Orexins A and B are hypothalamic peptides which are derived from the proteolytic cleavage of prepro-orexin and act via two subtypes of receptors, named OX1-R (that almost exclusively binds orexin-A) and OX2-R (nonselective for both orexins). Several lines of evidence show that other neuropeptides, which like orexins are involved in the central control of energy homeostasis (e.g. leptin and ghrelin), may play a role in the regulation of bone metabolism, acting via autocrine-paracrine or endocrine routes. Therefore, we studied by reverse transcription-polymerase chain reaction (RT-PCR) the expression of the orexin system in rat calvarial osteoblast-like (ROB) cells, whose osteoblastic lineage was immunocytochemically demonstrated by their osteonectin and collagen-1alpha content at day 14 of culture. Conventional PCR detected the mRNA expression of OX1-R, but not OX2-R and prepro-orexin in ROB cells at days 2, 7 and 21 of culture. Semiquantitative real time-PCR evidenced a gradual down-regulation of OX1-R mRNA in relation to the duration of culture. This novel finding suggests that rat osteoblasts could be a target for circulating orexin-A, especially during their early stages of differentiation into mature osteoblasts.