Serotonergic Regulation of the Orexin/Hypocretin Neurons through the 5-HT1A Receptor

Both orexin and serotonin (5-HT) have important roles in the regulation of sleep-wakefulness, as well as in feeding behavior. We examined the effects of 5-HT on orexin/hypocretin neurons, using hypothalamic slices prepared from orexin/enhanced green fluorescent protein (EGFP) transgenic mice in which EGFP is expressed exclusively in orexin neurons. Patch-clamp recording from EGFP-expressing cells showed that 5-HT hyperpolarized all orexin neurons in a concentration-dependent manner. The response was inhibited by the 5-HT1A receptor antagonist WAY100635. A 5-HT1A receptor agonist, 8-hydroxy-2-(dl-N-propyl-amino)tetralin, also evoked hyperpolarization on orexin neurons with potency comparable with 5-HT. A low concentration of Ba2+ (30 μm) inhibited 5-HT-induced hyperpolarization. Single-channel recording revealed that the conductance of 5-HT-induced channel activity was 33.8 pS, which is in good agreement with that of the G-protein-coupled inward rectifier potassium channel (GIRK). Moreover, 5-HT1A receptor-like immunoreactivity was observed on orexin neurons, and 5-HT transporter immunoreactive nerve endings are in close apposition to orexin neurons. Intracerebroventricular injection of the 5-HT1A receptor-selective antagonist WAY100635 (100 ng) increased locomotor activity during the latter half of dark phase in wild-type mice but not in orexin/ataxin-3 mice in which orexin neurons are specifically ablated, suggesting that activation of orexin neurons is necessary for the WAY100635-induced increase in locomotor activity. These results indicate that 5-HT hyperpolarizes orexin neurons through the 5-HT1A receptor and subsequent activation of the GIRK and that this inhibitory serotonergic input to the orexin neurons is likely to be important for the physiological regulation of this neuropeptide system.

[1]  F E Bloom,et al.  The hypocretins: hypothalamus-specific peptides with neuroexcitatory activity. , 1998, Proceedings of the National Academy of Sciences of the United States of America.

[2]  R. McCarley,et al.  Behavioral State Control through Differential Serotonergic Inhibition in the Mesopontine Cholinergic Nuclei: A Simultaneous Unit Recording and Microdialysis Study , 1998, The Journal of Neuroscience.

[3]  J. Leysen,et al.  Functional role of 5-HT2 receptors in the regulation of sleep and wakefulness in the rat , 2004, Psychopharmacology.

[4]  L. P. Morin,et al.  Reciprocal serotonergic connections between the hamster median and dorsal raphe nuclei , 2003, Brain Research.

[5]  R. Hen,et al.  Key Role of 5-HT1B Receptors in the Regulation of Paradoxical Sleep as Evidenced in 5-HT1B Knock-Out Mice , 1999, The Journal of Neuroscience.

[6]  P. Saxena,et al.  5-HT1-like receptor agonists enhance wakefulness , 1992, Neuropharmacology.

[7]  B. Jacobs,et al.  Raphe unit activity during REM sleep in normal cats and in pontine lesioned cats displaying REM sleep without atonia , 1981, Brain Research.

[8]  R. Ursin Serotonin and sleep. , 2002, Sleep medicine reviews.

[9]  Donghee Kim,et al.  Properties and Modulation of the G Protein‐Coupled K+ Channel in Rat Cerebellar Granule Neurons: ATP Versus Phosphatidylinositol 4,5‐Bisphosphate , 2003, The Journal of physiology.

[10]  A. Yamanaka,et al.  Orexins activate histaminergic neurons via the orexin 2 receptor. , 2002, Biochemical and biophysical research communications.

[11]  J. Siegel,et al.  Activity of dorsal raphe cells across the sleep–waking cycle and during cataplexy in narcoleptic dogs , 2004, The Journal of physiology.

[12]  T. Sakurai,et al.  Orexin-induced hyperlocomotion and stereotypy are mediated by the dopaminergic system 1 1 Published on the World Wide Web on 27 June 2000. , 2000, Brain Research.

[13]  J. Hobson,et al.  Forced activity alters sleep cycle periodicity and dorsal raphe discharge rhythm. , 1984, The American journal of physiology.

