Orx System Research Articles

Sex-Specific Effects of Neonatal Stress on Brainstem Expression of Orexin1-Receptors

Introduction and hypothesis: Located within the hypothalamus, orexin-producing neurons (ORX) regulate arousal, feeding, and cardiorespiratory function. Panic disorders (PD) affects ~5% of the North American population. Stress-related “hyperactivation” of the ORX system is a leading hypothesis to explain the pathophysiology of PD. One important sub-group of PD patients show significant respiratory symptoms, including an excessive ventilatory response to CO2 inhalation. Manifestations of PD are sex-specific: the prevalence of PD is ~2 times higher in women compared to men and the intensity of the ventilatory response to CO2 fluctuates with the hormonal cycle. Early life stress is a risk factor for PD and we recently demonstrated that in rats, neonatal maternal separation (NMS) augments the ventilatory response to CO2 (5%; 10 min) of adult females (but not males); this effect peaks with the rise of 17-b estradiol (E2) during proestrus. Because NMS-related enhancement of the hyperventilatory response to CO2 can be prevented by a selective ORX1-antagonist, we tested the hypothesis that sex, hormonal status and NMS interact to influence the expression of ORX-1 receptors in brainstem regions that regulate breathing. Methods: Following birth, pups were either exposed to NMS (3h/day from postnatal days 3 to 12) or raised under standard conditions. Rats were then raised until adulthood (8 weeks); experiments were performed on males, females in proestrus (high E2), and ovariectomised (low E2) females (2 weeks post-surgery). We used in situ hybridization histochemistry to quantify the expression of mRNA for ORX-1 receptors. The ORX-1 receptors signal was revealed by autoradiography emulsion and slides were examined under darkfield microscopy. Expression of ORX-1 receptors was quantified by measuring the pixel density of the hybridization signal in selected brainstem regions that regulate CO2 response. Results: The locus coeruleus (LC) contained the highest expression of ORX-1 receptors amongst the structures studied. In males, NMS was associated with significant decreases in ORX-1 expression in the LC and nucleus of the solitary tract (NTS). By comparison with OVX females, proestrus augmented ORX-1 expression in the LC of controls, but not NMS. By comparison with OVX, proestrus augmented ORX-1 expression in the raphe obscurus, especially in NMS females. Expression of ORX-1 was unchanged in the NTS of females. Conclusion and perspectives: We conclude that sex and the female’s hormonal status are important determinants of ORX-1 expression and that NMS can alter these relationships; however, those effects are structure-specific. In light of the evidence indicating that 5-HT dysfunction contributes to PD, the results observed in the raphe obscurus point to a plausible mechanism. This study was supported by ‘the Québec Heart and Lung Institute Research Foundation’. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Alternative use of suvorexant (Belsomra®) for the prevention of alcohol drinking and seeking in rats with a history of alcohol dependence.

Alcohol use disorder (AUD) is one of the most treatment-resistant medical conditions globally. The orexin (Orx) system regulates diverse physiological processes, including stress, and is a system of interest for the development of pharmaceuticals to treat substance use disorders, particularly AUD. The present study tested the ability of the dual orexin receptor antagonist suvorexant (SUV), marketed by Merck as Belsomra®, for the treatment of insomnia, to decrease alcohol self-administration and the stress-induced reinstatement of alcohol-seeking behavior in male Wistar rats with a history of alcohol dependence. Rats were trained to orally self-administer 10% alcohol (30 min/day for 3 weeks) and were either made dependent via chronic intermittent alcohol vapor exposure (14 h ON, 10 h OFF) for 6 weeks or exposed to air (non-dependent). Starting on week 7, the effect of SUV (0-20 mg/kg, p.o.) was tested on alcohol self-administration at acute abstinence (8 h after vapor was turned OFF) twice weekly. A separate cohort of rats that were prepared in parallel was removed from alcohol vapor exposure and then subjected to extinction training for 14 sessions. Once extinction was achieved, the rats received SUV (0 and 5 mg/kg, p.o.) and were tested for the footshock stress-induced reinstatement of alcohol-seeking behavior. Suvorexant at 5, 10, and 20 mg/kg selectively decreased alcohol intake in dependent rats. Furthermore, 5 mg/kg SUV prevented the stress-induced reinstatement of alcohol-seeking behavior in dependent rats only. These results underscore the significance of targeting the Orx system for the treatment of substance use disorders generally and suggest that repurposing SUV could be an alternative approach for the treatment of AUD.

