Heteroaryl derivatives of suvorexant as OX1R selective PET ligand candidates: Cu-mediated 18F-fluorination of boroxines, in vitro and initial in vivo evaluation.

BACKGROUND: The orexin receptor (OXR) plays a role in drug addiction and is aberrantly expressed in colorectal tumors. Subtype-selective OXR PET ligands suitable for in vivo use have not yet been reported. This work reports the development of 18F-labeled OXR PET ligand candidates derived from the OXR antagonist suvorexant and the OX1R-selective antagonist JH112. RESULTS: Computational analysis predicted that fluorine substitution (1e) and introduction of the fluorobenzothiazole scaffold (1f) would be suitable for maintaining high OX1R affinity. After multi-step synthesis of 1a-1f, in vitro OXR binding studies confirmed the molecular dynamics calculations and revealed single-digit nanomolar OX1R affinities for 1a-f, ranging from 0.69 to 2.5 nM. The benzothiazole 1f showed high OX1R affinity (Ki = 0.69 nM), along with 77-fold subtype selectivity over OX2R. Cu-mediated 18F-fluorination of boroxine precursors allowed for a shortened reaction time of 5 min to provide the non-selective OXR ligand [18F]1c and its selective OX1R congener [18F]1f in activity yields of 14% and 22%, respectively, within a total synthesis time of 52-76 min. [18F]1c and [18F]1f were stable in plasma and serum in vitro, with logD7.4 of 2.28 ([18F]1c) and 2.37 ([18F]1f), and high plasma protein binding of 66% and 77%, respectively. Dynamic PET imaging in rats showed similar brain uptake of [18F]1c (0.17%ID/g) and [18F]1f (0.15%ID/g). However, preinjection of suvorexant did not significantly block [18F]1c or [18F]1f uptake in the rat brain. Pretreatment with cyclosporine A to study the role of P-glycoprotein (P-gp) in limiting brain accumulation moderately increased brain uptake of [18F]1c and [18F]1f. Accordingly, in vitro experiments demonstrated that the P-gp inhibitor zosuquidar only moderately inhibited polarized, basal to apical transport of 1c (p < 0.05) and had no effect on the transport of 1f, indicating that P-gp does not play a relevant role in brain accumulation of [18F]1c and [18F]1f in vivo. CONCLUSIONS: The in vitro and in vivo results of [18F]1c and [18F]1f provide a solid basis for further development of suitable OXR PET ligands for brain imaging.

The restraint stress-induced antinociceptive effects decreased by antagonism of both orexin receptors within the CA1 region of the hippocampus.

Studies have indicated that stress-related symptoms can lead to hormonal and neural changes, affecting the pain threshold and nociceptive behaviors. The precise role of orexin receptors (OX1r and OX2r) in stress-induced analgesia (SIA) remains an inquiry yet to be comprehensively elucidated. The current investigation aimed to assess the impact of acute immobilization restraint stress on pain-related behavioral responses after administering antagonists targeting OX1r and OX2r in a rat model using the tail-flick test. After a period of five to seven days post-stereotaxic surgery in CA1, the baseline tail-flick latency (TFL) was recorded for each animal. Subsequently, rats were unilaterally administered varying doses of the OX1r antagonist (SB334867; 1, 3, 10, and 30 nmol), the OX2r antagonist (TCS OX2 29; 1, 3, 10, and 30 nmol), or a vehicle (0.5 µl solution containing 12% DMSO) through an implanted cannula. Following a 5-min interval, the animals were subjected to a restraint stress (RS) lasting for 3 h. The tail-flick test was conducted after the stress exposure, and the TFLs were assessed at 60-min intervals. The findings of this study revealed that RS elicits antinociceptive responses in the tail-flick test. Microinjection of OX1r and OX2r antagonists into the CA1 attenuated RS-induced analgesia during the tail-flick test. Furthermore, the results underscored the preeminent role of OX2 receptors in modulating SIA. In conclusion, the orexin system localized within the hippocampal CA1 region may, in part, contribute to the manifestation of SIA in the context of acute pain.

Jingqianshu granules mitigates premenstrual depression by regulating orexin signaling.

