Artículo destacado

Drug News & Perspectives
Vol. 16, No. 10, 2003, p. 669-681
ISSN 0214-0934
Copyright 2003 Prous Science, S.A.
CCC: 0214-0934/2003
http://www.prous.com

LOOKING AHEAD

Development of drugs targeting the orexin system will open novel therapeutic avenues for the treatment of many neuropsychiatric and neuroendocrine disorders of eating, pain, mood, sleep and memory.

Drugs to Interfere with Orexins (Hypocretins)

by Oliver Selbach, Krister S. Eriksson and Helmut L. Haas

Summary

Orexins (hypocretins) are bioactive peptides linking arousal, appetite and neuroendocrine-autonomic control. Dysfunction of the orexin system is associated with narcolepsy-cataplexy. Here, we review drugs interfering with orexins directly such as novel selective orexin receptor agonists and antagonists, as well as drugs interfering with orexins indirectly such as those used for the treatment of narcolepsy-cataplexy, and pharmacological targets within the complex network of endogenous neurohumoral signals integrated and relayed by the orexin system. These include amines, acetylcholine, purines, GABA and glutamate, as well as nutritional-metabolic, circadian-photic, im-munological and neuroendocrine-peptidergic influences. Basic and clinical evaluation of drugs interfering with the orexin system will lead to a better understanding of the molecular prerequisites that control behavioral state, stress responses, energy homeo- stasis and survival, and yield therapeutic advances for the treatment of narcolepsy and other disorders of sleep, eating, mood and memory. © 2003 Prous Science. All rights reserved.

Orexins, also named hypocretins, were discovered in 1998 by two independent groups using reverse genetic (subtractive cDNA cloning)¹ and pharmacological techniques (orphan receptor screening).² Research on orexins gained further impact by the discovery of their etiological role in the human sleep disorder narcolepsy and animal models of that disease.^3-7 Growing numbers of research articles published on orexins reflect their recognition as central regulators linking metabolic and behavioral state, and their implication in many body functions, including arousal, appetite, energy homeostasis and neuroendocrine-autonomic control. A number of excellent reviews on the multiple facets of orexins and narcolepsy have been published.^6-20 Here, we focus on drugs used in the treatment of narcolepsy-cataplexy, selective orexin receptor antagonists and agonists, and putative drug targets within the complex array of neurohumoral signals and behavioral control systems associated with orexins. We refer to hypocretins as orexins throughout our review, since a final nomenclature has not evolved.

Structure and functions of the orexin system

The main source of endogenous orexins in the central nervous system are orexinergic neurons located perifornically in the lateral hypothalamus,^21-24 a prominent feeding and arousal center implicated in appetitive and defense behaviors.^25,26 Orexin neurons project widely through the entire neuraxis.^27-29 Particularly dense innervations with orexinergic fibers are found in the neuroendocrine and aminergic-cholinergic autonomic centers in the brainstem, midbrain and hypothalamus but also accumulate in infralimbic cortical and subcortical structures including the spinal cord. The distribution of orexin receptors is also widespread but differential and often complementary within^30,31 and even out- side^2,32 the central nervous system, suggesting complex functions and heterogeneity. Indeed, orexin 2 receptors (OX₂Rs) are found mainly in infra- limbic structures involved in arousal control and sleep-wake transitions,^6,7,9 whereas OX₁Rs are closely associated with feeding and energy homeostasis,^{8,13-16,20,24} REM-sleep transitions³³ and autonomic outflow.^17-19,34-37 Notably, the distribution of orexins and their receptors matches the central representations of the sympathetic nervous system and structures related to intero-, viscero- and nociception,^38,39 and stress.⁴⁰ Indeed, 50% of the putative command neurons integrating somatomotor and cardiosympathetic functions have been identified as orexinergic.⁴¹ Apart from orexin neurons in the lateral hypothalamus, specific OX-B-like immunoreactivity has recently been described in clustered neuronal populations located in the lateral division of the central nucleus of the amygdala and in the bed nucleus of the stria terminalis.⁴² This is reminiscent of the corticotropin-releasing hormone, with which the orexin system exhibits close functional relations along all components of the HPA axis.^{19,34,35,43-50}

