title: Orexin neurons receive glycinergic innervations.
abstract: Glycine, a nonessential amino-acid that acts as an inhibitory neurotransmitter in the central nervous system, is currently used as a dietary supplement to improve the quality of sleep, but its mechanism of action is poorly understood. We confirmed the effects of glycine on sleep/wakefulness behavior in mice when administered peripherally. Glycine administration increased non-rapid eye movement (NREM) sleep time and decreased the amount and mean episode duration of wakefulness when administered in the dark period. Since peripheral administration of glycine induced fragmentation of sleep/wakefulness states, which is a characteristic of orexin deficiency, we examined the effects of glycine on orexin neurons. The number of Fos-positive orexin neurons markedly decreased after intraperitoneal administration of glycine to mice. To examine whether glycine acts directly on orexin neurons, we examined the effects of glycine on orexin neurons by patch-clamp electrophysiology. Glycine directly induced hyperpolarization and cessation of firing of orexin neurons. These responses were inhibited by a specific glycine receptor antagonist, strychnine. Triple-labeling immunofluorescent analysis showed close apposition of glycine transporter 2 (GlyT2)-immunoreactive glycinergic fibers onto orexin-immunoreactive neurons. Immunoelectron microscopic analysis revealed that GlyT2-immunoreactive terminals made symmetrical synaptic contacts with somata and dendrites of orexin neurons. Double-labeling immunoelectron microscopy demonstrated that glycine receptor alpha subunits were localized in the postsynaptic membrane of symmetrical inhibitory synapses on orexin neurons. Considering the importance of glycinergic regulation during REM sleep, our observations suggest that glycine injection might affect the activity of orexin neurons, and that glycinergic inhibition of orexin neurons might play a role in physiological sleep regulation.
authors.affiliation: Department of Molecular Neuroscience and Integrative Physiology, Faculty of Medicine, Kanazawa University, Kanazawa, Ishikawa, Japan.
authors.forename: Mari
authors.initials: M
authors.lastname: Hondo
authors.forename: Naoki
authors.initials: N
authors.lastname: Furutani
authors.forename: Miwako
authors.initials: M
authors.lastname: Yamasaki
authors.forename: Masahiko
authors.initials: M
authors.lastname: Watanabe
authors.forename: Takeshi
authors.initials: T
authors.lastname: Sakurai
mesh_terms: D000818:Animals; D004594:Electrophysiology; D005998:Glycine / Q000008:administration & dosage / Q000494:pharmacology*; D018684:Glycine Agents / Q000008:administration & dosage / Q000494:pharmacology*; D050487:Glycine Plasma Membrane Transport Proteins; D007124:Immunoenzyme Techniques; D007274:Injections, Intraperitoneal; D047908:Intracellular Signaling Peptides and Proteins; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D009433:Neural Inhibition; D009474:Neurons / Q000166:cytology / Q000187:drug effects*; D009479:Neuropeptides; D000068797:Orexins; D018009:Receptors, Glycine / Q000378:metabolism; D012890:Sleep / Q000187:drug effects / Q000502:physiology*; D014851:Wakefulness / Q000187:drug effects / Q000502:physiology*
chemical_list: D018684:Glycine Agents; D050487:Glycine Plasma Membrane Transport Proteins; D047908:Intracellular Signaling Peptides and Proteins; D009479:Neuropeptides; D000068797:Orexins; D018009:Receptors, Glycine; D005998:Glycine