Introduction: Premenstrual dysphoric disorder (PMDD), a severe form of premenstrual syndrome (PMS), is a serious health disorder that affects patient moods. It is caused by cyclic psychological symptoms and its pathogenesis is still unclear. Abnormalities in the basolateral amygdala (BLA) orexin system, which are important causes of the development of depressive mood, have not been reported in PMDD, so exploring its intrinsic mechanisms is meaningful for enriching the pathomechanisms of PMDD. Methods: High performance liquid chromatography was used for the determination of the active ingredients of Jingqianshu granules. Developing a rat model of premenstrual depression using the forced swimming test (FST). The experiment consisted of two parts. In Part 1, the rats were divided into the control group, the model group, the model + Jingqianshu group, and the model + fluoxetine group. The FST, open field test, and elevated plus maze test, were used to assess the behavior of the rats as well as to evaluate the effect of drug intervention. Immunofluorescence and RT-qPCR were used to detect the expression of orexin and its receptors OX1R and OX2R genes and proteins. The expression of Toll-like receptor 4, nuclear factor kappa-B, tumor necrosis factor-α, interleukin 6, and interleukin-1ß in the BLA brain region was detected by Western-Blot. In part 2, the rats were injected intracerebrally with orexin-A. Observe the behavioral activities of rats in the control group, model group, and model+orexin-A group. Immunofluorescence was used to detect microglia in the BLA area of rats, and the expression levels of the above inflammatory factors were detected by Western-Blot. Results: The five components of Jingqianshu granules are: paeoniflorin, erulic acid, liquiritin, hesperidin, and paeonol. During the estrous cycle, rats exhibited depressive-like behavior during the non-receptive phase of the behavioral test, which disappeared during the receptive phase. Immunofluorescence and RT-qPCR showed reduced gene and protein expression of orexin, OX1R, and OX2R in the BLA region of rats in the model group.WB showed elevated levels of inflammatory factors. All returned to control levels after drug treatment. In part 2, injection of orexin-A into the BLA brain region of model rats resulted in reduced immunoreactivity of microglia and decreased expression levels of inflammatory factors. Discussion: Jianqianshu granules can achieve the purpose of treating premenstrual depression by regulating orexin-mediated inflammatory factors, which provides a new idea for further research on the pathogenesis of PMDD. However, the current study is still preliminary and the pathogenesis of PMDD is complex. Therefore, more in-depth exploration is needed.

The protective effects of orexin-A in alleviating cell senescence against interleukin-1ß (IL-1ß) in chondrocytes.

Osteoarthritis (OA) is one of the most important causes of global disability, and dysfunction of chondrocytes is an important risk factor. The treatment of OA is still a challenge. Orexin-A is a hypothalamic peptide, and its effects in OA are unknown. In this study, we found that exposure to interleukin-1ß (IL-1ß) reduced the expression of orexin-2R, the receptor of orexin-A in TC-28a2 chondrocytes. Importantly, the senescence-associated ß-galactosidase (SA-ß-gal) staining assay demonstrated that orexin-A treatment ameliorates IL-1ß-induced cellular senescence. Importantly, the presence of IL-1ß significantly reduced the telomerase activity of TC-28a2 chondrocytes, which was rescued by orexin-A. We also found that orexin-A prevented IL-1ß-induced increase in the levels of Acetyl-p53 and the expression of p21. It is shown that orexin-A mitigates IL-1ß-induced reduction of sirtuin 3 (SIRT3). Silencing of SIRT3 abolished the protective effects of orexin-A against IL-1ß-induced cellular senescence. These results imply that orexin-A might serve as a promising therapeutic agent for OA.

Orexin B protects dopaminergic neurons from 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine-induced neurotoxicity associated with reduced extracellular signal-regulated kinase phosphorylation.

BACKGROUND: The loss of dopaminergic (DA) neurons in the substantia nigra pars compacta (SNpc) is a major pathological hallmark of Parkinson's disease (PD). Orexin B (OXB) has been reported to promote the growth of DA neurons. However, the roles of OXB in the degeneration of DA neurons still remained not fully clear. METHODS: An in vivo PD model was constructed by administrating 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) in mice. Pole test was performed to investigate the motor function of mice and the number of DA neurons was detected by immunofluorescence (IF). A PD cell model was established by treating SH-SY5Y cells with 1-methyl-4-phenylpyridinium (MPP+). OXB was added to the culture medium 2 h after MPP + treatment. Microscopic analysis was carried out to investigate the function of OXB in the cell model of PD 24 h after MPP + challenge. RNA-Seq analysis of the PD cell model was performed to explore the possible mechanisms. Western blot was used to detect the phosphorylation levels of extracellular signal-regulated kinase (ERK). RESULTS: OXB significantly decreased the DA neurons death caused by MPTP, alleviated MPP+-induced neurotoxicity in SH-SY5Y cells, and robustly enhanced the weight and motor ability of PD mice. Besides, RNA-Seq analysis demonstrated that the mitogen-activated protein kinase (MAPK) pathway was involved in the pathology of PD. Furthermore, MPP + led to increased levels of phosphorylation of ERK (p-ERK), OXB treatment significantly decreased the levels of p-ERK in MPP+-treated SH-SY5Y cells. CONCLUSIONS: This study demonstrated that OXB exerts a neuroprotective role associated with reduced ERK phosphorylation in the PD model. This suggests that OXB may have therapeutic potential for treatment of PD.