First considered to be primarily involved in body-weight control and energy homeostasis,^{8,14-16,20,24} orexins are now recognized as central regulators of arousal and sleep architecture.^3,6,7,9 Growing evidence indicates that the orexin system orchestrates many autonomic and somatic body functions including arousal, appetite, energy homeostasis,^{8,14-16,20,24} neuroendo-crine-autonomic control,^{19,34,35,44-50} thermoregulation,^51-54 reproduction,^55,56 nociception,^57-60 and learning or memory.^60-65 As an integral part of the brain-gut axis and hypothalamo-adipo-insular energy homeostat^13,66,67 as well as the hypothalamo-pituitary-adrenal stress axis,^{19,34,35,43-50} orexins may translate intero-, viscero- and nociceptive information^38,39 into systemic stress or defense responses subserving energy homeostasis and survival.^19,26,40 Orexins have been considered to maintain systemic bistability,¹¹ stabilizing metabolic and behavioral state through facilitation of appropriate autonomic and motor pattern generation and selection,³⁹ for example, opposing homeostatic wake- sleep transitions and sleep propensity during times of energy depletion and demand,^24,33 such as during exploration of novel environments or social stress.²⁶ Orexins may thus set thresholds of arousal^9,24 and appetitive-incentive behaviors^8,68,69 according to metabolic state and environmental challenges including circadian, photic and social cues.^26,70-76

Molecular prerequisites of the orexin system

The orexin system consists of three phylogenetically conserved genes. One encodes for the precursor peptide prepro-orexin, a member of the secretin-PACAP-VIP-glucagon family of peptides (Fig. 1).^1,9,77 Two other separate genes encode for the G-protein-coupled orexin receptors OX₁R and OX₂R.^2,8 The biologically active peptides orexin A (OX-A) and B (OX-B) are C-terminally RFamidated peptides derived from alternate splicing (convertase) and posttranslational modifications (cleavage).⁷⁸ They exhibit slight bombesin-like sequence homology.^2,8 OX-A is a 33-amino acid peptide (3,562 kDa) with two sets of disulfide bonds, playing a key role in receptor activation. OX-A also has a longer half-life than OX-B (28 amino acids; 2,937 kDa, 46% homology to OX-A). The orexin receptors OX₁ and OX₂ exhibit considerable differences with respect to their agonist affinities,^2,8 signal transduction^12,79-83 and distribution throughout and even outside the nervous system.^2,30,32 OX₁Rs exhibit about 10- to 100-fold higher selectivity for OX-A than OX-B, while OX₂Rs bind both peptides with equal affinity.^12,79 Binding of orexins is rather specific with respect to other receptors exhibiting structur- al and functional homology.^80-83 However, both OX-B and neuropeptide FF, a recently identified pain modulatory peptide, also bind to a novel orphan G-protein-coupled receptor.⁸⁴ One explanation may be the generally low level of discrimination of neuropeptide FF receptors for the few RFamide peptides such as orexins, neuropeptide Y (NPY) or prolactin-releasing peptide that have been identified in mammals and are abundant in invertebrates.

Fig. 1. Molecular basis of the orexin system.

In vitro evidence from studies in heterologous expression systems indicate that OX₁Rs belong to the G_q/11-subclass of heteromeric G-proteins coupling to Ca²⁺- and/or PKC-dependent intracellular signal transduction pathways,^12,79 whereas OX₂Rs may also couple to G_i/o.^35,36,82,85 However, studies in intact brain tissues indicate a much greater complexity in the signal transduction of OXRs,¹² including direct coupling to nonselective cationic channels,^86-89 to transient receptor potential channels,⁹⁰ to electrogenic sodium calcium exchangers,^63,91 to inhibition of GIRK channels that have previously been activated by somatostatin, nociception or the mu opioid agonist DAMGO,⁸⁵ and to unusual Ca²⁺-dependent signaling pathways associated with activation of mitogen-activated protein kinase,^12,79 and/or thapsigargin- and cAMP-PKA-sensitive pathways.^34,92,93 Similar to other G-protein-coupled receptors, OX₁Rs undergo homologous and heterologous desensitization and resensitization via ligand-induced b-arrestin 1 trafficking.⁹⁴ Notably, hypersensitization of OX₁Rs by cannabinoid CB1 receptors through putative heterooligomerization has recently been documented in heterologous expression systems, suggesting a tight molecular and functional coupling of endocannabinoid and orexinergic signaling.⁹⁵