Acute orexin antagonism selectively modulates anticipatory anxiety in humans: implications for addiction and anxiety

Research indicates that heightened anticipatory anxiety underlies several forms of psychopathology. Anticipatory anxiety can be reliably and objectively measured in the laboratory using the No-Predictable-Unpredictable (NPU) threat paradigm. The NPU paradigm is an ideal research tool for the NIH ‘Fast-Fail’ approach of screening promising compounds and testing human target engagement. Evidence from preclinical studies suggests that the hypocretin/orexin (ORX) hypothalamic neuropeptide system is a potential means for modulating anticipatory anxiety and disrupting stress-related alcohol use. The current study tested this question using a psychophysiological probe of the ORX system in humans. We examined whether a single dose of suvorexant (SUV; 10 mg; dual ORX receptor antagonist) can effectively and selectively target a well-validated human laboratory index of exaggerated anticipatory anxiety using a within-subjects placebo-controlled design. A total of twenty-one volunteers completed two laboratory sessions during acute administration of 10 mg SUV or placebo. Across sessions, we administered the NPU paradigm probing sustained anticipatory anxiety and fear while startle eyeblink was recorded as an index of aversive reactivity. Questionnaires assessing mood states and subjective drug effects were also collected. Results indicated SUV was well-tolerated. Compared with placebo, SUV was associated with decreased startle reactivity during anticipatory anxiety but not fear or no-threat conditions. Therefore, SUV selectively and effectively reduced objective indicators of anticipatory anxiety in humans and engaged our laboratory target of psychopathology. ORX antagonism may be a promising strategy for modulating human anxiety and potentially, stress-related alcohol use.

Cocaine-Seeking Behavior Induced by Orexin A Administration in the Posterior Paraventricular Nucleus of the Thalamus Is Not Long-Lasting: Neuroadaptation of the Orexin System During Cocaine Abstinence.

Hypothalamic orexin (Orx) projections to the paraventricular nucleus of the thalamus (PVT) have received growing interest because of their role in drug-seeking behavior. Using an established model of cocaine dependence (i.e., long access [LgA] to cocaine), we previously showed that OrxA injections in the posterior PVT (pPVT) reinstated extinguished cocaine-seeking behavior in rats after an intermediate period of abstinence (2–3 weeks). Considering the long-lasting nature of drug-seeking behavior, the present study examined whether the priming effect of intra-pPVT OrxA administration was preserved after a period of protracted abstinence (4–5 weeks) in rats that self-administered cocaine under LgA conditions. Furthermore, to better understand whether a history of cocaine dependence affects the Orx system—particularly the hypothalamic Orx↔pPVT connection—the number of Orx-expressing cells in the lateral hypothalamus (LH), dorsomedial hypothalamus (DMH), and perifornical area (PFA) and number of orexin receptor 1 (OrxR1)- and OrxR2-expressing cells in the pPVT were quantified. Orexin A administration in the pPVT induced cocaine-seeking behavior after intermediate abstinence, as reported previously. At protracted abstinence, however, the priming effect of OrxA was absent. A higher number of cells that expressed Orx was observed in the LH/DMH/PFA at both intermediate and protracted abstinence. In the pPVT, the number of OrxR2-expressing cells was significantly higher only at intermediate abstinence, with no changes in the number of OrxR1-expressing cells. These data build on our previous findings that the hypothalamic Orx↔pPVT connection is strongly recruited shortly after cocaine abstinence and demonstrate that the priming effect of OrxA is not long lasting. Furthermore, these findings suggest that throughout abstinence, the Orx↔pPVT connection undergoes neuroadaptive changes, reflected by alterations of the number of OrxR2-expressing cells in the pPVT.

Drug Seeking and Relapse: New Evidence of a Role for Orexin and Dynorphin Co-transmission in the Paraventricular Nucleus of the Thalamus.

The long-lasting vulnerability to relapse remains the main challenge for the successful treatment of drug addiction. Neural systems that are involved in processing natural rewards and drugs of abuse overlap. However, neuroplasticity that is caused by drug exposure may be responsible for maladaptive, compulsive, and addictive behavior. The orexin (Orx) system participates in regulating numerous physiological processes, including energy metabolism, arousal, and feeding, and is recruited by drugs of abuse. The Orx system is differentially recruited by drugs and natural rewards. Specifically, we found that the Orx system is more engaged by drugs than by non-drugs, such as sweetened condensed milk (SCM) or a glucose saccharin solution (GSS), in an operant model of reward seeking. Although stimuli (S+) that are conditioned to cocaine (COC), ethanol, and SCM/GSS equally elicited reinstatement, Orx receptor blockade reversed conditioned reinstatement for drugs vs. non-drugs. Moreover, the hypothalamic recruitment of Orx cells was greater in rats that were tested with the COC S+ vs. SCM S+, indicating of a preferential role for the Orx system in perseverative, compulsive-like COC seeking and not behavior that is motivated by palatable food. Accumulating evidence indicates that the paraventricular nucleus of the thalamus (PVT), which receives major Orx projections, mediates drug-seeking behavior. All Orx neurons contain dynorphin (Dyn), and Orx and Dyn are co-released. In the VTA, they play opposing roles in reward and motivation. To fully understand the physiological and behavioral roles of Orx transmission in the PVT, one important consideration is that Orx neurons that project to the PVT may co-release Orx with another peptide, such as Dyn. The PVT expresses both Orx receptors and κ opioid receptors, suggesting that Orx and Dyn act in tandem when released in the PVT, in addition to the VTA. The present review discusses recent findings that suggest the maladaptive recruitment of Orx/Dyn-PVT neurotransmission by drugs of abuse vs. a highly palatable food reward.