The differential effects of blocking retinal orexin receptors on the expression of retinal c-fos and hypothalamic Vip, PACAP, Bmal1, and c-fos in Male Wistar Rats.

Orexin A and B (OXA and OXB) and their receptors are expressed in the majority of retinal neurons in humans, rats, and mice. Orexins modulate signal transmission between the different layers of the retina. The suprachiasmatic nucleus (SCN) and the retina are central and peripheral components of the body's biological clocks; respectively. The SCN receives photic information from the retina through the retinohypothalamic tract (RHT) to synchronize bodily functions with environmental changes. In present study, we aimed to investigate the impact of inhibiting retinal orexin receptors on the expression of retinal Bmal1 and c-fos, as well as hypothalamic c-fos, Bmal1, Vip, and PACAP at four different time-points (Zeitgeber time; ZT 3, 6, 11, and ZT-0). The intravitreal injection (IVI) of OX1R antagonist (SB-334867) and OX2R antagonist (JNJ-10397049) significantly up-regulated c-fos expression in the retina. Additionally, compared to the control group, the combined injection of SB-334867 and JNJ-10397049 showed a greater increase in retinal expression of this gene. Moreover, the expression of hypothalamic Vip and PACAP was significantly up-regulated in both the SB-334867 and JNJ-10397049 groups. In contrast, the expression of Bmal1 was down-regulated. Furthermore, the expression of hypothalamic c-fos was down-regulated in all groups treated with SB-334867 and JNJ-10397049. Additionally, the study demonstrated that blocking these receptors in the retina resulted in alterations in circadian rhythm parameters such as mesor, amplitude, and acrophase. Finally, it affected the phase of gene expression rhythms in both the retina and hypothalamus, as identified through cosinor analysis and the zero-amplitude test. This study represents the initial exploration of how retinal orexin receptors influence expression of rhythmic genes in the retina and hypothalamus. These findings could provide new insights into how the retina regulates the circadian rhythm in both regions and illuminate the role of the orexinergic system expression within the retina.

A Review of Recent Advances in the Management of Alzheimer's Disease.

Alzheimer's disease (AD) is the most common neurodegenerative condition and a form of dementia encountered in medical practice. Despite many proposed and attempted treatments, this disease remains a major puzzle in the public health systems worldwide. The initial part of this article provides an overview and illustration of the primary mechanisms responsible for neuronal damage in AD. Subsequently, it offers a critical evaluation of the most noteworthy studies on pharmacological therapy for AD and outlines recent advancements and novel approaches to managing this condition. Main properties, categorization, Food and Drug Administration (FDA) status, mechanisms of action, benefits, and common side effects of the classical and the most recently proposed pharmacological treatments for AD are described. The conventional pharmacological agents revised comprise cholinesterase inhibitors, monoclonal antibodies, and other therapies, such as memantine, valproic acid, and rosiglitazone. The innovative reviewed pharmacological agents comprise the monoclonal antibodies: donanemab, gantenerumab, solanezumab, bapineuzumab, crenezumab, and semorinemab. Nutritional supplements such as alpha-tocopherol (vitamin E) and caprylidene are also revised. Tau and amyloid-targeting treatments include methylthioninium moiety (MT), leuco-methylthioninium bis (LMTM), an oxidized form of MT, and tramiprosate, which inhibits the beta-amyloid (Aß) monomer aggregation into toxic oligomers. Antidiabetic and anti-neuroinflammation drugs recently proposed for AD treatment are discussed. The antidiabetic drugs include NE3107, an anti-inflammatory and insulin sensitizer, and the diabetes mainstream drug metformin. The anti-neuroinflammatory AD therapies include the use of sodium oligomannate (GV-971), infusions with intravenous immunoglobulin aiming to decrease plasma levels of the constituents of Aß plaques, and masitinib, a tyrosine kinase inhibitor that impacts mast and microglia cells. Additional anti-inflammatory agents being currently tested in phase-2 clinical trials, such as atomoxetine (selective norepinephrine reuptake inhibitor), losartan (angiotensin 2 receptor agonist), genistein (anti-inflammatory isoflavone neuroprotective agent), trans-resveratrol (polyphenol antioxidant plant estrogen), and benfotiamine (synthetic thiamine precursor), were reviewed. Lastly, drugs targeting Alzheimer's-associated symptoms, such as brexpiprazole (serotonin dopamine activity modulator) and suvorexant (orexin receptor antagonist), respectively, used for agitation and insomnia in AD patients, are reviewed. As experimental investigations and clinical research progress, there is a possibility that a combination of newly tested medications and traditional ones may emerge as a promising treatment option for AD in the future.