Inactivation of bioactive peptides typically involves nonspecific proteolytic cleavage via cell surface metalloendopeptidases located in the proximity of the peptide receptors. A specific inactivation system for orexins is unknown. Thus, inhibitors of dipeptidyl peptidases⁹⁶ or proteolysis-resistant peptide analogues similar to those already synthesized for melanin-concentrating hormone (MCH)⁹⁷ or glucagon family peptides⁹⁸ may provide the means to prolong and boost orexinergic signaling in vivo. Another important mechanism controlling the availability and efficacy of peptides in the brain, particularly those with central and peripheral activity, is the dynamic of the blood-brain barrier. OX-A but not OX-B enters the brain by simple diffusion via the blood-brain barrier.⁹⁹ In addition, the abundance of orexins and their receptors in the olfactory bulb and throughout all parts of the central olfactory system¹⁰⁰ suggests that transnasal access¹⁰¹ may provide an alternative route for drugs to interfere with orexins. Moreover, the presence of prepro-orexin mRNA in ependymal cells, neuroblastomas¹⁰² and other neural-crest derivates^32,103 suggests yet unidentified roles of orexins as paracrine factors and "gatekeepers" controlling the neuroendocrine crosstalk between the brain and the periphery, and potentially also plasticity and repair.¹⁰⁴

Orexin receptor agonists and antagonists

The pharmacological targeting of OXRs and the development of selective and brain penetrant OXR agonists or antagonists are still in their infancy. Several prodrugs that directly interfere with orexins have been tested in animals in vitro and in vivo (Fig. 2, Table I).

Fig. 2. Chemical structures of drugs that interfere with orexins.

Orexin agonists

OX-A and OX-B are natural agonists at OXRs. Three l-leucine residues at positions 11, 14 and 15 in OX-B are important for selectivity to OX₂R over OX₁R.^105,106 Thus, prototype-selective OX₂R agonists [Ala¹¹]OX-B (SB-668875) and [Ala¹¹, d-Leu¹⁵]OX-B, derived from sys- tematic l-alanine and l-amino acid replacement of OX-B, retain agonist activity but are about 100-fold and 400-fold more selective for OX₂R over OX₁R, respectively, similar to OX-B from Xenopus frog.¹⁰⁷ In heterologous expression systems, [Ala¹¹, d-Leu¹⁵]-OX-B mobilized [Ca²⁺]_i with significant selectivity of OX₂R (EC₅₀ = 0.13 nM) over OX₁R (EC₅₀ = 52 nM). In vivo, SB-668875, apart from promoting arousal, produced marked and dose-dependent increases in locomotor activity, indicating that OX₂R activation is required for orexin-mediated effects on motor performance.

Orexin antagonists

Although OX₁Rs are rather selective for OX-A, the relative potencies of OX-A and OX-B are probably not sufficient to pharmacologically dissect different OXR functions, particularly not in vivo. SB-334867 is the first and so far only available selective OX₁R antagonist among several prodrugs, tested systematically in a variety of behavioral interaction studies and in vitro assays in rodents^{14,16,59,104,108-111} (Fig. 2, Table I). The compound is a competitive agonist (OX₁ pK_B = 7.4; OX₂ pK_B = 5.7) exhibiting about 50-fold selectivity for OX₁R over OX₂R and a range of other G-protein-coupled receptors. SB-334867 is bioavailable following intraperitoneal administration and is also brain penetrant, with brain levels peaking after 30 minutes and declining fairly rapidly thereafter. Another orally active and 100-fold more selective OX₁R antagonist, SB-408124 (OX1 pK_B = 7.8; OX₂ pK_B = 5.8), has a similar pharmacological profile to SB-334867 but much faster clearance from the brain, thus limiting its use in vivo.

