Orexin Hypocretin Research Articles

Major alteration of motor control during rapid eye movements sleep in mice models of sleep disorders.

Alteration of motor control during rapid eye movements (REM) sleep has been extensively described in sleep disorders, in particular in isolated REM sleep behavior disorder (iRBD) and narcolepsy type 1 (NT1). NT1 is caused by the loss of orexin/hypocretin (ORX) neurons. Unlike in iRBD, the RBD comorbid symptoms of NT1 are not associated with alpha-synucleinopathies. To determine whether the chronic absence of ORX neuropeptides is sufficient to induce RBD symptoms, we analyzed during REM sleep the EMG signal of the prepro-hypocretin knockout mice (ORX-/-), a recognized mouse model of NT1. Then, we evaluated the severity of motor alterations by comparing the EMG data of ORX-/- mice to those of mice with a targeted suppression of the sublaterodorsal glutamatergic neurotransmission, a recognized rodent model of iRBD. We found a significant alteration of tonic and phasic components of EMG during REM sleep in ORX-/- mice, with more phasic events and more REM sleep episodes without atonia compared to the control wild-type mice. However, these phasic events were fewer, shorter, and less complex in ORX-/- mice compared to the RBD-like ORX-/- mice. We thus show that ORX deficiency, as seen in NT1, is sufficient to impair muscle atonia during REM sleep with a moderate severity of alteration as compared to isolated RBD mice. As described in NT1 patients, we report a major interindividual variability in the severity and frequency of RBD symptoms in ORX-deficient mice.

Downregulation of hypocretin/orexin after H1N1 Pandemrix vaccination of adolescent mice.

Narcolepsy type 1 (NT1), characterized by the loss of hypocretin/orexin (HCRT) production in the lateral hypothalamus, has been linked to Pandemrix vaccination during the 2009 H1N1 pandemic, especially in children and adolescents. It is still unknown why this vaccination increased the risk of developing NT1. This study investigated the effects of Pandemrix vaccination during adolescence on Hcrt mRNA expression in mice. Mice received a primary vaccination (50 µL i.m.) during prepubescence and a booster vaccination during peri-adolescence. Hcrt expression was measured at three-time points after the vaccinations. Control groups included both a saline group and an undisturbed group of mice. Hcrt expression was decreased after both Pandemrix and saline injections, but 21 days after the second injection, the saline group no longer showed decreased Hcrt expression, while the Pandemrix group still exhibited a significant reduction of about 60% compared to the undisturbed control group. This finding suggests that Pandemrix vaccination during adolescence influences Hcrt expression in mice into early adulthood. The Hcrt mRNA level did not reach the low levels known to induce NT1 symptoms, instead, our finding supports the multiple-hit hypothesis of NT1 that states that several insults to the HCRT system may be needed to induce NT1 and that Pandemrix could be one such insult.

Inactivation of hypocretin receptor-2 signaling in dopaminergic neurons induces hyperarousal and enhanced cognition but impaired inhibitory control

Hypocretin/Orexin (HCRT/OX) and dopamine (DA) are both key effectors of salience processing, reward and stress-related behaviors and motivational states, yet their respective roles and interactions are poorly delineated. We inactivated HCRT-to-DA connectivity by genetic disruption of Hypocretin receptor-1 (Hcrtr1), Hypocretin receptor-2 (Hcrtr2), or both receptors (Hcrtr1&2) in DA neurons and analyzed the consequences on vigilance states, brain oscillations and cognitive performance in freely behaving mice. Unexpectedly, loss of Hcrtr2, but not Hcrtr1 or Hcrtr1&2, induced a dramatic increase in theta (7–11 Hz) electroencephalographic (EEG) activity in both wakefulness and rapid-eye-movement sleep (REMS). DAHcrtr2-deficient mice spent more time in an active (or theta activity-enriched) substate of wakefulness, and exhibited prolonged REMS. Additionally, both wake and REMS displayed enhanced theta-gamma phase-amplitude coupling. The baseline waking EEG of DAHcrtr2-deficient mice exhibited diminished infra-theta, but increased theta power, two hallmarks of EEG hyperarousal, that were however uncoupled from locomotor activity. Upon exposure to novel, either rewarding or stress-inducing environments, DAHcrtr2-deficient mice featured more pronounced waking theta and fast-gamma (52–80 Hz) EEG activity surges compared to littermate controls, further suggesting increased alertness. Cognitive performance was evaluated in an operant conditioning paradigm, which revealed that DAHcrtr2-ablated mice manifest faster task acquisition and higher choice accuracy under increasingly demanding task contingencies. However, the mice concurrently displayed maladaptive patterns of reward-seeking, with behavioral indices of enhanced impulsivity and compulsivity. None of the EEG changes observed in DAHcrtr2-deficient mice were seen in DAHcrtr1-ablated mice, which tended to show opposite EEG phenotypes. Our findings establish a clear genetically-defined link between monosynaptic HCRT-to-DA neurotransmission and theta oscillations, with a differential and novel role of HCRTR2 in theta-gamma cross-frequency coupling, attentional processes, and executive functions, relevant to disorders including narcolepsy, attention-deficit/hyperactivity disorder, and Parkinson’s disease.