Therapeutic effects of orexin-A in sepsis-associated encephalopathy in mice.

BACKGROUND: Sepsis-associated encephalopathy (SAE) causes acute and long-term cognitive deficits. However, information on the prevention and treatment of cognitive dysfunction after sepsis is limited. The neuropeptide orexin-A (OXA) has been shown to play a protective role against neurological diseases by modulating the inflammatory response through the activation of OXR1 and OXR2 receptors. However, the role of OXA in mediating the neuroprotective effects of SAE has not yet been reported. METHODS: A mouse model of SAE was induced using cecal ligation perforation (CLP) and treated via intranasal administration of exogenous OXA after surgery. Mouse survival, in addition to cognitive and anxiety behaviors, were assessed. Changes in neurons, cerebral edema, blood-brain barrier (BBB) permeability, and brain ultrastructure were monitored. Levels of pro-inflammatory factors (IL-1ß, TNF-α) and microglial activation were also measured. The underlying molecular mechanisms were investigated by proteomics analysis and western blotting. RESULTS: Intranasal OXA treatment reduced mortality, ameliorated cognitive and emotional deficits, and attenuated cerebral edema, BBB disruption, and ultrastructural brain damage in mice. In addition, OXA significantly reduced the expression of the pro-inflammatory factors IL-1ß and TNF-α, and inhibited microglial activation. In addition, OXA downregulated the expression of the Rras and RAS proteins, and reduced the phosphorylation of P-38 and JNK, thus inhibiting activation of the MAPK pathway. JNJ-10,397,049 (an OXR2 blocker) reversed the effect of OXA, whereas SB-334,867 (an OXR1 blocker) did not. CONCLUSION: This study demonstrated that the intranasal administration of moderate amounts of OXA protects the BBB and inhibits the activation of the OXR2/RAS/MAPK pathway to attenuate the outcome of SAE, suggesting that OXA may be a promising therapeutic approach for the management of SAE.

Transcranial direct current stimulation promotes angiogenesis and improves neurological function via the OXA-TF-AKT/ERK signaling pathway in traumatic brain injury.

Traumatic brain injury (TBI) and its resulting complications pose a major challenge to global public health, resulting in increased rates of disability and mortality. Cerebrovascular dysfunction is nearly universal in TBI cases and is closely associated with secondary injury after TBI. Transcranial direct current stimulation (tDCS) shows great potential in the treatment of TBI; however, the exact mechanism remains elusive. In this study, we performed in vivo and in vitro experiments to explore the effects and mechanisms of tDCS in a controlled cortical impact (CCI) rat model simulating TBI. In vivo experiments show that tDCS can effectively reduce brain tissue damage, cerebral edema and neurological deficits. The potential mechanism may be that tDCS improves the neurological function of rats by increasing orexin A (OXA) secretion, upregulating the TF-AKT/ERK signaling pathway, and promoting angiogenesis at the injury site. Cellular experiments showed that OXA promoted HUVEC migration and angiogenesis, and these effects were counteracted by the ERK1/2 inhibitor LY3214996. The results of Matrigel experiment in vivo showed that TNF-a significantly reduced the ability of HUVEC to form blood vessels, but OXA could rescue the effect of TNF-a on the ability of HUVEC to form blood vessels. However, LY3214996 could inhibit the therapeutic effect of OXA. In summary, our preliminary study demonstrates that tDCS can induce angiogenesis through the OXA-TF-AKT/ERK signaling pathway, thereby improving neurological function in rats with TBI.

