Introduction Loss of orexin neurons is associated with narcolepsy in the human, a sleep disorder characterized by excessive daytime sleepiness and cataplexy. Mice lacking orexin peptides, as well as those lacking orexin receptors (OX1R−/−; OX2R−/− mice), display a phenotype similar to narcolepsy, highlighting a critical role of orexin signaling in the maintenance of wakefulness. However, precise neural mechanisms downstream to orexin neurons have remained uncertain. Materials and methods We generated recombinant adeno-associated viruses (AAV) to express either OX1R or OX2R fused to EGFP and stereotaxically microinjected into the of OX1R−/−; OX2R−/− mice, then recorded EEG/EMG to score sleep/wakefulness states. The subtype of orexin receptors expressed was determined according to expression of endogenous orexin receptors in wild-type mice. Results We found that targeted restoration of orexin receptor expressions in noradrenergic neurons of the locus coeruleus and in serotonergic neurons of the dorsal raphe in OX1R−/−; OX2R−/− mice differentially inhibited pathological fragmentation of wakefulness (i.e., sleepiness) and direct transitions from wakefulness to REM sleep (cataplexy-like episode), respectively. Furthermore, pharmacogenetic activation of these neurons using DREADD technology significantly ameliorated narcolepsy of mice lacking orexin neurons. Conclusion These results suggest that orexin neurons consolidate wakefulness and suppress cataplexy by activating locus coeruleus noradrenergic and dorsal raphe serotonergic neurons, respectively. Our success in improving narcoleptic symptoms by DREADD may lead to a novel type of gene therapy. Acknowledgements This study was supported in part by Grants-in-Aid for Scientific Research (B) and for Challenging Exploratory Research from the Ministry of Education, Culture, Sports, Science, and Technology (MEXT) of Japan (M.M.), by the Mochida Memorial Foundation for Medical and Pharmaceutical Research (M.M.), and by the Cabinet Office, Government of Japan through its _gFunding Program for Next Generation World-Leading Researchers_h (T.S.). We thank Dr. Karl Deisseroth for pAAV-DIO-hChR2 (H134R)-EYFP-WPRE-pA; Dr. Scott M. Sternson for pAAV-FLEX-rev- ChR2mCherry; Dr. Kwang-Soo Kim for the PRSx8 promoter; Dr. Masahiko Watanabe for the anti-GFP antibody; and Dr. Arun Srivastava for pACG-2- Y730F.