Hypocretin / Orexin Receptor 1 Knockdown in GABA or Dopamine Neurons in the Ventral Tegmental Area Differentially Impact Mesolimbic Dopamine and Motivation for Cocaine

The hypocretins/orexins (HCRT) have been demonstrated to influence motivation for cocaine through actions on dopamine (DA) transmission. Pharmacological or genetic disruption of the hypocretin receptor 1 (Hcrtr1) reduces cocaine self-administration, blocks reinstatement of cocaine seeking, and decreases conditioned place preference for cocaine. These effects are likely mediated through actions in the ventral tegmental area (VTA) and resulting alterations in DA transmission. For example, HCRT drives VTA DA neuron activity and enhances the effects of cocaine on DA transmission, while disrupting Hcrtr1 attenuates DA responses to cocaine. These findings have led to the perspective that HCRT exerts its effects through Hcrtr1 actions in VTA DA neurons. However, this assumption is complicated by the observation that Hcrtr1 are present on both DA and GABA neurons in the VTA and HCRT drives the activity of both neuronal populations. To address this issue, we selectively knocked down Hcrtr1 on either DA or GABA neurons in the VTA and examined alterations in DA transmission and cocaine self-administration in female and male rats. We found that Hcrtr1 knockdown in DA neurons decreased DA responses to cocaine, increased days to acquire cocaine self-administration, and reduced motivation for cocaine. Although, Hcrtr1 knockdown in GABA neurons enhanced DA responses to cocaine, this manipulation did not affect cocaine self-administration. These observations indicate that while Hcrtr1 on DA versus GABA neurons exert opposing effects on DA transmission, only Hcrtr1 on DA neurons affected acquisition or motivation for cocaine – suggesting a complex interplay between DA transmission and behavior.

Epigenetic silencing of selected hypothalamic neuropeptides in narcolepsy with cataplexy

Narcolepsy with cataplexy isa sleep disorder caused by deficiency in the hypothalamic neuropeptide hypocretin/orexin (HCRT), unanimously believed to result from autoimmune destruction of hypocretin-producing neurons. HCRT deficiency can also occur in secondary forms of narcolepsy and be only temporary, suggesting it can occur without irreversible neuronal loss. The recent discovery that narcolepsy patients also show loss of hypothalamic (corticotropin-releasing hormone) CRH-producing neurons suggests that other mechanisms than cell-specific autoimmune attack, are involved. Here, we identify the HCRT cell-colocalized neuropeptide QRFP as the best marker of HCRT neurons. We show that if HCRT neurons are ablated in mice, in addition toHcrt, Qrfp transcript is also lost in the lateral hypothalamus, while in mice where only theHcrtgene is inactivated Qrfp is unchanged. Similarly, postmortem hypothalamic tissues of narcolepsy patients show preservedQRFPexpression, suggesting the neurons are present but fail to actively produce HCRT. We show that the promoter of theHCRTgene of patients exhibits hypermethylation at a methylation-sensitive and evolutionary-conserved PAX5:ETS1 transcription factor-binding site, suggesting the gene issubject to transcriptional silencing. We show also that in addition to HCRT,CRHand Dynorphin (PDYN) gene promoters, exhibit hypermethylation in the hypothalamus of patients. Altogether, we propose that HCRT,PDYN, andCRH are epigenetically silenced by a hypothalamic assault (inflammation) in narcolepsy patients,without concurrent cell death. Since methylation is reversible, our findings open the prospect of reversing or curing narcolepsy.