Taken together, studies with SB-334867 in vivo provide strong evidence that OX₁Rs are required for the effects of orexins on feeding and grooming,¹⁶ nociception,^59,111 arousal and REM sleep.¹¹⁰ In addition, the effects of SB-334867 in some studies suggest that, at least under certain conditions, an endogenous orexinergic tone may be particularly important for appetite regulation and antinociception. Selective OX₁R antagonists exhibit significant antiobesity and antidiabetic effects in leptin-deficient mice⁵² and prohyperalgesic properties under inflammatory conditions.^59,111 Conversely, selective OX₁R agonists may stimulate appetite (e.g., in cachexia, anorexia) and may be analgesic under inflammatory conditions, including (but probably not restricted to) arthritis, pulmonary or bowel diseases, and migraine.¹¹²

At present, no data with respect to specific OX₂R antagonists are available. Evidence from OX₂R knockout animals suggests that OX₂Rs in close association with histaminergic functions^33,73,91,113 may be required for the maintenance of wakefulness and the gating of slow-wave sleep. Selective OX₂R agonists are thus especially warranted for the treatment of hypersomnia in narcolepsy but may also apply to other conditions in which wake-promoting actions need to be dissociated from drug effects on energy metabolism and cardiovascular functions, for example, in anesthesiology or resuscitation. Moreover, the effect of SB-668875 on locomotor activity suggests that selective OX₂R agonists and antagonists, apart from their putative role as wake-promoting and hypnotic drugs in sleep disorders, may also add to the therapy of hypo- and hyper-kinetic movement disorders.

Pharmacotherapy of narcolepsy and cataplexy

Drug targets interfering with the orexin system indirectly may be deduced from the pharmacotherapy of human narcolepsy^4,6,7 and from animal models of this disease.^{2,3,24,33,114-116} Narcolepsy is a disabling neurological disorder characterized by excessive daytime sleepiness and abnormal REM sleep control including cataplexy, sleep paralysis and hypnagogic hallucinations. Most cases of late-onset human narcolepsy, affecting about 1 in 2000 individuals, are thought to result from a selective destruction of orexinergic neurons in the lateral hypothalamus.^117,118 Canine narcolepsy, however, is caused by gene mutations leading to dysfunction of OX₂ receptors.^3,115

Excessive daytime sleepiness in narcolepsy is commonly treated with sympathomimetic psychostimulants such as methamphetamine, methylphenidate, pemoline, mazindol or the novel wake-promoting drug modafinil.^3,7 Other therapeutic options include MAO inhibitors such as phenelzine, pargyline, tranylcypromine and the MAO-B selective inhibitor selegiline,¹¹⁹ as well as irreversible MAO-A selective inhibitors such as brofaromine.³ Abnormal REM sleep phenomena like sleep paralysis, vivid dreaming and hypnagogic hallucinations, are also treated with amphetamine-like compounds or a combination of antidepressants plus modafinil.^3,7

Cataplexy, a pathognomonic sudden loss of muscle tone and posture control in narcolepsy, is typically triggered by emotional arousal such as fear and joy (in humans) or rewarding stimuli such as food (in animals). It is thought to depend on a dysbalance of cholinergic (aggravation) and adrenergic (suppression) influences modulating poly- and monosynaptic reflex loops in upper and lower motor pathways^120-123 through activation of cholinergic M₂, adrenergic a_1b receptors and adrenergic/dopaminergic a₂/D₂ autoreceptors.³ Canine cataplexy is thus pharmacologically suppressed by antagonists of D₂ or M₂ receptors, and agonists of a₁D₁ or 5-HT_1A receptors, as well as TRH analogues, whereas it is aggravated by agonists of D₂ or M₂ receptors, physostigmine and a₁ receptor antagonists, and thalidomide.³ Likewise, injection of orexins into the aminergic and/or cholinergic centers in the brainstem suppresses cataplexy in narcoleptic dogs.^120,121 Most of the currently prescribed anticataplectic drugs in humans, including tricyclic antidepressants such as imipramine, desipramine and clomipramine, enhance the availability of noradrenaline by inhibiting its uptake and/or degradation.^3,7 Fluoxetine, a serotonin-reuptake inhibitor, as well as sibutramine, an approved antiobesity drug, and norepinephrine, serotonin and dopamine reuptake inhibitors are also effective,¹²⁴ emphasizing the molecular and functional convergence between aminergic, cholinergic and orexinergic signaling pathways⁸⁶ and food, mood, sleep and motor functions.

