Dexamethasone induces sleep disruption in male mice and is associated with hyperactivation of orexin neurons

Steroid therapy is widely used, but it frequently causes severe sleep disorders as a side effect. The underlying neurobiological mechanisms remain poorly understood, leading to a lack of clear evidence-based empirical drug selection. The aims of this study are: (1) to establish a mouse model of steroid-induced sleep disorder evoked by repeated dexamethasone administration, (2) to investigate the role of the orexin system in this disorder using neurophysiological techniques, and (3) to compare the efficacy of a benzodiazepine agonist (brotizolam) and a dual orexin receptor antagonist (suvorexant) in the model. Male C57BL/6J mice received intraperitoneal injections of dexamethasone (30 mg/kg) for five days before sleep architecture assessment by electroencephalography and electromyography. Orexin neuron activity was analyzed using fiber photometry in mice (Orexin-tTA) expressing GCaMP6 specifically in orexin neurons. Chronic dexamethasone administration in mice significantly increased wakefulness and reduced non-rapid eye movement sleep during the light-phase sleep period, and mimicked key features of clinical sleep disorders. Importantly, in vivo fiber photometry provided the first direct evidence that these states are driven by hyperactivity of orexin neurons during the arousal-promoting transition phase. Both brotizolam and suvorexant effectively reversed dexamethasone-induced sleep disruption and normalized sleep architecture. These findings indicate that steroid-induced sleep disturbance is mediated by hyperactivity of the orexinergic system. Furthermore, we show that both a benzodiazepine and a targeted orexin antagonist are effective in this model, providing a preclinical evidence base for the pharmacological management of this common and severe side effect.