The hypocretin/orexin system as a target for excessive motivation in alcohol use disorders.

The hypocretin/orexin (ORX) system has been repeatedly demonstrated to regulate motivation for drugs of abuse, including alcohol. In particular, ORX seems to be critically involved in highly motivated behaviors, as is observed in high-seeking individuals in a population, in the seeking of highly palatable substances, and in models of dependence. It seems logical that this system could be considered as a potential target for treatment for addiction, particularly alcohol addiction, as ORX pharmacological manipulations significantly reduce drinking. However, the ORX system also plays a role in a wide range of other behaviors, emotions, and physiological functions and is disrupted in a number of non-dependence-associated disorders. It is therefore important to consider how the ORX system might be optimally targeted for potential treatment for alcohol use disorders either in combination with or separate from its role in other functions or diseases. This review will focus on the role of ORX in alcohol-associated behaviors and whether and how this system could be targeted to treat alcohol use disorders while avoiding impacts on other ORX-relevant functions. A brief overview of the ORX system will be followed by a discussion of some of the factors that makes it particularly intriguing as a target for alcohol addiction treatment, a consideration of some potential challenges associated with targeting this system and, finally, some future directions to optimize new treatments.

Role of the Orexin System on the Hypothalamus-Pituitary-Thyroid Axis.

Hypocretin/orexin (ORX) are two hypothalamic neuropeptides discovered in 1998. Since their discovery, they have been one of the most studied neuropeptide systems because of their projecting fields innervating various brain areas. The orexinergic system is tied to sleep-wakefulness cycle, and narcolepsy is a consequence of their system hypofunction. Orexinergic system is also involved in many other autonomic functions such as feeding, thermoregulation, cardiovascular and neuroendocrine regulation. The main aim of this mini review article is to investigate the relationship between ORX and thyroid system regulation. Although knowledge about the ORX system is evolving, its putative effects on hypothalamic-pituitary-thyroid (HPT) axis still appear unclear. We analyzed some studies about ORX control of HPT axis to know better the relationship between them. The studies that were analyzed suggest Hypocretin/ORX to modulate the thyroid regulation, but the nature (excitatory or inhibitory) of this possible interaction remains actually unclear and needs to be confirmed.

Attenuation of saccharin-seeking in rats by orexin/hypocretin receptor 1 antagonist

The orexin (Orx)/hypocretin system has been implicated in reward-seeking, especially for highly salient food and drug rewards. We recently demonstrated that signaling at the OxR1 receptor is involved in sucrose reinforcement and reinstatement of sucrose-seeking elicited by sucrose-paired cues in food-restricted rats. Because sucrose reinforcement has both a hedonic and caloric component, it remains unknown what aspect of this reward drives its reinforcing value. The present study examined the involvement of the Orx system in operant responding for saccharin, a noncaloric, hedonic (sweet) reward, and in cue-induced reinstatement of extinguished saccharin-seeking in ad libitum-fed vs food-restricted male subjects. Male Sprague Dawley rats were fed ad libitum or food-restricted and trained to self-administer saccharin. We determined the effects of pretreatment with the OxR1 receptor antagonist SB-334867 (SB; 10-30 mg/kg) on fixed ratio (FR) saccharin self-administration and on cue-induced reinstatement of extinguished saccharin-seeking. SB decreased responding and number of reinforcers earned during FR responding for saccharin and decreased cue-induced reinstatement of extinguished saccharin-seeking. All of these effects were obtained similarly in food-restricted and ad libitum-fed rats. These results indicate that signaling at the OxR1 receptor is involved in saccharin reinforcement and reinstatement of saccharin-seeking elicited by saccharin-paired cues regardless of food restriction. These findings lead us to conclude that the Orx system contributes to the motivational effects of hedonic food rewards, independently of caloric value and homeostatic needs.