Amphetamines

Amphetamine-like psychostimulants such as d-amphetamine, cocaine and methylphenidate interfere with dopamine reuptake and/or release. They are the most potent wake-promoting,¹²⁵ appetite-suppressing and reinforcing drugs known in animals and humans.^68,126 Orexins excite dopaminergic neurons in the ventral tegmental area in vitro.^92,93 Moreover, insensitivity of dopamine transporter (DAT) knockout mice to the normally robust wake-promoting actions of amphetamines and modafinil¹²⁵ together with the discovery of wake-active neurons in the periaqueductal gray,¹²⁷ an extension of the mesolimbic dopaminergic reward system, provide strong evidence for a complex role of dopamine-orexin interactions (Fig. 3) in sleep-wake regulation and narcolepsy as well as natural reward⁶⁸ and memory functions and their usurpation in compulsion and addiction.^126,128,129

Fig. 3. Interactions of the orexin system.

Modafinil

Modafinil (Fig. 2), for which the exact mechanism of action is still not known, exhibits remarkable selectivity for hypothalamic arousal systems and has been shown to activate orexin neurons in rodents,^114,130 although the wake-promoting actions of both mo-dafinil and amphetamines are independent from OX₂R activation.^{33,114,125,130} Modafinil, similar to other psycho-stimulants, increases locomotor activity and has slight euphoric and reinforcing effects, but does not reduce canine cataplexy,³ and unlike other sympathomimetic agents has only marginal side effects on cardiovascular and hemodynamic parameters. Thus, mo-dafinil is biochemically and pharmacologically distinct from prototypical psychostimulants or sympathomimetic amines. Modafinil-induced wakefulness can be attenuated by the a₁-adrenergic receptor antagonist prazosin. However, modafinil at pharmacologically relevant concentrations does not bind to any of the receptors implicated in sleep-wake regulation, including those for norepinephrine, serotonin, dopamine, GABA, adenosine,¹³¹ histamine,⁹¹ melatonin¹³² and GABA,¹³³ and also does not inhibit MAO-B or phosphodiesterase II-V activities in vitro.

Recently, a sex-specific polymorphism of the gene encoding for catechol-O-methyltransferase (COMT), an enzyme that inactivates noradrenaline and dopamine,¹³⁴ has been found linking the severity of narcolepsy with its sensitivity to modafinil.¹³⁵ COMT gene polymorphism has also been implicated with obsessive-compulsive disorder, rapid cycling bipolar disorder, schizophrenia and mu opioid neurotransmitter responses to pain stressors,^136,137 pointing to a molecular and functional convergence of dopaminergic-noradrenergic signaling and the susceptibility to disorders of mood, thought, pain and sleep as well as to a putative mechanism of the action for modafinil. Modafinil binds to dopa-mine reuptake sites and causes increases in extracellular dopamine, but does not affect dopamine release. Wakefulness and hyperlocomotion induced by modafinil, in contrast to those induced by orexins and amphetamines, are not antagonized by dopamine receptor antagonists such as haloperidol.^{53,62,138-140} In addition, modafinil is considered to be less addictive and to have minimal abuse risk compared with amphetamines.¹⁴¹ However, further studies are warranted to establish the exact mechanisms of action of modafinil.

Caffeine

Caffeine, in doses equivalent to two cups of coffee, is the most commonly used wake-promoting drug world-wide¹⁴² and also effective in narcolepsy.^3,7 Mechanisms by which caffeine promotes wakefulness include antagonism of (inhibitory) A₁Rs in the cholinergic arousal centers of the basal forebrain¹⁴³ and of low-affinity (excitatory) A₂Rs in the sleep-active ventrolateral preoptic area¹⁴⁴ and striatum.¹⁴⁵ Moreover, A₁Rs are expressed in orexin neurons,¹³¹ suggesting that orexinergic signaling is controlled by purinergic influences and also mediate some of the pharmacological effects of caffeine. Notably, DAT knockout mice are insensitive to amphetamines and modafinil, but hypersensitive to caffeine,¹²⁵ emphasizing a crosstalk be- tween aminergic and adenosinergic pathways in behavioral state control. Moreover, dopamine-adenosine interactions have been implicated in addictive behaviors such as ethanol consumption¹⁴⁶ through intracellular path- ways converging on DARRP-32,¹⁴⁵ which in turn is also involved in the biochemical and behavioral effects of selective serotonin reuptake inhibitors such as fluoxetine.¹⁴⁷

