Orexin, Sleep, Sympathetic Neural Activity and Cardiovascular Function

Abstract

Inadequate sleep duration and quality are associated with reduced cardiovascular health and increased mortality. Experimental evidence points to the sympathetic nervous system as a key mediator in the observed relationship between poor sleep and cardiovascular dysfunction. However, brain mechanisms underpinning the impaired sympathetic function associated with poor sleep remain unclear. Recent evidence suggests the central orexin system, particularly orexins A and B and their receptors, have a key regulatory role for sleep in animal and human models. While orexin system activity has been observed to significantly impact sympathetic regulation in animals, the extension of these findings to humans has been difficult due to an inability to directly assess orexin system activity in humans. However, direct measures of sympathetic activity in populations with narcolepsy and chronic insomnia, two sleep disorders associated with deficient and excessive orexin neural activity, have allowed indirect assessment of the relationships between orexin, sleep, and sympathetic regulation. Further, the recent pharmaceutical development of dual orexin receptor antagonists for use in clinical insomnia populations offers an unprecedented opportunity to examine the mechanistic role of orexin in sleep and cardiovascular health in humans. The current review assesses the role of orexin in both sleep and sympathetic regulation from a translational perspective, spanning animal and human studies. The review concludes with future research directions necessary to fully elucidate the mechanistic role for orexin in sleep and sympathetic regulation in humans.

Introduction

A significant proportion of the general population suffers from poor and/or insufficient sleep.^1–3 Reduced sleep quantity and quality is associated with a number of cardiovascular and metabolic disorders, including an increased risk of hypertension as reported in cross-sectional¹ and longitudinal analyses.⁴ Sleep disorders such as insomnia are associated with cardiometabolic dysfunction.^{5, 6} While the preponderance of epidemiological studies indicate a clear role for poor sleep efficiency and quantity in the pathogenesis of cardiovascular dysfunction, the mechanism(s) underlying these associations remain elusive.

One posited mechanism by which poor sleep contributes to cardiovascular dysfunction is dysregulation of the sympathetic nervous system. Experimental sleep deprivation and sleep restriction paradigms have reported elevated levels of sympathetic activity following acute sleep impairment.^7–9 Dysfunction of the sympathetic nervous system is similarly observed within sleep disorders,^{10, 11} as well as cardiometabolic disorders such as hypertension and¹² heart failure.¹³ Despite these findings, the central regulatory mechanisms underpinning these associations has not been established in humans.

Recent attention has shifted towards the potential role for the central orexin system in sleep and cardiovascular regulation.^14–17 In an attempt to build upon prior findings, the current review assesses the crucial role that central orexinergic activity has on poor sleep and sympathetic dysregulation within animal and human models. Further, the potential for pharmaceutical interventions targeting the orexin system to more adequately assess its impact in human models, and to improve upon the translational efficacy of studies to date, will be discussed. Specifically, the recent development of dual orexin receptor antagonists offers a unique opportunity to assess the dynamic impact of orexin function on sleep and sympathetic neural control, not only in rodents and small animals, but also in humans.

Anatomical Situation of Orexin Neurons and Axonal Projections

The orexin peptides, also called hypocretins, were discovered simultaneously by two research groups just over two decades ago.^{18, 19} The orexin peptides consist of two types, orexin-A and B, which are encoded by the hypocretin neuropeptide precursor (HCRT) gene, and subsequently produced from the same precursor, prepro-orexin. Orexin producing neurons are contained within the lateral, dorsomedial, posterior and perifornical areas of the hypothalamus.^{18, 19} The isolation of orexin-producing neurons within specific hypothalamic regions suggests a limited scope of influence by the orexin system on other brain regions and physiological processes. However, orexin neurons extend a network of axonal projections which innervate numerous brain regions,^{20, 21} allowing a profound effect on complex physiological processes.

