Orexinergic (OX) neurons in the lateral hypothalamus (LH), perifornical area (PFA) and dorsomedial hypothalamus (DMH) play a role in the hypercapnic ventilatory response, presumably through direct inputs to central pattern generator sites and/or through interactions with other chemosensitive regions. OX neurons can produce and release orexins, excitatory neuropeptides involved in many functions, including physiological responses to changes in CO2/pH. Thus, in the present study, we tested the hypothesis that different nuclei (LH, PFA and DMH) where the orexinergic neurons are located, show distinct activation by CO2 during the light-dark cycle phases. For this purpose, we evaluated the Fos and OXA expression by immunohistochemistry to identify neurons that co-localize Fos+OXA in the LH, LPeF, MPeF and DMH in the light-inactive and dark-active phase in Wistar rats subjected to 3 hours of normocapnia or hypercapnia (7% CO2). Quantitative analyses of immunoreactive neurons show that hypercapnia caused an increase in the number of neurons expressing Fos in the LH, LPeF, MPeF and DMH in the light and dark phases. In addition, the number of Fos+OXA neurons increased in both phases of the diurnal cycle in the LPeF and DMH, whereas in the MPeF, this increase was observed exclusively in the light phase. Thus, we suggest that OX neurons are selectivity activated by hypercapnia throughout the diurnal cycle, reinforcing the differential role of nuclei in the hypothalamus during central chemosensitivity.