g-Hydroxybutyric acid

An endogenous psychoactive GABA metabolite, g-hydroxybutyric acid (GHB), is another recently approved treatment option for narcolepsy and is also used in resuscita- tion, anesthesia and addiction therapy.^148,149 GHB strongly increases and consolidates fragmented slow-wave sleep and cataplexy in narcolepsy. However, euphoric, addictive ("liquid ecstasy") and anabolic side effects (GH release) of GHB, implying a high risk for drug abuse, limit its therapeutic use.^148,149 In vitro, GHB binds with high affinity to specific receptors located preferentially in target structures of dopaminergic transmission and with lower affinity to widely distributed GABA_B receptors.¹⁴⁸ Endogenous GHB accumulates during hepatic failure, alcoholic intoxication or defects of succinate semialdehyde dehydrogenase, the enzyme that degrades GHB. Mice with a deficiency in succinate semialdehyde dehydrogenase exhibit lethal tonic-clonic seizures, which interestingly can be rescued by dietary supplementation of the amino acid taurine¹⁵⁰ or pharmacological intervention with vigabatrin, or the GABA_B receptor antagonist CGP-35348. Because of its strong link to GABA metabolism, GHB may also act in the endogenous GABAergic sleep pathway^151,152 from the VLPO to the wake-promoting histaminergic system in the tuberomamillary nucleus.^{91,133,153,154}

Together, the pharmacotherapy of narcolepsy emphasizes a high degree of molecular and functional convergence between orexins and aminergic, cholinergic, purinergic and GABAergic signaling pathways. Moreover, the overlap of neuroendocrine-peptidergic mechanisms controlling energy homeostasis and behavioral state as well as the clinical symptomatology and co-morbidity of narcolepsy with other neuropsychiatric diseases such as depression and migraine support a functional link of the orexin system with disorders of mood, eating, sleep, memory and pain. In fact, most of the drugs used to treat narcolepsy have high abuse potential in normal subjects but rarely cause dependence or addiction in narcoleptic patients.

Aminergic, cholinergic, purinergic, GABAergic and glutamatergic behavioral state control systems

Orexins strongly interact with the aminergic and cholinergic behavioral state control systems^11,27,28,114 in the locus coeruleus (noradrenaline),^155,156 dorsal raphe (serotonin),^86,87 ventral tegmental area/substantia nigra (dopamine/GABA),^92,93 lateral dorsal tegmentum/basal forebrain (acetylcholine/GABA)^157-159 and tuberomamillary nucleus (histamine).⁹¹ These transmitter systems, together with glutamatergic, GABAergic and purinergic signals, in turn influence the intrinsic properties of hypothalamic orexin neurons, providing feedback and feed-forward mechanisms regulating orexinergic activity and functions.^21-23 Orexin neurons in vitro are strongly excited by glutamate and inhibited by GABA, noradrenaline and serotonin but not histamine or acetylcholine, suggesting differential and probably state-dependent¹⁵⁹ feedback mechanisms. Many psychoactive drugs interfere with these transmitters, and thus, depending on their pharmacological profile, are likely to interfere with orexinergic activity and function as well. Atypical neuroleptics with known side effects on body weight have recently been shown to activate a subset of orexinergic neurons in the lateral hypothalamus.¹³⁸ In line with a negative noradrenergic and serotonergic feedback onto orexin neurons in hypothalamic slices,²¹ the effect of the selective serotonin reuptake inhibitor fluoxetine on food intake was not associated with an upregulation of orexinergic activity markers.¹⁶⁰ In addition, interference of psychoactive drugs targeting noradrenergic serotonergic, as well as dopaminergic and purinergic pathways with orexins, is evidenced by their efficacy in the treatment of narcolepsy-cataplexy (see above).

