Single-Cell Profiling Uncovers Evolutionary Divergence of Hypocretin/Orexin Neuronal Subpopulations.

Brain nuclei are traditionally defined by their anatomy, activity, and expression of specific markers. The hypothalamus contains discrete neuronal populations that coordinate fundamental behavioral functions, including sleep and wakefulness, in all vertebrates. Particularly, the diverse roles of hypocretin/orexin (Hcrt)-releasing neurons suggest functional heterogeneity among Hcrt neurons. Using single-cell RNA sequencing (scRNA-seq) and high-resolution imaging of the adult male and female zebrafish hypothalamic periventricular zone, we identified 21 glutamatergic and 28 GABAergic cell types. Integration of zebrafish and mouse scRNA-seq revealed evolutionary conserved and divergent hypothalamic cell types. The expression of specific genes, including npvf, which encodes a sleep-regulating neuropeptide, was enriched in subsets of glutamatergic Hcrt neurons in both larval and adult zebrafish. The genetic profile, activity, and neurite processing of the neuronal subpopulation that coexpresses both Hcrt and Npvf (Hcrt+Npvf+) differ from other Hcrt neurons. These interspecies findings provide a unified annotation of hypothalamic cell types and suggest that the heterogeneity of Hcrt neurons enables multifunctionality, such as consolidation of both wake and sleep by the Hcrt- and Npvf-releasing neuronal subpopulation.

Effect of optogenetic excitation of non-orexinergic neurons in the hypothalamic perifornical area on motor behaviors and cardiovascular parameters in rats.

Central command, a motor volition originating in the rostral part of the brain, plays a pivotal role in the precise regulation of autonomic nervous and cardiovascular systems. Central neuronal substrates responsible for transmitting central command signals remain incompletely understood. This study aimed to investigate the effect of optogenetic excitation of non-orexinergic (NOrx) neurons in the hypothalamic perifornical area (PeFA), where orexinergic neurons are densely distributed, on motor behaviors and cardiovascular parameters in rats. An adeno-associated viral serotype 2 vector carrying the human synapsin promoter encoding channelrhodopsin 2 (ChR2) fused to EYFP was injected into the PeFA of Sprague-Dawley rats, resulting in selective expression of ChR2-EYFP in NOrx PeFA neurons. In conscious rats, optogenetic excitation of NOrx PeFA neurons rapidly elicited walking or biting behavior, simultaneously causing pressor and tachycardiac responses regardless of the observed behavioral patterns. Under anesthesia, this excitation rapidly increased renal sympathetic nerve activity, immediately followed by sympathoinhibition. These findings suggest that NOrx PeFA neurons transmit central command signals, concurrently regulating somatomotor and autonomic nervous systems for locomotor exercise or biting behavior.

Neurochemical and genetic factors in panic disorder: a systematic review.

This systematic review addresses the complex nature of Panic Disorder (PD), characterized by recurrent episodes of acute fear, with a focus on updating and consolidating knowledge regarding neurochemical, genetic, and epigenetic factors associated with PD. Utilizing the PRISMA methodology, 33 original peer-reviewed studies were identified, comprising 6 studies related to human neurochemicals, 10 related to human genetic or epigenetic alterations, and 17 animal studies. The review reveals patterns of altered expression in various biological systems, including neurotransmission, the Hypothalamic-Pituitary-Adrenal (HPA) axis, neuroplasticity, and genetic and epigenetic factors leading to neuroanatomical modifications. Noteworthy findings include lower receptor binding of GABAA and serotonin neurotransmitters in the amygdala. The involvement of orexin (ORX) neurons in the dorsomedial/perifornical region in triggering panic reactions is highlighted, with systemic ORX-1 receptor antagonists blocking panic responses. Elevated Interleukin 6 and leptin levels in PD patients suggest potential connections between stress-induced inflammatory changes and PD. Brain-derived neurotrophic factor (BDNF) and tyrosine receptor kinase B (TrkB) signaling are implicated in panic-like responses, particularly in the dorsal periaqueductal gray (dPAG), where BDNF's panicolytic-like effects operate through GABAA-dependent mechanisms. GABAergic neurons' inhibitory influence on dorsomedial and posterior hypothalamus nuclei is identified, potentially reducing the excitability of neurons involved in panic-like responses. The dorsomedial hypothalamus (DMH) is highlighted as a specific hypothalamic nucleus relevant to the genesis and maintenance of panic disorder. Altered brain lactate and glutamate concentrations, along with identified genetic polymorphisms linked to PD, further contribute to the intricate neurochemical landscape associated with the disorder. The review underscores the potential impact of neurochemical, genetic, and epigenetic factors on the development and expression of PD. The comprehensive insights provided by this systematic review contribute to advancing our understanding of the multifaceted nature of Panic Disorder and pave the way for targeted therapeutic strategies.