Sex differences in activation of extra-hypothalamic forebrain areas during hedonic eating.

Palatable foods can stimulate appetite without hunger, and unconstrained overeating underlies obesity and binge eating disorder. Women are more prone to obesity and binge eating than men but the neural causes of individual differences are unknown. In an animal model of hedonic eating, a prior study found that females were more susceptible than males to eat palatable food when sated and that the neuropeptide orexin/hypocretin (ORX) was crucial in both sexes. The current study examined potential extra-hypothalamic forebrain targets of ORX signaling during hedonic eating. We measured Fos induction in the cortical, thalamic, striatal, and amygdalar areas that receive substantial ORX inputs and contain their receptors in hungry and sated male and female rats during palatable (high-sucrose) food consumption. During the test, hungry rats of both sexes ate substantial amounts, and while sated males ate much less than hungry rats, sated females ate as much as hungry rats. The Fos induction analysis identified sex differences in recruitment of specific areas of the medial prefrontal cortex, paraventricular nucleus of the thalamus (PVT), nucleus accumbens (ACB), and central nucleus of the amygdala (CEA), and similar patterns across sexes in the insular cortex. There was a striking activation of the infralimbic cortex in sated males, who consumed the least amount food and unique correlations between the insular cortex, PVT, and CEA, as well as the prelimbic cortex, ACB, and CEA in sated females but not sated males. The study identified key functional circuits that may drive hedonic eating in a sex-specific manner.

Characterization of T cell receptors reactive to HCRTNH2, pHA273-287, and NP17-31 in control and narcolepsy patients

Narcolepsy type 1 (NT1), a disorder caused by hypocretin/orexin (HCRT) cell loss, is associated with human leukocyte antigen (HLA)-DQ0602 (98%) and T cell receptor (TCR) polymorphisms. Increased CD4+ T cell reactivity to HCRT, especially DQ0602-presented amidated C-terminal HCRT (HCRTNH2), has been reported, and homology with pHA273-287 flu antigens from pandemic 2009 H1N1, an established trigger of the disease, suggests molecular mimicry. In this work, we extended DQ0602 tetramer and dextramer data to 77 cases and 44 controls, replicating our prior finding and testing 709 TCRs in Jurkat 76 T cells for functional activation. We found that fewer TCRs isolated with HCRTNH2 (∼11%) versus pHA273-287 or NP17-31 antigens (∼50%) were activated by their ligand. Single-cell characterization did not reveal phenotype differences in influenza versus HCRTNH2-reactive T cells, and analysis of TCR CDR3αβ sequences showed TCR clustering by responses to antigens but no cross-peptide class reactivity. Our results do not support the existence of molecular mimicry between HCRT and pHA273-287 or NP17-31.

Effects of single and dual hypocretin-receptor blockade or knockdown of hypocretin projections to the central amygdala on alcohol drinking in dependent male rats

Hypocretin/orexin (HCRT) is a neuropeptide that is associated with both stress and reward systems in humans and rodents. The different contributions of signaling at hypocretin-receptor 1 (HCRT-R1) and hypocretin-receptor 2 (HCRT-R2) to compulsive alcohol drinking are not yet fully understood. Thus, the current studies used pharmacological and viral-mediated targeting of HCRT to determine participation in compulsive alcohol drinking and measured HCRT-receptor mRNA expression in the extended amygdala of both alcohol-dependent and non-dependent male rats. Rats were made dependent through chronic intermittent exposure to alcohol vapor and were tested for the acute effect of HCRT-R1-selective (SB-408124; SB-R1), HCRT-R2-selective (NBI-80713; NB-R2), or dual HCRT-R1/2 (NBI-87571; NB-R1/2) antagonism on alcohol intake. NB-R2 and NB-R1/2 antagonists each dose-dependently decreased overall alcohol drinking in alcohol-dependent rats, whereas, SB-R1 decreased alcohol drinking in both alcohol-dependent and non-dependent rats at the highest dose (30 mg/kg). SB-R1, NB-R2, and NB-R1/2 treatment did not significantly affect water drinking in either alcohol-dependent or non-dependent rats. Additional PCR analyses revealed a significant decrease in Hcrtr1 mRNA expression within the central amygdala (CeA) of dependent rats under acute withdrawal conditions compared to non-dependent rats. Lastly, a shRNA-encoding adeno-associated viral vector with retrograde function was used to knockdown HCRT in CeA-projecting neurons from the lateral hypothalamus (LH). LH-CeA HCRT knockdown significantly attenuated alcohol self-administration in alcohol-dependent rats. These observations suggest that HCRT signaling in the CeA is necessary for alcohol-seeking behavior during dependence. Together, these data highlight a role for both HCRT-R1 and -R2 in dependent alcohol-seeking behavior.