Orexin-A mediates glioblastoma proliferation inhibition by increasing ferroptosis triggered by unstable iron pools and GPX4 depletion.

Glioblastoma (GBM) represents a prevalent form of primary malignant tumours in the central nervous system, but the options for effective treatment are extremely limited. Ferroptosis, as the most enriched programmed cell death process in glioma, makes a critical difference in glioma progression. Consequently, inducing ferroptosis has become an appealing strategy for tackling gliomas. Through the utilization of multi-omics sequencing data analysis, flow cytometry, MDA detection and transmission electron microscopy, the impact of orexin-A on ferroptosis in GBM was assessed. In this report, we provide the first evidence that orexin-A exerts inhibitory effects on GBM proliferation via the induction of ferroptosis. This induction is achieved by instigating an unsustainable increase in iron levels and depletion of GPX4. Moreover, the regulation of TFRC, FTH1 and GPX4 expression through the targeting of NFE2L2 appears to be one of the potential mechanisms underlying orexin-A-induced ferroptosis.

A Comparative Analysis of Orexins in the Physio-Pathological Processes of the Male Genital Tract: New Challenges? A Review.

Orexins A (OXA) and B (OXB) and their specific receptors, receptor 1 (OX1R) and 2 (OX2R) for orexins, are hypothalamic peptides involved in orchestrating several functions in the central nervous system and peripheral organs, including sleep, excitement, nutrition, reward, circadian rhythm, anxiety, cognition, and reproduction. The aim of this narrative review is, in particular, to speculate the role of orexins in the male genital tract of animal species and human beings. The experimental evidence collected in recent years assumed that in the testes of the animal species here described, orexins are directly involved in steroidogenesis and spermatogenesis regulation. In the epididymis, these peptides are locally synthesized, thus suggesting their role governing the fertilizing capability of the immature male gamete. In addition to playing a physiological role, orexins are involved in numerous inflammatory and/or neoplastic pathologies too. The expression of the orexinergic system in prostate cancer suggests that they might play a potential therapeutic function. Overall, the future directions of this literature review allow us to hypothesize a role of the orexinergic complex not only as a marker for the diagnosis of certain tumors affecting the male genital tract but also for the treatment of hypo/infertility condition.

Potential immunological triggers for narcolepsy and idiopathic hypersomnia: Real-world insights on infections and influenza vaccinations.

OBJECTIVE: It is hypothesized that narcolepsy type 1 (NT1) develops in genetically susceptible people who encounter environmental triggers leading to immune-mediated hypocretin-1 deficiency. The pathophysiologies of narcolepsy type 2 (NT2) and idiopathic hypersomnia (IH) remain unknown. The main aim of this study was to collect all reported immunological events before onset of a central disorder of hypersomnolence. METHODS: Medical records of 290 people with NT1, and 115 with NT2 or IH were retrospectively reviewed to extract infection and influenza vaccination history. Prevalence, distribution of immunological events, and time until hypersomnolence onset were compared between NT1 and the combined group of NT2 and IH. RESULTS: Immunological events were frequently reported before hypersomnolence disorder onset across groups. Flu and H1N1 influenza vaccination were more common in NT1, and Epstein-Barr virus and other respiratory and non-respiratory infections in NT2 and IH. Distributions of events were comparable between NT2 and IH. Rapid symptom onset within one month of infection was frequent across groups, especially after flu infection in NT1. Hypersomnolence disorder progression after an immunological event was reported in ten individuals. CONCLUSIONS: Our findings suggest a variety of immunological triggers potentially related to NT1, including H1N1 influenza infection or vaccination, infection with other flu types, and other respiratory and non-respiratory infections. Frequent reports of immunological events (other than those reported in NT1) immediately prior to the development of NT2 and IH support the specificity of triggers for NT1, and open important new research avenues into possible underlying immunological mechanisms in NT2 and IH.

Mutual Effects of Orexin and Bone Morphogenetic Proteins on Catecholamine Regulation Using Adrenomedullary Cells.