Histamine

Although known for a long time, the histaminergic system in the tuberomamillary nucleus of the hypothalamus is a fairly neglected regulator of behavioral state.¹⁶¹ Because of its close anatomical and functional relationship with the orexin system,^{33,73,91,113,162,163} histamine experiences a renaissance as an important mediator of arousal and hunger-related anticipatory locomotor activity.¹⁶⁴ Mice lacking the histamine-synthesizing enzyme histidine-decarboxylase similar to those lacking H₁Rs¹⁶³ have difficulties maintaining wakefulness in novel environments,¹⁶⁵ suggesting that central histamine, similar to what has been attributed to orexins, gates cortical arousal according to metabolic and environmental challenges or stress. Indeed, the wake-promoting action of orexins¹⁶³ as well as the effect of leptin and amylin on food intake are absent in H₁R knockout mice,^166,167 and good evidence from a large-scale analysis of knockout mice indicates a positive feedback loop between histamine and orexin neurons.⁷³ Centrally active drugs with an H₁R antagonistic profile are long known for their sedative properties and secondary effects on body weight. Moreover, the GABAergic projections from the VLPO to the histaminergic tuberomamillary neurons^133,153 have recently been identified as an endogenous sleep pathway and major target of the sedating effects of commonly used anesthetics and adrenergic a₂R agonists.^151,152 Notably, this pathway is gated by orexins and dynorphin through presynaptic OX₂ and opioidergic k receptors.^91,154 The strong anatomical and functional interactions between histamine, orexin (Fig. 4) and other neuroendocrine-peptidergic systems not only identify histamine as a key regulator in metabolic and behavioral state control but also as a major effector pathway and relay station for neurohumoral signals. Histamine may thus be a promising drug target for disorders of food, mood, sleep and memory.¹⁶¹

Fig. 4. Interactions between histaminergic and orexinergic neurons.

Glutamate

Glutamate and GABA are the most abundant and important excitatory and inhibitory neurotransmitters in the nervous system. They play major roles in the control of neuronal activity and plasticity in most neuronal networks, including the hypothalamus. Orexin neurons are immunoreactive for glutamate,^168,169 and a glutamatergic feed-forward mechanism has been proposed to drive activity of orexin neurons in hypothalamic slices.^21-23 Moreover, orexins, through presynaptic mechanisms of action, facilitate glutamate release in other subcortical and cortical structures as well,^1,170-173 suggesting that orexins may synchronize glutamatergic transmission according to metabolic and behavioral state throughout the entire neuraxis. Drugs interfering with ionotropic and metabotropic glutamate receptors and/or glutamate transporters such as centrally active antiepileptics, analgesics (e.g., ketamine), and muscle relaxing or neuroprotective NMDAR antagonists (memantine, amantadine), used in the treatment of neurodegen-erative diseases such as Parkinson's or Alzheimer's disease, may also in-terfere with orexinergic activity and functions.

g-Amino butyric acid

Hypothalamic orexin neurons in vitro are strongly inhibited by the GABA agonist muscimol, reflecting direct or indirect GABAergic feedback control mechanisms.^21-23 GABAergic projections to the lateral hypothalamus arise from cortical and subcortical structures such as the lateral septum, amygdala and nucleus accumbens, but also via the endogenous sleep pathway from the VLPO, providing convergent input from emotional and behavioral state control systems. Sensitivity of orexin neurons to GABA may relay some of the sedating, amnesic and addictive properties of benzodiazepines, anesthetics and narcotics including ethanol.^133,151-153 Orexins in turn, preferentially through post- and presynaptic OX₂Rs, directly excite GABAergic neurons and facilitate GABA release in various targets, in-cluding the nucleus raphe,⁸⁷ substantia nigra/ventral tegmental area,⁹³ pineal gland,¹³² basal forebrain,¹⁵⁸ medial septum,^63,174 nucleus accumbens¹⁷⁵ and inputs from the VLPO to the wake-promoting histaminergic neurons in the tuberomammillary nucleus (Fig. 3).¹⁵⁴

Metabolic-nutritional, circadian-photic, immunological and neuroendocrine-peptidergic signals

Hypothalamic neurons integrate nutritional-metabolic,^{8,14-16,20,24,176-178} circadian-photic,^71,72,74-76 immunological^179,180 and neuroendocrine-peptidergic^69,181 influences. They finally relay this information to the major central gateways that control autonomic outflow including the aminergic^{86,91,92,156,182} and cholinergic^157,158 centers in the brainstem and hypothalamus,^19,183 as well as to the peripheral components of the autonomous nervous system^{17,19,26,184-186} and endocrine glands.^32,50,187