Activation of the Orexin 1 Receptor is a Critical Component of CO2-Mediated Anxiety and Hypertension but not Bradycardia

Acute hypercapnia (elevated arterial CO(2)/H(+)) is a suffocation signal that is life threatening and rapidly mobilizes adaptive changes in breathing and behavioral arousal in order to restore acid-base homeostasis. Severe hypercapnia, seen in respiratory disorders (eg, asthma or bronchitis, chronic obstructive pulmonary disease (COPD)), also results in high anxiety and autonomic activation. Recent evidence has demonstrated that wake-promoting hypothalamic orexin (ORX: also known as hypocretin) neurons are highly sensitive to local changes in CO(2)/H(+), and mice lacking prepro-ORX have blunted respiratory responses to hypercapnia. Furthermore, in a recent clinical study, ORX-A, which crosses blood-brain barrier easily, was dramatically increased in the plasma of patients with COPD and hypercapnic respiratory failure. This is consistent with a rodent model of COPD where chronic exposure to cigarette smoke led to a threefold increase in hypothalamic ORX-A expression. In the present study, we determined the role of ORX in the anxiety-like behavior and cardiorespiratory responses to acute exposure to a threshold panic challenge (ie, 20% CO(2)/normoxic gas). Exposing conscious rats to such hypercapnic, but not atmospheric air, resulted in respiratory, pressor, and bradycardic responses, as well as anxiety-like behavior and increased cellular c-Fos responses in ORX neurons. Systemically, pre-treating rats with a centrally active ORX1 receptor antagonist (30 mg/kg SB334867) attenuated hypercapnic gas-induced pressor and anxiety responses, without altering the robust bradycardia response, and only attenuated breathing responses at offset of the CO(2) challenge. Our results show that the ORX system has an important role in anxiety and sympathetic mobilization during hypercapnia. Furthermore, ORX1 receptor antagonists may be a therapeutic option rapidly treating increased anxiety and sympathetic drive seen during panic attacks and in hypercapnic states such as COPD.

A key role for orexin in panic anxiety

Introductory paragraphPanic disorder is a severe anxiety disorder with recurrent, debilitating panic attacks. In subjects with panic disorder there is evidence of decreased central GABAergic activity as well as marked increases in autonomic and respiratory responses following intravenous infusions of 0.5M sodium lactate1–3. In an animal model of panic disorder, chronic inhibition of GABA synthesis in the dorsomedial/perifornical hypothalamus of rats produces anxiety-like states and a similar vulnerability to sodium lactate-induced cardioexcitatory responses4–9. The dorsomedial/perifornical hypothalamus is enriched in orexin (ORX, also known as hypocretin)-containing neurons10 that play a critical role in arousal10,11, vigilance10 and central autonomic mobilization12, all of which are key components of panic. Here, we demonstrate that activation of the ORX neurons is necessary for developing a panic-prone state in the animal model, and either silencing the hypothalamic ORX gene (Hcrt) product with RNA interference or systemic ORX1 antagonists blocks the panic responses. Moreover, we show that subjects with panic anxiety have elevated levels of ORX in the cerebrospinal fluid compared to subjects without panic anxiety. Taken together our results suggest that the ORX system may be involved in the pathophysiology of panic anxiety, and that ORX antagonists constitute a potential novel treatment strategy for panic disorder.

Orexin-1 receptor antagonism decreases ethanol consumption and preference selectively in high-ethanol–preferring Sprague–Dawley rats

Work from our laboratory has shown that orexin (ORX; or hypocretin) neurons in the lateral hypothalamus are involved in preference for morphine, cocaine, and food. Other groups have demonstrated a connection between the ORX system and ethanol-related behaviors. Here, we extended those results to investigate, in outbred Sprague–Dawley rats, the relationship between ethanol preference and the ORX system. In Experiment 1, rats were trained to drink 10% ethanol using the intermittent access (IA) technique. In Experiment 2, different groups of rats were trained to drink 10% ethanol using either IA or the sucrose-fade (SF) technique. Following ethanol-drinking acquisition, ethanol preference was assessed using two-bottle-choice tests. The rats were then tested for changes in preference with additional two-bottle-choice tests following administration of the orexin-1 receptor antagonist SB-334867 (SB; 30 mg/kg, intraperitoneally). Differences in ethanol preference were observed across individuals, with a significantly higher ethanol preference observed in rats trained to drink using IA compared with SF. In both Experiments 1 and 2, SB reduced ethanol preference selectively in rats with high ethanol preference. These results demonstrate a strong, causal relationship between the ORX system and ethanol preference in outbred rats. These findings provide additional evidence that the ORX system provides opportunities to develop novel treatments for alcohol abuse.