Some of the densest orexinergic axonal projections synapse in wake-promoting brain areas such as the noradrenergic locus coeruleus (LC), among others.^{20, 21} Orexin neurons receive additional input from the sleep-promoting regions,^{22, 23} which contribute to the modulatory role of orexin in sleep/wake transitions. In addition to its clear impact on sleep, orexin impacts areas involved in sympathetic autonomic control, including the nucleus of the solitary tract (NTS), rostral ventrolateral medulla (RVLM), and the parvocellular neurons of the paraventricular nucleus of the hypothalamus (PVN).^{20, 24–26} Many of these projections are reciprocated via afferent neural synapses on orexin-producing neurons from numerous autonomic and sleep/wakefulness related brain nuclei.^{22, 23} The interconnectedness of orexin producing neurons within both sleep and autonomic related brain regions indicate a crucial role of orexin as a link between insufficient sleep and impaired sympathetic control.²⁷

Activity of Orexin at Cellular Targets

Orexin neuropeptides A and B bind selectively to the orexin receptors 1 and 2 (OX1R and OX2R). OX1R and OX2R are seven-transmembrane G-protein coupled receptors with differing affinities for orexin-A and orexin-B. Specifically, orexin A has equal affinity for both receptors, while orexin B has higher affinity for OX2R.¹⁹ This is particularly important as the expression of OX1R and OX2R differs throughout brain regions. For example, OX1R is more heavily distributed within arousal promoting areas such as the LC, while OX2R is more prominent in the histaminergic tuberomammillary nucleus (TMN).²⁸ Conversely, other regions of the brain, including the serotonergic dorsal raphe (DR), the PVN, and the RVLM, express both OX1R and OX2R.^28–33

In the brain, orexin receptors are primary distributed in neurons.³² One recent study reported that PVN OX1Rs are also co-localized with astrocytes.³² In vitro studies have consistently observed a robust cyclic AMP (cAMP) production in rat cerebral cortex astrocyte cultures upon stimulation with orexin A, and this increase in cAMP is mediated by OX1R but not OX2R, suggesting that OX1R is expressed in the brain astrocyte and its activation activates G_s-adenylyl cyclase-cAMP signaling.³⁴ Whether these astrocytes expressing OX1R are involved in blood pressure and sleep regulation remains unknown.

Orexins A and B have been observed to elicit primarily excitatory influences on brain regions involved in sleep/wake^35–42 and sympathetic^43–46 regulation. However, the mechanisms by which orexins carry out this excitatory response are vast and complex (for reviews see Kukkonen et al.⁴⁷, Scammell et al.³³, and Dale et al.⁴⁸). Orexin receptors have been shown to act variably through coupling to G-protein families including G_q/11, G_i/o, and G_s,^{33, 47} whereby binding of orexins subsequently regulate phospholipases, ion channels, protein kinases, or adenylyl cyclase, ultimately triggering the activation of various downstream signaling pathways. One of the primary means of orexin-mediated neuronal excitation is through a rise in intracellular Ca²⁺ at the target cell.¹⁹ Orexin administration results in an augmented intracellular Ca²⁺ concentration that appears to be primarily mediated through extracellular Ca²⁺ influx into the cytosol.^{49, 50} A role for transient receptor potential canonical channels (TRPCs) in facilitating the entry of extracellular Ca²⁺ into the cytosol has been proposed.⁵¹ While Ca²⁺ influx from extracellular space appears to be the primary mechanism for cellular activation, intracellular release of Ca²⁺ from the endoplasmic reticulum via activation of the G_q/11 mediated phospholipase C inositol trisphosphate (IP3) pathway may serve as a secondary mediator of neuronal excitability.^{49, 50} Ca²⁺ is an important secondary messenger, which can in turn activate many Ca²⁺-sensitive enzymes including calcium/calmodulin-dependent protein kinase II (CaMKII).³² Previous studies report that orexin-mediated CaMKII activation within the rat PVN can result in elevations of sympathetic nerve activity and blood pressure.^{32, 52} Orexins may also act through activation of the sodium-calcium exchanger⁵³ or suppression of potassium efflux⁵⁴ to further post-synaptically modify neuronal excitability. Many orexin neurons are glutamatergic, and can also release glutamate to facilitate neuronal excitation.⁵⁵ Lastly, orexin neurons have been observed to modify neuronal excitability through pre-synaptic modulation of glutamate and γ-amino-butyric acid (GABA) release,^{50, 53} outlining a complex regulatory role for orexin in neuronal excitability through both pre- and post-synaptic mechanisms (Figure 1).