Increased Expression of Orexin-A in Patients Affected by Polycystic Kidney Disease.

Orexin-A is a neuropeptide product of the lateral hypothalamus that acts on two receptors, OX1R and OX2R. The orexinergic system is involved in feeding, sleep, and pressure regulation. Recently, orexin-A levels have been found to be negatively correlated with renal function. Here, we analyzed orexin-A levels as well as the incidence of SNPs in the hypocretin neuropeptide precursor (HCRT) and its receptors, HCRTR1 and HCRTR2, in 64 patients affected by autosomal dominant polycystic kidney disease (ADPKD) bearing truncating mutations in the PKD1 or PKD2 genes. Twenty-four healthy volunteers constituted the control group. Serum orexin-A was assessed by ELISA, while the SNPs were investigated through Sanger sequencing. Correlations with the main clinical features of PKD patients were assessed. PKD patients showed impaired renal function (mean eGFR 67.8 ± 34.53) and a statistically higher systolic blood pressure compared with the control group (p < 0.001). Additionally, orexin-A levels in PKD patients were statistically higher than those in healthy controls (477.07 ± 69.42 pg/mL vs. 321.49 ± 78.01 pg/mL; p < 0.001). Furthermore, orexin-A inversely correlated with blood pressure (p = 0.0085), while a direct correlation with eGFR in PKD patients was found. None of the analyzed SNPs showed any association with orexin-A levels in PKD. In conclusion, our data highlights the emerging role of orexin-A in renal physiology and its potential relevance to PKD. Further research is essential to elucidate the intricate mechanisms underlying orexin-A signaling in renal function and its therapeutic implications for PKD and associated cardiovascular complications.

Knockdown of type 2 orexin receptor in adult mouse testis potentiates testosterone production and germ cell proliferation.

Orexins (OXs) are neuropeptides which regulate various physiological processes. OXs exist in two different forms, mainly orexin A (OXA) and orexin B (OXB) and their effects are mediated via OX1R and OX2R. Presence of OXB and OX2R in mouse testis is also reported. However, the role of OXB/OX2R in the male gonad remains unexplored. Herein we investigated the role of OXB/OX2R system in testicular physiology under in vivo and ex vivo conditions. Adult mice were given a single dose of bilateral intratesticular injection of siRNA targeting OX2R and were sacrificed 96 h post-injection. OX2R-knockdown potentiated serum and intratesticular testosterone levels with up-regulation in the expressions of major steroidogenic proteins. Germ cell proliferation also increased in siRNA-treated mice. Results of the ex vivo experiment also supported the findings of the in vivo study. In conclusion, OX2R may regulate testosterone production and thereby control the fine-tuning between steroidogenesis and germ cell dynamics.

Single cell transcriptomics of cerebrospinal fluid cells from patients with recent-onset narcolepsy.

Narcolepsy is a rare cause of hypersomnolence and may be associated or not with cataplexy, i.e. sudden muscle weakness. These forms are designated narcolepsy-type 1 (NT1) and -type 2 (NT2), respectively. Notable characteristics of narcolepsy are that most patients carry the HLA-DQB1*06:02 allele and NT1-patients have strongly decreased levels of hypocretin-1 (synonym orexin-A) in the cerebrospinal fluid (CSF). The pathogenesis of narcolepsy is still not completely understood but the strong HLA-bias and increased frequencies of CD4+ T cells reactive to hypocretin in the peripheral blood suggest autoimmune processes in the hypothalamus. Here we analyzed the transcriptomes of CSF-cells from twelve NT1 and two NT2 patients by single cell RNAseq (scRNAseq). As controls, we used CSF cells from patients with multiple sclerosis, radiologically isolated syndrome, and idiopathic intracranial hypertension. From 27,255 CSF cells, we identified 20 clusters of different cell types and found significant differences in three CD4+ T cell and one monocyte clusters between narcolepsy and multiple sclerosis patients. Over 1000 genes were differentially regulated between patients with NT1 and other diseases. Surprisingly, the most strongly upregulated genes in narcolepsy patients as compared to controls were coding for the genome-encoded MTRNR2L12 and MTRNR2L8 peptides, which are homologous to the mitochondria-encoded HUMANIN peptide that is known playing a role in other neurological diseases including Alzheimer's disease.