Food restriction or fasting, concurrent with falling blood-glucose levels or insulin-induced hypoglycemia as well as leptin-deficiency are major triggers of orexinergic activity, evidenced by increased c-Fos and prepro-orexin expression^2,8,176,188 as well as increased excitability of orexin neurons in vitro.^14,24 Food restriction or fasting inversely changes blood plasma levels of orexin-A and leptin¹⁸⁹ and increases vigilance and locomotor activity both in animals and humans,^190,191 supporting histamine-, dopamine- and opioid-dependent food seeking behavior¹⁶⁴ and natural reward.⁶⁸ Moreover, (short-term) fasting has antinociceptive, antiinflammatory, antihypertensive, antiaging and antidepressive, euphoric and cognitive enhancing effects. Conversely, postprandial somnolence is a commonly observed phenomenon, and high-caloric food intake is associated with a variety of pathological conditions that converge in metabolic syndromes and immune dysfunctions including obesity, hypertension and diabetes mellitus.^192,193 Notably, human narcolepsy is also associated with metabolic abnormalities, including increased body-mass index and frequency of non-insulin-dependent diabetes mellitus^194-196 as well as migraine,¹¹² a periodically recurrent pain disorder associated with neurogenic inflammation. Similarly, orexin neuron-ablated knockout mice are obese in spite of eating less and typically lack fasting-induced effects on vigilance and locomotor effects usually observed in healthy animals.^2,24,116 Together, these observations strongly suggest that orexins are a molecular prerequisite linking metabolic and behavioral state^2,24 by setting thresholds of arousal and motivated appetitive-incentive behaviors.^9,11 Thus, even behavioral and lifestyle changes relayed through the endogenous orexin system may affect neuroendocrine-autonomic and somatic body functions, providing promising yet underestimated therapeutic avenues for many disorders of eating, mood, sleep and memory.^{68,69,126,181,197,198}

Systematic high-throughput screen- ing for orphan receptors and their ligands by modern reverse genetic¹ and pharmacologic² approaches is expected to bring up more putative drug targets and candidate molecules to interfere with orexins.^69,181,198 Evidence indicating close neuroanatomical and functional interactions with orexins exists for MCH,¹⁹⁹ ghrelin,^24,200,201 leptin,^{15,18,24,78,179,180} NPY, CRF and vasopressin-oxytocin,^19,35,44-49 and gonado-tropin-releasing hormone.^56,187 Here, we will briefly focus on dynorphin since it is highly co-expressed in orexin neurons²⁰² and thus probably has specific impact for orexinergic activity and function.

Dynorphin

Recent in vitro evidence suggests that dynorphins together with orexins gate the GABAergic input from the VLPO to the wake-active histaminergic neurons in the tuberomamillary nucleus through presynaptic mechanisms.¹⁵⁴ This pathway is a prominent target of commonly used anesthetics and adrenergic a₂-agonists.^151,152 Moreover, orexin but not MCH-induced feeding responses depend on opioids.²⁰³ Finally, the orexin system is selectively activated by withdrawal from drugs of abuse such as morphine and cocaine,^128,129 suggesting that it plays a role not only in natural reward⁶⁸ but also compulsion and addiction.¹²⁶

Conclusion

Drugs interfering directly with the orexins, such as selective OXR antagonists and agonists, not only improve our knowledge of orexinergic signaling, but also allow a pharmacologic segregation of OX₁ and OX₂ receptors according to their functions in health and disease. Drugs targeting the complex array of neurohumoral signals associated with the orexin system can influence the bistable control of behavioral state, body weight and other homeostatic body functions. Further research is needed to specify the exact role of orexins within this context. However, development of drugs targeting the orexin system both directly or indirectly will not only lead to a better pharmacotherapy of narcolepsy but will also open novel therapeutic avenues for the treatment of a number of neuropsychiatric and neuroendocrine disorders of eating, pain, mood, sleep and memory.

Acknowledgments

This work was supported by EU-QLRT 826 and DFG-SFB 575.

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Oliver Selbach* and Krister S. Eriksson are Researchers, and Helmut L. Haas is Director in the Department of Neurophysiology, Heinrich-Heine-University, Dusseldorf, Germany. *Correspondence: Oliver Selbach, Department of Neurophysiology, Heinrich-Heine-University, P.O. Box 101007, D-40001 Dusseldorf, Ger-many; Tel: +49 211 8112687, Fax: +49 211 8114231; E-mail: selbach@ uni-duesseldorf.de.

Drug News & Perspectives Vol. 16, No. 10, 2003, p. 669-681
ISSN 0214-0934 Copyright 2003 Prous Science, S.A. CCC: 0214 0934/2003 http://www.prous.com

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