Figure 1:

Actions of orexins A and B at cellular targets. Orexins elicit cellular excitation at targets through numerous means. Orexin receptors have been shown to act variably through coupling to G-protein families including Gq/11, Gi/o, and Gs which ultimately lead to numerous downstream signaling cascades. Orexin binding also leads to an influx of Ca²⁺ through cation channels, an inhibition of K⁺ efflux, and pre-synaptic modulation of glutamate and GABA release. OXA, orexin A; OXB, orexin B; OX1R, orexin receptor 1; OX2R, orexin receptor 2; TRPC, transient receptor potential canonical channels; GIRK, G protein-activated inwardly rectifying K+ channels; GABA, γ-amino butyric acid; cAMP, cyclic adenosine monophosphate; ER, endoplasmic reticulum; CaMKII, calcium/calmodulin-dependent protein kinase II.

Natural Orexin Oscillations in Sleep and Wakefulness

Strong evidence of orexin’s behavior-state dependent activity come from two studies utilizing single-unit recordings of orexin neurons in head-fixed⁵⁶ and freely moving, unrestrained rats.⁵⁷ Both studies reported similar findings, namely that orexin neuronal discharge is at its peak during wakefulness in freely moving rodents, and subsequently decreases when entering resting wakefulness, slow wave sleep (SWS), and rapid eye movement (REM) sleep.^{56, 57} Orexin activities are further elevated during active exploration.⁵⁷ Similar findings have reported that orexin neuronal discharge levels are highest during goal-oriented tasks, such as foraging behavior and in response to food restriction,^{58, 59} indicating a role for orexin neuronal activation in achieving central arousal levels necessary for motivated behaviors based on external demands and energy balance. However, discharge of orexin neurons is abruptly reduced upon goal attainment. For instance, in the case of food consumption, orexin neuron activity in freely moving mice was diminished less than 1 second after contact was made with the food source.⁵⁹ As sleep ensues, orexin discharge rates are reduced with increased depth of sleep.^{56, 57} However, during REM sleep, while orexin neuronal activation is nearly absent in tonic REM, brief bursts of orexin neuronal firing occur in tandem with phasic events and muscle twitches.⁵⁷

This pattern of reduced orexin neuronal firing is similarly observed in other wake-promoting nuclei, such as the noradrenergic LC that is virtually silenced during REM sleep.⁶⁰ The activity of orexin neurons precedes muscle movement and arousals causing transitions from REM to wakefulness,⁵⁶ suggesting an important role in transitions from sleep to wakefulness. This is relevant to disorders such as insomnia, where fragmented or poor REM sleep serve as a key underlying characteristic associated with the perceived poor sleep^61–63 and subsequent emotional disturbance.^{64, 65} The findings of orexin activation preceding arousal during REM sleep, as well as the role for REM disruption in the manifestation of insomnia, suggest a link between orexin hyperactivity and the clinical traits observed within insomnia disorder.

Causal Role for Orexin in Sleep-Wake Transitions

A key role for orexin stabilization of wake and sleep states was inferred by the discovery of orexin neuron destruction in the development of narcolepsy in animals^{66, 67} and humans.^{68, 69} Experimental manipulations of orexin activity have advanced the causal role for orexin in sleep, wakefulness, and sleep state transitions. De Lecea and colleagues have shown that selective, optogenetic stimulation of orexin producing neurons evokes transitions from SWS and REM to wakefulness.^{70, 71} Activation of orexin neurons selectively reduces the latency to awakening from sleep, indicating a key role for orexin neurons in sleep-state transitons,⁷² potentially through indirect effects on other arousal promoting brain regions.^{71, 73} Due to the differential expression of OX1R and OX2R in nuclei involved in sleep/wake regulation, it is likely that OX1R and OX2R exhibit differing effects on sleep regulation. Intracerebroventricular injection of orexin-A, which binds to both receptor types, promotes wakefulness in wildtype mice, although this effects is diminished to a greater extent following selective OX2R versus OX1R depletion.³⁰ Similarly, antagonism of OX2R, but not OX1R, prior to optogenetic stimulation of orexin-producing neurons results in a significant reduction in orexin-mediated wakefulness.⁷⁴

In the case of narcolepsy, the impact of orexin on maintaining wakefulness via projections to wake-promoting brain nuclei is thought to be a primary means by which narcolepsy occurs. In the ‘flip-flop’ model of sleep-state switching proposed by Saper and colleagues,⁷² orexin neurons send excitatory input to wake-promoting neurons, which act through a negative feedback mechanism to inhibit orexin neuronal activity. In narcolepsy, due to the removal of orexin neurons from this system, wake and sleep promoting brain centers exhibit a mutual inhibitory circuit, whereby any slight increase in activity in one system exceeding the other causes abrupt changes in arousal state due to self-disinhibition, leading to transitions from wakefulness to sleep.⁷² Orexin receptor restoration in orexin-receptor deficient mice,⁷⁵ as well as intranasal orexin administration in narcoleptic humans^{76, 77} improve narcoleptic symptomology, supporting the role of proper orexin signaling in maintained sleep/wake regulation.