Twenty-Three Years of Hypocretins: The "Rosetta Stone" of Sleep/Arousal Circuits.

The discovery of the hypocretins/orexins (HCRTs) has revolutionized sleep science in the last two decades. A combination of anatomical tracing methods, optogenetics, and pharmacology is delineating a blueprint of functional inputs and outputs of the HCRT system. Here, we discuss several models of HCRT action that involve the integration between physiological variables, circadian output, and sleep homeostasis. Generation of activity maps during the sleep-wake cycle at the cellular level will allow investigators to decipher computational frameworks modeling operations of HCRT networks.

Hedonic Eating: Sex Differences and Characterization of Orexin Activation and Signaling

Palatable taste can stimulate appetite in the absence of hunger, and individual differences in hedonic eating may be critical to overeating. Women are more prone to obesity and binge eating than men, which warrants comparisons of hedonic versus physiological consumption and the underlying neural substrates in both sexes. The current study examined palatable (high-sugar) food consumption in male and female rats under physiological hunger and satiety, and the role of the neuropeptide orexin/hypocretin (ORX). Across multiple tests, females consistently consumed similar amounts of palatable food regardless of whether they were hungry or sated prior to testing. In contrast, males typically adjusted their consumption according to their hunger/satiety state. This difference was specific to palatable food consumption, as both sexes ate standard chow according to their hunger state. ORX is important in food motivation and reward behaviors. Thus, to begin to determine the neuronal mechanisms of hedonic eating, we examined activation and signaling of ORX neurons. We systematically characterized Fos induction patterns of ORX neurons across the entire rostrocaudal extent of the lateral hypothalamus and found that they were activated by food and by fasting in both sexes. Then, we showed that systemic blockade of ORX receptor 1 signaling with SB-334867 decreased palatable food consumption in hungry and sated rats of both sexes. These results demonstrate sex differences in hedonic eating; increased susceptibility in females to overeat palatable food regardless of hunger state, and that ORX is a critical neuropeptide mechanism of hedonic eating in both sexes.

Cannabidiol Partially Blocks the Excessive Sleepiness in Hypocretindeficient Rats: Preliminary Data.

Excessive daytime sleepiness and cataplexy are among the symptoms of narcolepsy, a sleep disorder caused by the loss of hypocretin/orexin (HCRT/OX) neurons placed into the Hypothalamus (LH). Several treatments for managing narcolepsy include diverse drugs to induce alertness, such as antidepressants, amphetamine, or modafinil, etc. Recent evidence has shown that cannabidiol (CBD), a non-psychotropic derived from Cannabis sativa, shows positive therapeutic effects in neurodegenerative disorders, including Parkinson´s disease. Furthermore, CBD provokes alertness and enhances wake-related neurochemicals in laboratory animals. Thus, it is plausible to hypothesize that excessive somnolence observed in narcolepsy might be blocked by CBD. Here, we determined whether the systemic injection of CBD (5mg/kg, i.p.) would block the excessive sleepiness in a narcoleptic model. To test this idea, the neurotoxin hypocretin-2-saporin (HCRT2/SAP) was bilaterally injected into the LH of rats to eliminate HCRT leading to the establishment of narcoleptic-like behavior. Since excessive somnolence in HCRT2/SAP lesioned rats has been observed during the lights-off period, CBD was administered at the beginning of the dark phase. Hourly analysis of sleep data showed that CBD blocked the sleepiness during the lights-off period across 7h post-injection in lesioned rats. Taking together, these preliminary findings suggest that CBD might prevent sleepiness in narcolepsy.

Involvement of orexin/hypocretin in the expression of social play behaviour in juvenile rats

ABSTRACTSocial play is a highly rewarding and motivated behaviour displayed by juveniles of many mammalian species. We hypothesized that the orexin/hypocretin (ORX) system is involved in the expression of juvenile social play behaviour because this system is interconnected with brain regions that comprise the social behaviour and mesocorticolimbic reward networks. We found that exposure to social play increased recruitment of ORX-A neurons in juvenile rats. Furthermore, central administration of ORX-A decreased social play duration, while central blockade of ORX-1 receptors differentially altered social play duration in juvenile rats with low versus high baseline levels of social play (increasing social play in low baseline social play individuals and decreasing social play in high baseline social play individuals). Together, our results provided the first evidence of a role for the ORX system in the modulation of juvenile social play behaviour.

