We analyzed the effect of chronic (4 weeks) ethanol feeding on 24-h variation of pituitary-testicular function in peripubertal male Wistar rats by measuring circulating concentrations of prolactin, follicle-stimulating hormone, luteinizing hormone, testosterone, and thyrotropin. Animals were maintained under a 12-h light: 12-h dark photoperiod and received a liquid diet for 4 weeks, starting on day 35 of life. The ethanol-fed group received a diet similar to that provided to control animals, except that maltose was replaced isocalorically with ethanol. Ethanol replacement provided 36% of the total caloric content of the diet. Rats were killed at one of six times around the clock, beginning at zeitgeber time (ZT) 1 (ZT 0 = lights on). In ethanol-fed rats globally, secretion of prolactin was augmented, whereas secretion of follicle-stimulating hormone, luteinizing hormone, testosterone, and thyrotropin was decreased. Significant changes in the 24-h secretory pattern of circulating hormones occurred in rats receiving ethanol, including the appearance of two peaks (at ZT 1 and ZT 9), rather than one peak, of follicle-stimulating hormone during the inactive phase of the daily cycle, suppression of the maximum plasma luteinizing hormone concentration during the first part of the inactive phase, and appearance of a second peak of testosterone and prolactin during the second part of the inactive phase (at ZT 5 and ZT 9, respectively) and of a second peak of plasma thyrotropin during the first part of the active phase (at ZT 13). The significant positive correlation between testosterone and individual luteinizing hormone and prolactin concentrations in control animals was no longer observed after ethanol administration. Chronic ethanol administration presumably affects the endogenous clock that modulates the circadian variation of the pituitary-gonadal axis and thyrotropin release in growing male rats.