Conversely, mice with transgenic overexpression of prepro-orexin experience augmented levels of sleep fragmentation, particularly during REM sleep.⁷⁸ Further, abnormalities are observed in muscle tone, with chronic overexpression of orexin resulting in intrusive muscle activation during REM sleep when atonia is normally observed.⁷⁸ These findings are relevant given the associations between REM fragmentation and sleep disturbance observed in individuals with chronic insomnia.^61–63 Thus, experimental evidence supports a causal role for orexin hyperactivity in the pathogenesis of insomnia.

Recently, there has been a surge in pharmaceutical development of dual orexin receptor antagonists for the treatment of insomnia,^79–81 which have been shown to improve sleep outcomes while not drastically impacting sleep architecture. While invasive measures investigating the impact of orexin neuronal activity on sleep in humans are not feasible, the use of dual orexin receptor antagonists to improve sleep offers evidence of a key role for proper orexin signaling to maintain adequate sleep quality and offers a new research avenue to assess mechanistic roles of orexin in sleep and wakefulness in humans.

Orexin’s Role in Autonomic Nervous System Function

Orexin Regulation of Cardiac Activity

While the primary topic of this review surrounds the key role for orexin in sympathetic function and dysfunction, evidence has shown an additional role for the orexin system in parasympathetic regulation. Orexin-producing neurons synapse on brain regions involved in vagal control of the heart.^82–84 Orexin axonal projections synapse on preganglionic cardiac vagal neurons, and primarily exhibit excitatory influences through activation of glutamatergic post-synaptic currents.⁸⁴ In support of this, acute microinjection of orexin-A into regions involved in parasympathetic control of cardiac function such as the nucleus ambiguus (NAmb)⁸² and certain subnuclei of the NTS⁸³ results in a robust dose-dependent bradycardic response which is abolished following muscarinic blockade or surgical vagotomy. However, this response is dependent on the brain region effected. For instance, orexin-A administration into the commissural nucleus of the NTS elicits a paradoxical pressor and tachycardic response.^{83, 85} Similarly, orexin-A injection into the rostral ventromedial medulla elicits tachycardia, a response which is partially abolished by muscarinic blockade, and fully ameliorated following nicotinic receptor inhibition,²⁴ highlighting the complex role for orexin in the regulation of both parasympathetic and sympathetic influences at the level of the heart.

Orexin Regulation of Peripheral Sympathetic Outflow

Orexin axonal projections synapse in numerous areas involved with central regulation of sympathetic outflow to the periphery,^{20, 24–26} and receptor expression appears to be primarily found in autonomic neurons rather than glial populations.³² Early studies established a clear pressor response and associated augmented peripheral sympathetic outflow following central injection of orexin peptides in healthy animals,^{86, 87} likely through interactions at the level of pre-sympathetic neurons within the RVLM^{26, 88, 89} or indirectly through input to cardiovascular relevant regions such as the PVN.^{31, 46, 90, 91} In both brain regions, orexin-A and orexin-B are effective in eliciting membrane depolarization on target cells,^44–46 although this effect within the RVLM may be facilitated primarily through the OX2R.⁴⁵ Recent studies have additionally observed augmented baroreflex gain or sensitivity in response to orexin administration,^{29, 92, 93} indicating that orexin not only elicits a sympathoexcitatory response, but also enhances sympathetic responsiveness to blood pressure fluctuations. Finally, orexin hyperactivity has been observed in numerous models of hypertension including salt-sensitive,^{94, 95} obesity related,^{90, 91} stress-induced,⁹⁶ and others (for review see Huber et al.¹⁶).