Orexins are neuronal peptides that play a prominent role in sleep behavior and feeding behavior in the central nervous system, though their receptors also exist in peripheral organs, including the adrenal gland. In this study, the effects of orexins on catecholamine synthesis in the rat adrenomedullary cell line PC12 were investigated by focusing on their interaction with the adrenomedullary bone morphogenetic protein (BMP)-4. Orexin A treatment reduced the mRNA levels of key enzymes for catecholamine synthesis, including tyrosine hydroxylase (Th), 3,4-dihydroxyphenylalanie decarboxylase (Ddc) and dopamine ß-hydroxylase (Dbh), in a concentration-dependent manner. On the other hand, treatment with BMP-4 suppressed the expression of Th and Ddc but enhanced that of Dbh with or without co-treatment with orexin A. Of note, orexin A augmented BMP-receptor signaling detected by the phosphorylation of Smad1/5/9 through the suppression of inhibitory Smad6/7 and the upregulation of BMP type-II receptor (BMPRII). Furthermore, treatment with BMP-4 upregulated the mRNA levels of OX1R in PC12 cells. Collectively, the results indicate that orexin and BMP-4 suppress adrenomedullary catecholamine synthesis by mutually upregulating the pathway of each other in adrenomedullary cells.

Emerging and future directions of migraine research and treatment.

Despite many migraine-specific treatments that became available over the past 5 years, many patients still suffer from debilitating migraine. Emerging and future directions of migraine research and treatment should consider different aspects including revising the headache diagnostic criteria to reflect disease burden and prognosis, developing biomarkers, including genetic, serum, imaging, and deep phenotyping biomarkers to facilitate personalized medicine for headache treatment. Additionally, research should also emphasize identifying novel treatment targets for drug development. In this chapter, we provide an overview of current studies and controversies in the diagnosis of migraine and available research on potential migraine biomarkers. We also discuss potential treatment targets for migraine, including CGRP, PACAP, orexin, non-µ opioid receptors, nitric oxide, BKCa channel, KATP channel, amylin, TRP channels, prolactin, PAR-2, and other potential targets.

Improvement of autistic-like behaviors in adult rats prenatally exposed to valproic acid through early suppression of orexin receptor.

Introduction: Autism spectrum disorder (ASD) is a disabling psychiatric disease characterized by impairments in communication and social skills. The pathophysiology of autism is complex and not fully known. Considering the incidence of sleep disorders in individuals with ASD and the important role of orexin in sleep, it is possible to hypothesize that an alteration of the orexinergic system could be implicated in the pathogenesis of autism symptoms. The present study was conducted to investigate the effect of suvorexant [dual orexin receptor antagonists (DORAs)] on autism-like behavior in prenatally valproic acid (VPA)-exposed rats]. Methods: Wistar female rats were administered VPA [600 mg/kg, intraperitoneally (i.p.)] or normal saline (10 ml/kg, i.p.; vehicle control) on gestational day 12.5. Thirty-two male offspring were divided into four groups: Control, VPA, Suvorexant+VPA, and VPA+Risperidone. The pups were given suvorexant [20 ml/kg, by mouth/orally (p.o.)] or risperidone (1 ml/kg, p.o.) daily from postnatal day (PND) (40-54). The offspring were tested for repetitive behaviors and cognitive ability with a Y-maze task on PND 55, and social interaction was assessed by play behavior in the open field on PND 56. And anxiety with using the three-chamber social assay on PND 56. Results: In the Y-maze apparatus, spontaneous alteration significantly decreased in the prenatal VPA-treated rats compared to control rats showing autistic-like behavior, and 2-week suvorexant increased the alternation, indicating the beneficial effect of suvorexant. Prenatal treatment with VPA, impaired play behavior (sniffing, grooming, and darting), and increased anxiety-related behavior. Suvorexant treatment attenuated the problems in male offspring's social behavior. Conclusion: Our results showed that suvorexant improved ASD-associated behaviors in the VPA-treated rats, and the orexinergic system may be associated with the pathogenesis of autism symptoms.

Imaging flow cytometry of tumoroids: A new method for studying GPCR expression.

