The dual orexin receptor antagonist almorexant induces sleep and decreases orexin-induced locomotion by blocking orexin 2 receptors.

STUDY OBJECTIVES: Orexin peptides activate orexin 1 and orexin 2 receptors (OX(1)R and OX(2)R), regulate locomotion and sleep-wake. The dual OX(1)R/OX(2)R antagonist almorexant reduces activity and promotes sleep in multiple species, including man. The relative contributions of the two receptors in locomotion and sleep/wake regulation were investigated in mice. DESIGN: Mice lacking orexin receptors were used to determine the contribution of OX(1)R and OX(2)R to orexin A-induced locomotion and to almorexant-induced sleep. SETTING: N/A. PATIENTS OR PARTICIPANTS: C57BL/6J mice and OX(1)R(+/+), OX(1)R(-/-), OX(2)R(+/+), OX(2)R(-/-) and OX(1)R(-/-)/OX(2)R(-/-) mice. INTERVENTIONS: Intracerebroventricular orexin A; oral dosing of almorexant. MEASUREMENTS AND RESULTS: Almorexant attenuated orexin A-induced locomotion. As in other species, almorexant dose-dependently increased rapid eye movement sleep (REM) and nonREM sleep in mice. Almorexant and orexin A were ineffective in OX(1)R(-/-)/OX(2)R(-/-) mice. Both orexin A-induced locomotion and sleep induction by almorexant were absent in OX(2)R(-/-) mice. Interestingly, almorexant did not induce cataplexy in wild-type mice under conditions where cataplexy was seen in mice lacking orexins and in OX(1)R(-/-)/OX(2)R(-/-) mice. Almorexant dissociates very slowly from OX(2)R as measured functionally and in radioligand binding. Under non equilibrium conditions in vitro, almorexant was a dual antagonist whereas at equilibrium, almorexant became OX(2)R selective. CONCLUSIONS: In vivo, almorexant specifically inhibits the actions of orexin A. The two known orexin receptors mediate sleep induction by almorexant and orexin A-induced locomotion. However, OX(2)R activation mediates locomotion induction by orexin A and antagonism of OX(2)R is sufficient to promote sleep in mice.

Neuropeptide S receptor deficiency modulates spontaneous locomotor activity and the acoustic startle response.

The present study investigated the phenotype of heterozygous and homozygous neuropeptide S receptor (Npsr) deficient C57BL/6 mice in NPS- and cocaine induced hyperactivity, spontaneous and reactive locomotor activity, elevated plus maze, conditioned fear, and prepulse inhibition of the acoustic startle response. In Npsr-deficient mice, a strong reduction of spontaneous locomotor activity and of the startle magnitude was observed; heterozygous mice had an intermediate phenotype. In the other experiments, Npsr deficiency leads to no or only a very modest phenotype. These results support an important role of neuropeptide S in regulating locomotor activity.

Intra-amygdala injections of neuropeptide S block fear-potentiated startle.

Injections of neuropeptide S (NPS) into the lateral ventricle induce a strong hyperactivity. Since most behavioral paradigms are dependent of spontaneous locomotor activity, this makes it difficult to interpret the role of NPS in such paradigms. The aim of the present experiment was to investigate the effects of NPS in fear-potentiated startle, a behavioral fear paradigm which we believe is less sensitive to general changes in locomotor activity. Furthermore, NPS was directly injected into the amygdala, the central site of the neural fear circuitry. Our data shows that intra-amygdala NPS injections dose-dependently block the expression of conditioned fear and that this effect is independent of NPS effects on locomotor activity. This strongly supports a crucial role of amygdaloid NPS in conditioned fear.

Fear-reducing effects of intra-amygdala neuropeptide Y infusion in animal models of conditioned fear: an NPY Y1 receptor independent effect.

RATIONALE: Neuropeptide Y (NPY) and its receptors are densely localized in brain regions involved in the mediation and modulation of fear, including the amygdala. Several studies showed that central NPY is involved in the modulation of fear and anxiety. OBJECTIVES: In the present study, we investigated (1) whether intra-amygdala injections of NPY affect the expression of conditioned fear and (2) whether NPY Y1 receptors (Y1R) mediates the effects of these intra-amygdaloid NPY injections. RESULTS: Intra-amygdala NPY injections robustly decreased the expression of conditioned fear measured by conditioned freezing and fear-potentiated startle. These NPY effects were not mimicked by intra-amygdala injections of the Y1R agonists Y-28 or Y-36, and co-infusion of the Y1R antagonist BIBO 3304 did not block the NPY effects. Furthermore, we tested Y1R-deficient mice in conditioned freezing and found no differences between wild type and mutant littermates. Finally, we injected NPY into the amygdala of Y1R-deficient mice. Y1R deficiency had no effect on the fear-reducing effects of intra-amygdala NPY. CONCLUSIONS: These data show an important role of the transmitter NPY within the amygdala for the expression of conditioned fear. Y1R do not appear to be involved in the mediation of the observed intra-amygdala NPY effects suggesting that these effects are mediated via other NPY receptors.