Depletion of hypothalamic hypocretin/orexin neurons correlates with impaired memory in a Parkinson's disease animal model

Parkinson's disease (PD) is a progressive and chronic neurodegenerative disorder characterized by loss of dopaminergic neurons in the substantia nigra pars compacta (SNpc) and affects multiple neurotransmission systems such as hypocretin/orexin (HO) release and can lead to cognitive and memory deficits. The HO neurons located in lateral hypothalamus/perifornical area (LH/PeF) are involved with consolidation and memory processes. Here we verified the involvement of HO deficit in learning and memory process in an animal model of PD induced by bilateral intra-striatal injections of 6-hydroxydopamine (6-OHDA). The present study performed a working memory test by object recognition task and spatial memory test using the Morris water maze in control and PD-induced animals after depletion of HO neurons. In addition, our results indicate that HO system in degenerative disorders such as PD may modulate the declarative and spatial memory (assessed by object recognition and Morris water maze tests, respectively). A significant reduction of HO neurons in the LH/PeF and HO degeneration process in the hippocampus (CA1 and dentate gyrus areas) were noticed. Our data suggest that the HO system degeneration could be associated to memory dysfunction in PD.

Molecular codes and in vitro generation of hypocretin and melanin concentrating hormone neurons

Hypocretin/orexin (HCRT) and melanin concentrating hormone (MCH) neuropeptides are exclusively produced by the lateral hypothalamus and play important roles in sleep, metabolism, reward, and motivation. Loss of HCRT (ligands or receptors) causes the sleep disorder narcolepsy with cataplexy in humans and in animal models. How these neuropeptides are produced and involved in diverse functions remain unknown. Here, we developed methods to sort and purify HCRT and MCH neurons from the mouse late embryonic hypothalamus. RNA sequencing revealed key factors of fate determination for HCRT (Peg3, Ahr1, Six6, Nr2f2, and Prrx1) and MCH (Lmx1, Gbx2, and Peg3) neurons. Loss of Peg3 in mice significantly reduces HCRT and MCH cell numbers, while knock-down of a Peg3 ortholog in zebrafish completely abolishes their expression, resulting in a 2-fold increase in sleep amount. We also found that loss of HCRT neurons in Hcrt-ataxin-3 mice results in a specific 50% decrease in another orexigenic neuropeptide, QRFP, that might explain the metabolic syndrome in narcolepsy. The transcriptome results were used to develop protocols for the production of HCRT and MCH neurons from induced pluripotent stem cells and ascorbic acid was found necessary for HCRT and BMP7 for MCH cell differentiation. Our results provide a platform to understand the development and expression of HCRT and MCH and their multiple functions in health and disease.

Role of Orexin-B/Orexin 2 receptor in myocardial protection.

Emerging evidence attributes to orexins/hypocretins (ORs) a protective function in the regulation of cardiovascular responses, heart rate, and hypertension. However, little is known about any direct effect of orexins in the heart function. This is of special relevance considering that cardiovascular diseases, including myocardial infarction and heart failure, are one of the major causes of mortality in the world. In the article published inClinical Science (2018)(vol.132,2547-2564), Patel and colleagues investigated the role of orexins in myocardial protection. Intriguingly, they revealed a source of orexin-A (OR-A) and orexin-B (OR-B) in the heart and cardiomyocytes of the rat. More interestingly, these peptides exert a direct effect on the heart rate by acting in an autocrine/paracrine manner on their respective receptors (OXRs). Indeed, OR-B, but not OR-A, by acting through orexin receptor-2 (OX2R), exerts direct cardioprotective effects in heart failure models. OR-B/OX2R signalling enhances myosin light chain (MLC) and troponin-I (TnI) phosphorylation in a dose-dependent manner, leading to an increase in the strength of their twitch contraction. This effect is mediated by extracellular signal-regulated kinase 1/2 (ERK1/2) and Akt phosphorylation, both in the rat myocardial tissue and human heart samples. A negative correlation between OX2R expression and clinical severity of symptoms has been found in patients with heart failure. Thus, in addition to the known central effects of orexins/OX2R, the work of Patel and colleagues (Clinical Science (2018) 132,2547-2564) reports a direct action of OR-B on the heart rate pinpointing to OX2R as a potential therapeutic target for prevention and treatment of cardiovascular disease.