Characterization of a putative orexin receptor in Ciona intestinalis sheds light on the evolution of the orexin/hypocretin system in chordates.

Tunicates are evolutionary model organisms bridging the gap between vertebrates and invertebrates. A genomic sequence in Ciona intestinalis (CiOX) shows high similarity to vertebrate orexin receptors and protostome allatotropin receptors (ATR). Here, molecular phylogeny suggested that CiOX is divergent from ATRs and human orexin receptors (hOX1/2). However, CiOX appears closer to hOX1/2 than to ATR both in terms of sequence percent identity and in its modelled binding cavity, as suggested by molecular modelling. CiOX was heterologously expressed in a recombinant HEK293 cell system. Human orexins weakly but concentration-dependently activated its Gq signalling (Ca2+ elevation), and the responses were inhibited by the non-selective orexin receptor antagonists TCS 1102 and almorexant, but only weakly by the OX1-selective antagonist SB-334867. Furthermore, the 5-/6-carboxytetramethylrhodamine (TAMRA)-labelled human orexin-A was able to bind to CiOX. Database mining was used to predict a potential endogenous C. intestinalis orexin peptide (Ci-orexin-A). Ci-orexin-A was able to displace TAMRA-orexin-A, but not to induce any calcium response at the CiOX. Consequently, we suggested that the orexin signalling system is conserved in Ciona intestinalis, although the relevant peptide-receptor interaction was not fully elucidated.

A vertebrate family without a functional Hypocretin/Orexin arousal system.

The Hypocretin/Orexin signaling pathway suppresses sleep and promotes arousal, whereas the loss of Hypocretin/Orexin results in narcolepsy, including the involuntary loss of muscle tone (cataplexy).1 Here, we show that the South Asian fish species Chromobotia macracanthus exhibits a sleep-like state during which individuals stop swimming and rest on their side. Strikingly, we discovered that the Hypocretin/Orexin system is pseudogenized in C. macracanthus, but in contrast to Hypocretin-deficient mammals, C. macracanthus does not suffer from sudden behavioral arrests. Similarly, zebrafish mutations in hypocretin/orexin show no evident signs of cataplectic-like episodes. Notably, four additional species in the Botiidae family also lack a functional Hypocretin/Orexin system. These findings identify the first vertebrate family that does not rely on a functional Hypocretin/Orexin system for the regulation of sleep and arousal.

A direct estrogenic involvement in the expression of human hypocretin.

Hypocretin is synthesized exclusively in the hypothalamus and distributes inputs to several areas of the brain, which may play an important role in depression. Our previous study showed that hypocretin-1 was increased in the lateral hypothalamus in female patients with depression compared to female controls. Estrogen acts through estrogen receptor (ER)α and ERß. We studied the possibility of a direct action of estrogen receptors on the expression of human hypocretin. We found that hypocretin-1 plasma levels were significantly higher in female patients with depression than in female controls. Female depression estrogen receptors and hypocretin are colocalized in the human lateral hypothalamus, PC12, and SK-N-SH cells. The estrogen receptor response elements (ERE) that exist in the hypocretin promoter region may directly regulate the gene expression of hypocretin. The synchronicity of change of hypocretin and estradiol both in hypothalamus and plasma was verified in female rats. In the presence of estradiol, specific binding occurs between the recombinant human ER and hypocretin-ERE. Expression of ER combined with estradiol repressed hypocretin promoter activity via the ERE. In conclusion, we found that estradiol may directly affect hypocretin neurons in the human hypothalamus via ER binding to the hypocretin-ERE, which may lead to the sex-specific pathogenesis of depression.

Embryonic ethanol exposure and optogenetic activation of hypocretin neurons stimulate similar behaviors early in life associated with later alcohol consumption.

The initiation of alcohol use early in life is one of the strongest predictors of developing a future alcohol use disorder. Clinical studies have identified specific behaviors during early childhood that predict an increased risk for excess alcohol consumption later in life. These behaviors, including increased hyperactivity, anxiety, novelty-seeking, exploratory behavior, impulsivity, and alcohol-seeking, are similarly stimulated in children and adolescent offspring of mothers who drink alcohol during pregnancy. Here we tested larval zebrafish in addition to young pre-weanling rats and found this repertoire of early behaviors along with the overconsumption of alcohol during adolescence to be increased by embryonic ethanol exposure. With hypocretin/orexin (Hcrt) neurons known to be stimulated by ethanol and involved in mediating these alcohol-related behaviors, we tested their function in larval zebrafish and found optogenetic activation of Hcrt neurons to stimulate these same early alcohol-related behaviors and later alcohol intake, suggesting that these neurons have an important role in producing these behaviors. Together, these results show zebrafish to be an especially useful animal model for investigating the diverse neuronal systems mediating behavioral changes at young ages that are produced by embryonic ethanol exposure and predict an increased risk for developing alcohol use disorder.

