The aim of this study was to investigate thyroid hormone levels in horses stabled in two different locations on the island of Sicily. The study was carried out on a total of 72 clinically healthy Sanfratellano horses ranging in age from 5 to 9 years and weighing 585 ± 40 kg. The results showed higher thyroxine values (P < 0.02) in horses stabled in an endemic goitre area (group II) than those observed in horses in a non-endemic area (group I). Unexpectedly, the T4/T3 and the fT4/fT3 rations were both lower in group I than in group II. The percentages of fT4 to T4 and of fT3 to T3 were both higher in group I than the percentages for group II. On the basis of gender, comparison between the two groups showed higher T4 (P < 0.01) and fT4 levels (P < 0.001) in males, and lower fT3 (P < 0.001) and fT4 levels (P < 0.005) in females stabled in the goitre endemic area. On the basis of age, younger horses (<7 years old) showed the highest thyroid hormone levels in both groups. Results suggest a physiological adaptive response of the equine species to an endemic goitre environment. The possibility that hypothyroidism is present in these horses is thus excluded and is supported not only by the lack of clinical signs, but also by the rarity of cases previously reported.