Thyroid hormones play an important role in the regulation of energy balance, sleep and emotional behaviors. Neuropeptide S (NPS) is a recently discovered neuropeptide, regulating feeding, sleep and anxiety. Here, we examined the effect of hyperthyroidism on the gene and protein expression of neuropeptide S and its receptor (NPS-R) in the hypothalamus, brainstem and amygdala of rats. Our results showed that the expression of NPS and NPS-R was differentially modulated by hyperthyroidism in the rat brain. NPS and NPS-R mRNA and protein levels were decreased in the hypothalamus of hyperthyroid rats. Conversely NPS-R expression was highly increased in the brainstem and NPS and NPS-R expression were unchanged in the amygdala of these rats. These data suggest that changes in anxiety and food intake patterns observed in hyperthyroidism could be associated with changes in the expression of NPS and NPS-R. Thus, the NPS/NPS-R system may be involved in several hyperthyroidism-associated comorbidities.