We measured luteinizing hormone (LH) and follicle stimulating hormone (FSH) by immunofluorometric assays and alpha-inhibin by radioimmunoassay in serum sampled every 10 min throughout the night (2100-0500 h) from 44 normal girls. Mean overnight LH values rose log-linearly from a mean of 0.2 IU/l in prepubertal girls to 3.0 IU/l in late pubertal girls. Log2 mean overnight FSH rose rapidly through early puberty and then remained constant; mean FSH rose from 1.0 IU/l in prepubertal girls to approximately 2.8 IU/l in Tanner III-V girls. Mean overnight inhibin increased through puberty, rising from 151 ng/l in prepubertal girls to 432 ng/l in fully pubescent girls. Within each of the first three Tanner stages, LH differed approximately 100-fold between the smallest and largest mean concentrations but differed <10-fold within stages IV or V. Such within-pubertal stage variability was less pronounced for FSH, which differed approximately 16-fold among Tanner I subjects and 4-10-fold at later stages, and for inhibin, which varied approximately 4-fold within each Tanner stage. The frequency of LH pulses during overnight sampling increased significantly during puberty, but the frequency of FSH and inhibin pulses remained constant. We compared the results from girls to those from 50 normal boys [Manasco et al. (1995) J. Clin. Endocrinol. Metab., 80, 20462052]. At each pubertal stage, girls had approximately the same mean overnight LH values as boys; girls had higher mean overnight FSH, particularly during Tanner stages II-IV; and boys had mean overnight alpha-inhibin immunoreactivity approximately 1.5 times that of girls at each pubertal stage. Still, hormone concentrations for individuals of both sexes intergraded at each pubertal stage.