In order to investigate the effects of long-term exercise training on brain endorphin systems, and the latter''s possible effects on the hypothalamic-pituitary-gonadal axis, female Wistar rats were subjected to daily treadmill running. A sedentary control group was also employed. After 8 wk of training, and just prior to sacrifice, 1/2 of each group received a final fatiguing bout of exercise. Thus, the final 4 groups consisted of a trained-fatigued (TF), trained-nonfatigued (TN), control-fatigued (CF), and control-nonfatigued (CN) group. Regional brain levels of .beta.-endorphin (.beta.E), methionine enkephalin and leucine enkephalin (LE) were assayed with independent RIA [radioimmunoassay] from the nucleus accumbens, cortex, caudate-putamen, septum, amygdala, anterior and posterior hypothalamus, substantia nigra and ventral tegmentum. Diestrus serum levels of luteinizing hormone(LH), FSH and prolactin (PRL) were also determined. Fatiguing resulted in a decrease in serum LH levels as well as an increase in .beta.E content in the nucleus accumbens, and LE content in the ventral tegmentum. TF animals exhibited less LE in the amygdala than the TN rats. These changes in brain endorphins may indicate an acute, fatigue-running modulation of the hypothalamic-pituitary-gonadal axis.