To delineate the contributions of genetic and environmental factors in the regulation of human prolactin (PRL) secretion, the 24-h profile of plasma PRL was obtained at 15-min intervals in 10 pairs of monozygotic and 10 pairs of dizygotic twins. Sleep was monitored polygraphically. PRL secretory rates were derived from plasma concentrations by deconvolution. Diurnal (24-h) variations were quantified by a regression curve to define nadir, acrophase, and amplitude. Pulses of PRL secretion were identified using a computerized algorithm. A procedure specifically developed for twin studies was used to partition the variance into genetic and environmental contributions. Significant genetic effects were identified for daytime PRL concentrations, rhythm amplitude, and overall waveshape of the daily PRL profile. In contrast, environmental effects were dominant for mean concentrations during sleep, total secretory output during sleep, overall 24-h concentrations, and total 24-h secretion. However, when interindividual variations in sleep fragmentation were taken into account, the estimates of genetic variance for PRL concentrations and secretion during sleep approached statistical significance. Significant genetic influences were identified for slow-wave sleep (SWS). Because SWS is associated with increased nocturnal PRL secretion, it is possible that genetic effects on PRL secretion during sleep reflect genetic influences on SWS. In conclusion, genetic factors determine partially both the basal daytime concentrations of PRL and the temporal organization of PRL secretion over the 24-h cycle in normal young men.