels occurred during the prolonged preovulatory period. The proestrous surge of LH in plasma was completely suppressed on both days of treatment with Nembutal, but exhibited a normal preovulatory peak on the following day. FSH showed a small but significant rise on both days of treatment. Peak concentrations of FSH on the following day were not different in magnitude from the normal preovulatory surge, although the duration of this surge was shortened by 12 h. Prolactin and progesterone concentrations in plasma were only partially suppressed on the days of treatment, while on the following day preovulatory levels of prolactin were significantly greater than in controls. The proestrous peak levels of estradiol occurred prior to the first Nembutal injection and declined more gradually after this treatment. Plasma levels of estradiol remained above basal levels during the period of delayed ovulation with peaks of short duration at 15u0 h on the day of second treatment and the day prior to ovulation. Plasma levels of these 5 hormones during the first 4 days of gestation were not altered by delayed ovulation.