Abstract # 3107 A role for hypocretin/orexin in metabolic and sleep abnormalities in a mouse model of non-metastatic breast cancer

Among cancer patients, metabolic and sleep abnormalities are significant problems that are associated with decreased quality of life and increased mortality. In spite of the abundance of metabolic and sleep abnormalities in cancer patients, the mechanisms by which tumors alter metabolism and sleep remain unknown. Utilizing a non-metastatic murine breast cancer model, we investigated the relationships among immune, metabolic, and sleep abnormalities. Tumor bearing mice displayed increased serum interleukin-6 concentrations, concurrent with hyperglycemia, altered gluconeogenesis/glycolysis gene expression, and sleep abnormalities. To test whether tumor induced elevations in IL-6 were responsible for the metabolic and sleep abnormalities observed in our study, we reduced IL-6 signaling by administering a monoclonal antibody against IL-6. Despite successfully suppressing IL-6 signaling, metabolic and sleep abnormalities remained. Next, we investigated hypocretin/orexin (HO) signaling in the lateral hypothalamus as this population of neurons is known to regulate both metabolism and sleep, and observed increased HO activity. We administered a dual HO receptor antagonist (almorexant) to test whether the increased HO signaling was driving the altered hepatic glucose production and hyperglycemia. Administration of almorexant rescued the tumor-induced deficits in hyperglycemia, hepatic glucose production, and improved sleep quality, independent of IL-6. Together, our data demonstrate a novel mechanism by which non-metastatic tumors can promote energy availability, and suggest that targeting HO system may represent a potential intervention for breast cancer patients with metabolic dysfunctions.

Autoimmunity to hypocretin and molecular mimicry to flu in type 1 narcolepsy

Type 1 narcolepsy (T1N) is caused by hypocretin/orexin (HCRT) neuronal loss. Association with the HLA DQB1*06:02/DQA1*01:02 (98% vs. 25%) heterodimer (DQ0602), T cell receptors (TCR) and other immune loci suggest autoimmunity but autoantigens are unknown. Onset is seasonal and associated with influenza A, notably pandemic 2009 H1N1 (pH1N1) infection and vaccination (Pandemrix). Peptides derived from HCRT and influenza A, including pH1N1, were screened for DQ0602 binding and presence of cognate DQ0602 tetramer-peptide-specific CD4+ T cells tested in 35 T1N cases and 22 DQ0602 controls. Higher reactivity to influenza pHA273-287 (pH1N1 specific), PR8 (H1N1 pre-2009 and H2N2)-specific NP17-31 and C-amidated but not native version of HCRT54-66 and HCRT86-97 (HCRTNH2) were observed in T1N. Single-cell TCR sequencing revealed sharing of CDR3β TRBV4-2-CASSQETQGRNYGYTF in HCRTNH2 and pHA273-287-tetramers, suggesting molecular mimicry. This public CDR3β uses TRBV4-2, a segment modulated by T1N-associated SNP rs1008599, suggesting causality. TCR-α/β CDR3 motifs of HCRT54-66-NH2 and HCRT86-97-NH2 tetramers were extensively shared: notably public CDR3α, TRAV2-CAVETDSWGKLQF-TRAJ24, that uses TRAJ24, a chain modulated by T1N-associated SNPs rs1154155 and rs1483979. TCR-α/β CDR3 sequences found in pHA273-287, NP17-31, and HCRTNH2 tetramer-positive CD4+ cells were also retrieved in single INF-γ-secreting CD4+ sorted cells stimulated with Pandemrix, independently confirming these results. Our results provide evidence for autoimmunity and molecular mimicry with flu antigens modulated by genetic components in the pathophysiology of T1N.