Fluorescence confocal microscopy is commonly used to analyze the regulation membrane proteins expression such as G protein-coupled receptors (GPCRs). With this approach, the internal movement of GPCRs within the cell can be observed with a high degree of resolution. However, these microscopy techniques led to complex and time-consuming analysis and did not allow a large population of events to be sampled. A recent approach termed imaging flow cytometry (IFC), which combines flow cytometry and fluorescence microscopy, had two main advantages to study the regulation of GPCRs expression such as orexins receptors (OXRs): the ability (1) to analyze large numbers of cells and; (2) to visualize cell integrity and fluorescent markers localization. Here, we compare these two technologies using the orexin A (OxA) ligand coupled to rhodamine (OxA-rho) to investigate anti-tumoral OX1R expression in human digestive cancers. IFC has been adapted for cancer epithelial adherent cells and also to 3D cell culture tumoroids which partially mimic tumoral structures. In the absence of specific antibody, expression of OX1R is examined in the presence of OxA-rho. 2D-culture of colon cancer cells HT-29 exhibits a maximum level of OX1R internalization induced by OxA with 19% ± 3% colocalizing to early endosomes. In 3D-culture of HT-29 cells, internalization of OX1R/OxA-rho reached its maximum at 60 min, with 30.7% ± 6.4% of OX1R colocalizing with early endosomes. This is the first application of IFC to the analysis of the expression of a native GPCR, OX1R, in both 2D and 3D cultures of adherent cancer cells.

Orexins and Prostate Cancer: State of the Art and Potential Experimental and Therapeutic Perspectives.

Prostate cancer (PCa) is the second most common cancer in humans. Peptides have recently been used as targeted therapeutics in cancers, due to their extensive multi-functional applications. Two hypothalamic peptides, orexins A (OXA) and B (OXB) and their specific receptors, orexin receptor 1 (OX1R) and 2 (OX2R), orchestrate several biological processes in the central nervous system and peripheral organs. However, in addition to their role in physiological responses, orexins are involved in numerous inflammatory and/or neoplastic pathologies. The presence and expression of orexins in different cancer models, including prostate cancer, and their role in inducing pro- or anti-apoptotic responses in tumor cell lines, suggest that the orexinergic system might have potential therapeutic action or function as a diagnostic marker in PCa. In addition to the traditional animal models for studying human PCa, the canine model might also serve as an additional tool, due to its clinical similarities with human prostate cancer.

Alterations in Appetite-Regulating Hormones in Girls with Central Early or Precocious Puberty.

The prevalence of central precocious puberty (CPP) in girls has increased worldwide and is often associated with obesity in childhood as well as high fat/high glycemic index diets. Evidence suggests that subjects with obesity present with alterations in appetite-regulating hormones. The arcuate and paraventricular nuclei of the hypothalamus are the centers of action of appetite hormones, as well as the location of gonadotropin-releasing hormone (GnRH) neurons, the activation of which results in the onset of puberty. This anatomical proximity raises the question of possible alterations in appetite-regulating hormones in patients with CPP. Furthermore, diet-induced hypothalamic inflammation constitutes a probable mechanism of the pathophysiology of CPP, as well as alterations in appetite-regulating hormones in young children. In this article, we summarize the evidence investigating whether girls with CPP present with alterations in appetite-regulating hormones. We present evidence that leptin concentrations are elevated in girls with CPP, ghrelin concentrations are lower in girls with CPP, nesfatin-1 and orexin-A concentrations are elevated among girls with premature thelarche, and insulin concentrations are increased in girls with early menarche.

Ramelteon and suvorexant for postoperative delirium in elderly patients with esophageal cancer.

BACKGROUND: There is no clear evidence on the prevention of postoperative delirium with pharmacotherapy in elderly patients with esophageal cancer. This retrospective study aimed to evaluate the efficacy of ramelteon and suvorexant in preventing postoperative delirium in this patient group. METHODS: Data on 251 patients who received radical esophagectomy for thoracic esophageal cancer were collected from January 2010 to September 2021. In total, 74 patients did not receive preventive intervention, and 177 received ramelteon and suvorexant. After propensity score matching, the rate of postoperative delirium was compared between the two groups. RESULTS: Seventy-two well-balanced patients in each group demonstrated similar clinical and pathological characteristics. The mean ages of the intervention and control groups were 70.8 and 70.3 years, respectively. All the patients underwent McKeown esophagectomy, and in the volume of intraoperative blood loss or operative time did not significantly differ between the two groups. The incidence rates of postoperative hyperactive delirium were 7% (5/72) in the intervention group and 32% (23/72) in the control group (p < 0.001). No severe adverse event potentially attributable to the intervention drug was observed. The multivariate analysis showed that the use of ramelteon and suvorexant was the only independent protective factor against postoperative delirium (hazard ratio 0.157, 95% CI 0.055-0.448, p < 0.001). CONCLUSIONS: Ramelteon and suvorexant may play an important role in reducing postoperative delirium in elderly patients with esophageal cancer.