Objective To observe the effects of high fat diet and 6-week voluntary wheel running on the structure of hypothalamus, pituitary and testis and sex hormone levels in pubertal male rats.Methods Forty 3-week-old male Sprague-Dawley rats were randomly divided into following groups: the control group fed with normal diet (C), training group fed with normal diet (CE), control group fed with high fat diet (D), and training group fed with high fat diet (DE), 10 rats in each group. The groups C and D were bred for 8 weeks freely, and the groups CE and DE were assigned to have voluntary wheel running twice/d, 1 h/time, 5 d/w, for 6 weeks after a 2-week adaptive feeding. Eight weeks later, blood sample was collected to detect the serum T, E₂, FSH and LH, and the E₂ and T of testis were also detected. The histology of hypothalamic, pituitary, and testis tissues was observed by light microscopy.Results (l) Compared with the group C, the group D had significantly decreased levels of serum T and E₂ and testicular T (P<0.05), and significantly increased serum E₂, FSH and LH levels (P<0.05). In the group D, vacuolar lipid droplets were increased in the hypothalamus, eosinophils and basophils were reduced in the pituitary, and the area of seminiferous tubules, percentage of sperm cells, and quantity of Leygid cells were significantly decreased in the testis. (2) Compared with the group D, the serum T and testicular T concentrations were increased, but the serum E₂, FSH and LH were decreased significantly (P<0.05) in the group DE, and vacuolar lipid drops were increased in the hypothalamus, eosinophil cells were increased in the pituitary gland, and the area of seminiferous tubules was increased but not significantly in the testis. Conclusions 6-week voluntary wheel exercise can improve the high fat diet-induced abnormal secretion of sex hormones, but not effectively improve the histological changes in hypothalamus, pituitary gland and testis.