Bidirectional and context-dependent changes in theta and gamma oscillatory brain activity in noradrenergic cell-specific Hypocretin/Orexin receptor 1-KO mice

Noradrenaline (NA) and hypocretins/orexins (HCRT), and their receptors, dynamically modulate the circuits that configure behavioral states, and their associated oscillatory activities. Salient stimuli activate spiking of locus coeruleus noradrenergic (NALC) cells, inducing NA release and brain-wide noradrenergic signalling, thus resetting network activity, and mediating an orienting response. Hypothalamic HCRT neurons provide one of the densest input to NALC cells. To functionally address the HCRT-to-NA connection, we selectively disrupted the Hcrtr1 gene in NA neurons, and analyzed resulting (Hcrtr1Dbh-CKO) mice’, and their control littermates’ electrocortical response in several contexts of enhanced arousal. Under enforced wakefulness (EW), or after cage change (CC), Hcrtr1Dbh-CKO mice exhibited a weakened ability to lower infra-θ frequencies (1–7 Hz), and mount a robust, narrow-bandwidth, high-frequency θ rhythm (~8.5 Hz). A fast-γ (55–80 Hz) response, whose dynamics closely parallelled θ, also diminished, while β/slow-γ activity (15–45 Hz) increased. Furthermore, EW-associated locomotion was lower. Surprisingly, nestbuilding-associated wakefulness, inversely, featured enhanced θ and fast-γ activities. Thus HCRT-to-NA signalling may fine-tune arousal, up in alarming conditions, and down during self-motivated, goal-driven behaviors. Lastly, slow-wave-sleep following EW and CC, but not nestbuilding, was severely deficient in slow-δ waves (0.75–2.25 Hz), suggesting that HCRT-to-NA signalling regulates the slow-δ rebound characterizing sleep after stress-associated arousal.

A Role for Hypocretin/Orexin in Metabolic and Sleep Abnormalities in a Mouse Model of Non-metastatic Breast Cancer

We investigated relationships among immune, metabolic, and sleep abnormalities in mice with non-metastatic mammary cancer. Tumor-bearing mice displayed interleukin-6 (IL-6)-mediated peripheral inflammation, coincident with altered hepatic glucose processing and sleep. Tumor-bearing mice were hyperphagic, had reduced serum leptin concentrations, and enhanced sensitivity to exogenous ghrelin. We tested whether these phenotypes were driven by inflammation using neutralizing monoclonal antibodies against IL-6; despite the reduction in IL-6 signaling, metabolic and sleep abnormalities persisted. We next investigated neural populations coupling metabolism and sleep, and observed altered activity within lateral-hypothalamic hypocretin/orexin (HO) neurons. We used a dual HO-receptor antagonist to test whether increased HO signaling was causing metabolic abnormalities. This approach rescued metabolic abnormalities and enhanced sleep quality in tumor-bearing mice. Peripheral sympathetic denervation prevented tumor-induced increases in serum glucose. Our results link metabolic and sleep abnormalities via the HO system, and provide evidence that central neuromodulators contribute to tumor-induced changes in metabolism.

Knockdown of hypocretin attenuates extended access of cocaine self-administration in rats.

The hypocretin/orexin (HCRT) neuropeptide system regulates feeding, arousal state, stress responses, and reward, especially under conditions of enhanced motivational relevance. In particular, HCRT neurotransmission facilitates drug-seeking behavior in circumstances that demand increased effort and/or motivation to take the drug. The present study used a shRNA-encoding adeno-associated viral vector to knockdown Hcrt expression throughout the dorsal hypothalamus in adult rats and determine the role of HCRT in cocaine self-administration. Chronic Hcrt silencing did not impact cocaine self-administration under short-access conditions, but robustly attenuated cocaine intake under extended access conditions, a model that mimics key features of compulsive cocaine taking. In addition, Hcrt silencing decreased motivation for both cocaine and a highly palatable food reward (i.e., sweetened condensed milk; SCM) under a progressive ratio schedule of reinforcement, but did not alter responding for SCM under a fixed ratio schedule. Importantly, Hcrt silencing did not affect food or water consumption, and had no consequence for general measures of arousal and stress reactivity. At the molecular level, chronic Hcrt knockdown reduced the number of neurons expressing dynorphin (DYN), and to a smaller extent melanin-concentrating hormone (MCH), in the dorsal hypothalamus. These original findings support the hypothesis that HCRT neurotransmission promotes operant responding for both drug and non-drug rewards, preferentially under conditions requiring a high degree of motivation. Furthermore, the current study provides compelling evidence for the involvement of the HCRT system in cocaine self-administration also under low-effort conditions in rats allowed extended access, possibly via functional interactions with DYN and MCH signaling.