Transient targeting of hypothalamic orexin neurons alleviates seizures in a mouse model of epilepsy.

Lateral hypothalamic (LH) hypocretin/orexin neurons (HONs) control brain-wide electrical excitation. Abnormally high excitation produces epileptic seizures, which affect millions of people and need better treatments. HON population activity spikes from minute to minute, but the role of this in seizures is unknown. Here, we describe correlative and causal links between HON activity spikes and seizures. Applying temporally-targeted HON recordings and optogenetic silencing to a male mouse model of acute epilepsy, we found that pre-seizure HON activity predicts and controls the electrophysiology and behavioral pathology of subsequent seizures. No such links were detected for HON activity during seizures. Having thus defined the time window where HONs influence seizures, we targeted it with LH deep brain stimulation (DBS), which inhibited HON population activity, and produced seizure protection. Collectively, these results uncover a feature of brain activity linked to seizures, and demonstrate a proof-of-concept treatment that controls this feature and alleviates epilepsy.

Bi-allelic variants in HCRT cause autosomal recessive narcolepsy.

Narcolepsy with cataplexy is a complex disease with both genetic and environmental risk factors. To gain further insight into the homozygous HCRT-related narcolepsy, we present a case series of five patients from two consanguineous families, each harboring a novel homozygous variant of HCRT c.17_18del. All affected individuals exhibited severe cataplexy accompanied by narcolepsy symptoms during infancy. Additionally, cataplexy symptoms improved or disappeared in the majority of patients over time. Pathogenic variants in HCRT cause autosomal recessive narcolepsy with cataplexy. Genetic testing of the HCRT gene should be conducted in specific subgroups of narcolepsy, particularly those with early onset, familial cases, and a predominantly cataplexy phenotype.

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.

The transcriptomics profiling of blood CD4 and CD8 T-cells in narcolepsy type I.

Background: Narcolepsy Type I (NT1) is a rare, life-long sleep disorder arising as a consequence of the extensive destruction of orexin-producing hypothalamic neurons. The mechanisms involved in the destruction of orexin neurons are not yet elucidated but the association of narcolepsy with environmental triggers and genetic susceptibility (strong association with the HLA, TCRs and other immunologically-relevant loci) implicates an immuno-pathological process. Several studies in animal models and on human samples have suggested that T-cells are the main pathogenic culprits. Methods: RNA sequencing was performed on four CD4 and CD8 T-cell subsets (naive, effector, effector memory and central memory) sorted by flow cytometry from peripheral blood mononuclear cells (PBMCs) of NT1 patients and HLA-matched healthy donors as well as (age- and sex-) matched individuals suffering from other sleep disorders (OSD). The RNAseq analysis was conducted by comparing the transcriptome of NT1 patients to that of healthy donors and other sleep disorder patients (collectively referred to as the non-narcolepsy controls) in order to identify NT1-specific genes and pathways. Results: We determined NT1-specific differentially expressed genes, several of which are involved in tubulin arrangement found in CD4 (TBCB, CCT5, EML4, TPGS1, TPGS2) and CD8 (TTLL7) T cell subsets, which play a role in the immune synapse formation and TCR signaling. Furthermore, we identified genes (GZMB, LTB in CD4 T-cells and NLRP3, TRADD, IL6, CXCR1, FOXO3, FOXP3 in CD8 T-cells) and pathways involved in various aspects of inflammation and inflammatory response. More specifically, the inflammatory profile was identified in the "naive" subset of CD4 and CD8 T-cell. Conclusion: We identified NT1-specific differentially expressed genes, providing a cell-type and subset specific catalog describing their functions in T-cells as well as their potential involvement in NT1. Several genes and pathways identified are involved in the formation of the immune synapse and TCR activation as well as inflammation and the inflammatory response. An inflammatory transcriptomic profile was detected in both "naive" CD4 and CD8 T-cell subsets suggesting their possible involvement in the development or progression of the narcoleptic process.