[14]  C. Idzikowski,et al.  5-Hydroxytryptamine-2 antagonist increases human slow wave sleep , 1986, Brain Research.

[15]  E. Lainey,et al.  Effects of ipsapirone, a 5‐HT1A agonist, on sleep/wakefulness cycles: probable post‐synaptic action , 1993, Journal of sleep research.

[16]  A. Yamanaka,et al.  Regulation of orexin neurons by the monoaminergic and cholinergic systems , 2004 .

[17]  Takeshi Sakurai,et al.  Distribution of orexin neurons in the adult rat brain 1 Published on the World Wide Web on 17 March 1999. 1 , 1999, Brain Research.

[18]  B. Jacobs,et al.  Structure and function of the brain serotonin system. , 1992, Physiological reviews.

[19]  C. Sinton,et al.  Electrophysiological evidence for a functional differentiation between subtypes of the 5-HT1 receptor. , 1988, European journal of pharmacology.

[20]  Takeshi Sakurai,et al.  Hypocretin/Orexin Excites Hypocretin Neurons via a Local Glutamate Neuron—A Potential Mechanism for Orchestrating the Hypothalamic Arousal System , 2002, Neuron.

[21]  S. Han,et al.  5-HT1A receptor-mediated activation of G-protein-gated inwardly rectifying K+ current in rat periaqueductal gray neurons , 2001, Neuropharmacology.

[22]  D. Bajic,et al.  Two different inward rectifier K+ channels are effectors for transmitter-induced slow excitation in brain neurons , 2002, Proceedings of the National Academy of Sciences of the United States of America.

[23]  T. Akasu,et al.  5-Hydroxytryptamine-induced outward currents mediated via 5-HT1A receptors in neurons of the rat dorsolateral septal nucleus , 2000, Neuroscience Research.

[24]  Sebastiaan Overeem,et al.  Hypocretin (orexin) deficiency in human narcolepsy , 2000, The Lancet.

[25]  K. Kunii,et al.  Involvement of the serotonergic system in orexin-induced behavioral alterations in rats , 2002, Regulatory Peptides.

[26]  S. Watson,et al.  Comparative anatomical distribution of 5-HT1A receptor mRNA and 5-HT1A binding in rat brain — a combined in situ hybridisation/in vitro receptor autoradiographic study , 1991, Brain Research.

[27]  M. I. Smith,et al.  Orexin A activates locus coeruleus cell firing and increases arousal in the rat. , 1999, Proceedings of the National Academy of Sciences of the United States of America.

[28]  L. Descarries,et al.  Somatodendritic localization of 5‐HT1A and preterminal axonal localization of 5‐HT1B serotonin receptors in adult rat brain , 2000, The Journal of comparative neurology.

[29]  S. Carr,et al.  Orexins and Orexin Receptors: A Family of Hypothalamic Neuropeptides and G Protein-Coupled Receptors that Regulate Feeding Behavior , 1998, Cell.

[30]  D. Clapham,et al.  Recombinant G-protein beta gamma-subunits activate the muscarinic-gated atrial potassium channel. , 1994, Nature.

[31]  A. N. van den Pol,et al.  Neurons Containing Hypocretin (Orexin) Project to Multiple Neuronal Systems , 1998, The Journal of Neuroscience.

[32]  L. Dinardo,et al.  Midbrain Raphe Modulation of Nonphotic Circadian Clock Resetting and 5-HT Release in the Mammalian Suprachiasmatic Nucleus , 2003, The Journal of Neuroscience.

[33]  Jun Lu,et al.  Afferents to the Ventrolateral Preoptic Nucleus , 2002, The Journal of Neuroscience.

[34]  H. Lester,et al.  Evidence that neuronal G-protein-gated inwardly rectifying K+ channels are activated by G beta gamma subunits and function as heteromultimers. , 1995, Proceedings of the National Academy of Sciences of the United States of America.

[35]  Jon T. Willie,et al.  Genetic Ablation of Orexin Neurons in Mice Results in Narcolepsy, Hypophagia, and Obesity , 2001, Neuron.

