HyLine Brown laying hens at 30 weeks of age were treated twice daily with Fadrozole, a non-steroidal aromatase inhibitor (AI; 1 mg/kg body weight; i.m.) for six consecutive days; control hens received saline. Blood was collected every day 0.5 h after oviposition, i.e. just before AI treatment. Ovarian steroids: progesterone (P4), testosterone (T) and estradiol (E2), and iodothyronines: thyroxine (T4), triiodothyronine (T3) and reverse-triiodothyronine (rT3) were measured in blood plasma by radioimmunoassay methods. In AI-treated hens a gradual delay in oviposition time was observed. AI significantly decreased P4 and E2 levels, maximally by 43% on day 4 and by 74% on day 5, respectively, and elevated T level, maximally by 248% on day 4. Simultaneously, the increases in T4 and T3 levels with no changes in rT3 levels were observed. The maximal effect of AI on T4 and T3 levels was found on day 4 (60% increase) and day 5 (312% increase), respectively. Moreover, statistically significant, negative coefficient of correlation between E2 and T3 (r = -0.51), and positive coefficient of correlation between T and T3 (r = 0.42) in AI-treated hens were found. The results obtained indicate that in mature laying hens there is a strong relationship between ovarian steroids and thyroid hormones, and suppression of E2 synthesis not only disrupts ovarian function but also affects the activity of the thyroid gland and peripheral metabolism of thyroid hormones.