Deficiency of orexin signaling during sleep is involved in abnormal REM sleep architecture in narcolepsy.

Narcolepsy is a sleep disorder caused by deficiency of orexin signaling. However, the neural mechanisms by which deficient orexin signaling causes the abnormal rapid eye movement (REM) sleep characteristics of narcolepsy, such as cataplexy and frequent transitions to REM states, are not fully understood. Here, we determined the activity dynamics of orexin neurons during sleep that suppress the abnormal REM sleep architecture of narcolepsy. Orexin neurons were highly active during wakefulness, showed intermittent synchronous activity during non-REM (NREM) sleep, were quiescent prior to the transition from NREM to REM sleep, and a small subpopulation of these cells was active during REM sleep. Orexin neurons that lacked orexin peptides were less active during REM sleep and were mostly silent during cataplexy. Optogenetic inhibition of orexin neurons established that the activity dynamics of these cells during NREM sleep regulate NREM-REM sleep transitions. Inhibition of orexin neurons during REM sleep increased subsequent REM sleep in "orexin intact" mice and subsequent cataplexy in mice lacking orexin peptides, indicating that the activity of a subpopulation of orexin neurons during the preceding REM sleep suppresses subsequent REM sleep and cataplexy. Thus, these results identify how deficient orexin signaling during sleep results in the abnormal REM sleep architecture characteristic of narcolepsy.

Diurnal Characteristics of the Orexin System Genes and Its Effects on Pathology at Early Stage in 3xTg-AD Mice.

Orexin and its receptors are closely related to the pathogenesis of Alzheimer's disease (AD). Although the expression of orexin system genes under physiological condition has circadian rhythm, the diurnal characteristics of orexin system genes, and its potential role in the pathogenesis in AD are unknown. In the present study, we hope to elucidate the diurnal characteristics of orexin system genes at the early stage of AD, and to investigate its potential role in the development of AD neuropathology. We firstly detected the mRNA levels of orexin system genes, AD risk genes and core clock genes (CCGs) in hypothalamus and hippocampus in 6-month-old male 3xTg-AD mice and C57BL/6J (wild type, WT) control mice, then analyzed diurnal expression profiles of all genes using JTK_CYCLE algorithm, and did the correlation analysis between expression of orexin system genes and AD risk genes or CCGs. In addition, the expression of ß-amyloid protein (Aß) and phosphorylated tau (p-tau) protein were measured. The results showed that the diurnal mRNA expression profiles of PPO, OX1R, OX2R, Bace2, Bmal1, Per1, Per2 and Cry1 in the hypothalamus, and gene expression of OX1R, OX2R, Bace1, Bmal1, Per1 and Cry2 in the hippocampus in 3xTg-AD mice were different from that in WT mice. Furthermore, there is positive correlation between orexin system genes and AD risk genes or CCGs in the brain in 3xTg-AD mice. In addition, the expression of Aß and p-tau in hippocampus in 3xTg-AD mice were significantly increased, and the expression of p-tau is higher in night than in day. These results indicate that the abnormal expression profiles of orexin system genes and its interaction with AD risk genes or CCGs might exert important role in the pathogenesis of AD, which will increase the expression of Aß and p-tau, and accelerate the development of AD.

A series of 7 cases of patients with narcolepsy with hypocretin deficiency without the HLA DQB1*06:02 allele.

STUDY OBJECTIVES: We report data collected from 2 reference European sleep centers on a series of patients with narcolepsy with hypocretin-1 deficiency and absence of the human leukocyte antigens (HLA) DQB1*06:02 allele. METHODS: Clinical data, HLA DQ markers, and cerebrospinal fluid assessments were collected retrospectively from Caucasian patients with a diagnosis of narcolepsy type 1 with cerebrospinal fluid hypocretin-1 deficiency (< 110 pg/ml) and absence of the HLA DQB1*06:02 allele, with follow-up with at least 1 visit within the last 4 years, consecutively admitted to 2 European sleep centers (Lugano, Switzerland and Montpellier, France). RESULTS: Seven patients (3 of 29 patients in Lugano and 4 of 328 in Montpellier) were diagnosed with narcolepsy with hypocretin-1 deficiency and absence of HLA DQB1*06:02 (ie, 2% of patients with narcolepsy type 1). Regarding the HLA-DQB1 genotyping, 4 cases were positive for HLA DQB1*03:01, 1 for DQB1*03:02, and 3 for DQB1*02:01. Three patients had atypical cataplexy and 1 had no cataplexy. Only 2 patients had both a mean sleep latency of less than 8 minutes and more than 2 sleep onset rapid eye movement periods on the Multiple Sleep Latency Test, indicative of a less severe condition. CONCLUSIONS: Although rare, this series of 7 cases confirms that hypocretin-deficient narcolepsy should not be excluded in the absence of HLA DQB1*06:02, especially if patients are carriers of other high-risk HLA-DQB1 alleles (DQB1*03:01, *03:02, *02:01). These data support the hypothesis that narcolepsy type 1 is a wider disease spectrum linked to the loss of hypocretin peptide. CITATION: Miano S, Barateau L, De Pieri M, et al. A series of 7 cases of patients with narcolepsy with hypocretin deficiency without the HLA DQB1*06:02 allele. J Clin Sleep Med. 2023;19(12):2053-2057.