[36]  G. Aston-Jones,et al.  Hypocretin (orexin) activation and synaptic innervation of the locus coeruleus noradrenergic system , 1999, The Journal of comparative neurology.

[37]  T. Yakushiji,et al.  Characterization of the K+ current mediated by 5-HT1A receptor in the acutely dissociated rat dorsal raphe neurons , 1997, Brain Research.

[38]  Ronald M. Harper,et al.  Dorsal raphe neurons: depression of firing during sleep in cats , 1976, Brain Research.

[39]  J. Adrien Neurobiological bases for the relation between sleep and depression. , 2002, Sleep medicine reviews.

[40]  N. Akaike,et al.  α2‐Adrenoceptor‐mediated potassium currents in acutely dissociated rat locus coeruleus neurones , 1998, The Journal of physiology.

[41]  M. Stryker,et al.  Sleep and Sleep Homeostasis in Mice Lacking the 5-HT2c Receptor , 2002, Neuropsychopharmacology.

[42]  M. Jouvet Sleep and Serotonin: An Unfinished Story , 1999, Neuropsychopharmacology.

[43]  N. Vrang,et al.  Neuronal projections from the mesencephalic raphe nuclear complex to the suprachiasmatic nucleus and the deep pineal gland of the golden hamster (Mesocricetus auratus) , 1998, The Journal of comparative neurology.

[44]  Takeshi Sakurai,et al.  Hypothalamic Orexin Neurons Regulate Arousal According to Energy Balance in Mice , 2003, Neuron.

[45]  H. Haas,et al.  Convergent Excitation of Dorsal Raphe Serotonin Neurons by Multiple Arousal Systems (Orexin/Hypocretin, Histamine and Noradrenaline) , 2002, The Journal of Neuroscience.

[46]  T. Petrov,et al.  The hypothalamic paraventricular and lateral parabrachial nuclei receive collaterals from raphe nucleus neurons: A combined double retrograde and immunocytochemical study , 1992, The Journal of comparative neurology.

[47]  Ming-Fung Wu,et al.  Release of Hypocretin (Orexin) during Waking and Sleep States , 2002, The Journal of Neuroscience.

[48]  Sebastiaan Overeem,et al.  A mutation in a case of early onset narcolepsy and a generalized absence of hypocretin peptides in human narcoleptic brains , 2000, Nature Medicine.

[49]  Mustapha Riad,et al.  Immunocytochemical localization of serotonin1A receptors in the rat central nervous system , 1996, The Journal of comparative neurology.

[50]  R. Hen,et al.  Involvement of 5-HT1A Receptors in Homeostatic and Stress-Induced Adaptive Regulations of Paradoxical Sleep: Studies in 5-HT1A Knock-Out Mice , 2002, The Journal of Neuroscience.

[51]  A. N. van den Pol,et al.  Hypocretins (Orexins) Regulate Serotonin Neurons in the Dorsal Raphe Nucleus by Excitatory Direct and Inhibitory Indirect Actions , 2002, The Journal of Neuroscience.

[52]  M. Jouvet,et al.  The role of monoamines and acetylcholine-containing neurons in the regulation of the sleep-waking cycle. , 1972, Ergebnisse der Physiologie, biologischen Chemie und experimentellen Pharmakologie.

[53]  D. Clapham,et al.  Recombinant G-protein βγ-subunits activate the muscarinic-gated atrial potassium channel , 1994, Nature.

[54]  Jon T. Willie,et al.  Narcolepsy in orexin Knockout Mice Molecular Genetics of Sleep Regulation , 1999, Cell.

[55]  Barry L. Jacobs,et al.  Raphe unit activity in freely moving cats: Correlation with level of behavioral arousal , 1979, Brain Research.

[56]  D. Rainnie,et al.  Microdialysis perfusion of 8-hydroxy-2-(di-n-propylamino)tetralin (8-OH- DPAT) in the dorsal raphe nucleus decreases serotonin release and increases rapid eye movement sleep in the freely moving cat , 1996, The Journal of neuroscience : the official journal of the Society for Neuroscience.

[57]  C. Saper,et al.  Differential expression of orexin receptors 1 and 2 in the rat brain , 2001, The Journal of comparative neurology.