Anterior cingulate cortex orexin signaling mediates early-life stress-induced social impairment in females.

Early-life stress has long-term impacts on the structure and function of the anterior cingulate cortex (ACC), and raises the risk of adult neuropsychiatric disorders including social dysfunction. The underlying neural mechanisms, however, are still uncertain. Here, we show that, in female mice, maternal separation (MS) during the first three postnatal weeks results in social impairment accompanied with hypoactivity in pyramidal neurons (PNs) of the ACC. Activation of ACC PNs ameliorates MS-induced social impairment. Neuropeptide Hcrt, which encodes hypocretin (orexin), is the top down-regulated gene in the ACC of MS females. Activating ACC orexin terminals enhances the activity of ACC PNs and rescues the diminished sociability observed in MS females via an orexin receptor 2 (OxR2)-dependent mechanism. Our results suggest orexin signaling in the ACC is critical in mediating early-life stress-induced social impairment in females.

Orexin pathway in Parkinson's disease: a review.

Parkinson's disease (PD) is a progressive neurodegenerative disease (NDD) caused by dopaminergic neuron degeneration in the substantia nigra (SN). Orexin is a neuropeptide that plays a role in the pathogenesis of PD. Orexin has neuroprotective properties in dopaminergic neurons. In PD neuropathology, there is also degeneration of orexinergic neurons in the hypothalamus, in addition to dopaminergic neurons. However, the loss of orexinergic neurons in PD began after the degeneration of dopaminergic neurons. Reduced activity of orexinergic neurons has been linked to developing and progressing motor and non-motor symptoms in PD. In addition, the dysregulation of the orexin pathway is linked to the development of sleep disorders. The hypothalamic orexin pathway regulates various aspects of PD neuropathology at the cellular, subcellular, and molecular levels. Finally, non-motor symptoms, particularly insomnia and disturbed sleep, promote neuroinflammation and the accumulation of neurotoxic proteins as a result of defects in autophagy, endoplasmic reticulum (ER) stress, and the glymphatic system. As a result, this review aimed to highlight the potential role of orexin in PD neuropathology.

Immortal orexin cell transplants restore motor-arousal synchrony during cataplexy.

Waking behaviors such as sitting or standing require suitable levels of muscle tone. But it is unclear how arousal and motor circuits communicate with one another so that appropriate motor tone occurs during wakefulness. Cataplexy is a peculiar condition in which muscle tone is involuntarily lost during normal periods of wakefulness. Cataplexy therefore provides a unique opportunity for identifying the signaling mechanisms that synchronize motor and arousal behaviors. Cataplexy occurs when hypothalamic orexin neurons are lost in narcolepsy; however, it is unclear if motor-arousal decoupling in cataplexy is directly or indirectly caused by orexin cell loss. Here, we used genomic, proteomic, chemogenetic, electrophysiological, and behavioral assays to determine if grafting orexin cells into the brain of cataplectic (i.e., orexin-/-) mice restores normal motor-arousal behaviors by preventing cataplexy. First, we engineered immortalized orexin cells and found that they not only produce and release orexin but also exhibit a gene profile that mimics native orexin neurons. Second, we show that engineered orexin cells thrive and integrate into host tissue when transplanted into the brain of mice. Next, we found that grafting only 200-300 orexin cells into the dorsal raphe nucleus-a region densely innervated by native orexin neurons-reduces cataplexy. Last, we show that real-time chemogenetic activation of orexin cells restores motor-arousal synchrony by preventing cataplexy. We suggest that orexin signaling is critical for arousal-motor synchrony during wakefulness and that the dorsal raphe plays a pivotal role in coupling arousal and motor behaviors.