Despite several studies examining the role of orexin in sympathetic regulation within animal models, only one study has assessed orexin’s sympathomimetic effects in healthy humans using intranasal administation.⁹⁷ Orexin administered intravenously does not appear to significantly impact cardiovascular measures, indicating a primarily central impact.⁸⁸ For this reason, a significant barrier exists in assessing the effect of orexin on sympathetic regulation in humans. However, intranasal administration of orexin-A results in elevations of cerebrospinal fluid (CSF) orexin-A concentrations in non-human primates,⁹⁸ indicating that nasal administration of orexins offers a feasible option to assess the effects of central orexin neuronal activity on peripheral sympathetic activation. In the only study of its kind, Meusel et al.⁹⁷ utilized intranasal administration of orexin-A in healthy adults to assess its effects on muscle sympathetic nerve activity (MSNA), a direct measure of post-ganglionic efferent sympathetic outflow in humans.^99–101 In response to intranasal orexin-A, the authors observed a significant, albeit modest, increase in MSNA independent of changes in other cardiovascular measures.⁹⁷ In contrast to studies in rodents,^{29, 92, 93} orexin-A administration did not impact baroreflex sensitivity,⁹⁷ although it appeared to elevate the set-point of sympathetic outflow to the periphery. In tandem with results from animal models, the findings of Meusel et al.⁹⁷ suggest a sympathomimetic effect of central orexin, supporting the concept that hyperactive orexinergic activity is a key player in the exaggerated sympathetic activation observed in human sleep disorders.¹⁰

Orexin as a Link Between Sleep Disorders and Sympathetic Dysregulation in Humans

Sleep disruption and discontinuity are associated with sympathetic dysregulation in humans.²⁷ While blood pressure is elevated in response to sleep deprivation, experimental evidence has shown that total sleep deprivation significantly increases peripheral sympathetic outflow dependent upon age and sex.^{7, 102} Similarly, levels of urinary and plasma norepinephrine have been observed to increase following semi-chronic models of sleep restriction with⁹ and without⁸ circadian misalignment in otherwise healthy adults. In an observational study assessing correlates of sleep disruption and corresponding associations with MSNA, Taylor et al.¹⁰³ observed that the frequency of nocturnal arousals, or sleep disruptions, was most highly associated with peripheral sympathetic outflow in individuals with and without obstructive sleep apnea. These findings suggest an association between inadequate sleep quality/quantity and excessive sympathetic activation in humans. However, the role that orexin plays in these associations remains difficult to disentangle in humans due to methodological limitations. While acute intranasal administration of orexin-A evokes a modest increase in sympathetic activity in healthy adults,⁹⁷ the mechanisms of its action on central regulatory centers of peripheral sympathetic outflow can only be inferred from animal models. However, human models of chronic sleep disorders known to be impacted by orexin activity (i.e., narcolepsy and insomnia) offer further insight into the role of orexin in chronic sympathetic disturbance.

Narcolepsy Types 1 and 2.

Perhaps the most compelling evidence for a role of orexin in human sympathetic function comes from a study by Donadio et al.¹¹ where MSNA was monitored in a group of patients with narcolepsy type 1 (NT1) and healthy controls. The authors reported a reduced level of peripheral sympathetic outflow in participants with NT1. These findings are consistent with early studies in orexin-knockout mice,¹⁰⁴ whereby blood pressure and cardiovascular reactivity to stress were significantly reduced compared to wild-type mice. Further, these differences were abolished following systemic application of an α-adrenergic receptor antagonist, offering evidence of reduced peripheral sympathetic outflow in orexin knockout mice.¹⁰⁴ The concentration of circulating orexin-A within the cerebrospinal fluid (CSF) of the patients with NT1 was positively associated with MSNA levels, indicating that lower levels of orexin expression was associated with blunted sympathetic activity¹¹ (Figure 2).

Figure 2:

Relationships between orexin cerebrospinal fluid concentrations and resting A) muscle sympathetic nerve activity (MSNA) and B) heart rate (HR) in individuals with narcolepsy. HCRT, hypocretin; MSNA, muscle sympathetic nerve activity (bursts/100 heart beats); HR, heart rate.Obtained with permission from Donadio et al.¹¹

While sympathetic outflow during wakefulness mimics what might be expected based on findings in animals, nocturnal sympathetic activity does not appear to be disturbed in individuals with NT1.¹⁰⁵ Rather, MSNA in NT1 patients is reduced as NREM sleep is initiated, and increased during REM sleep,¹⁰⁵ similar to healthy adults.¹⁰⁶ Despite the maintained reductions in nocturnal NREM sympathetic activity, narcolepsy is associated with elevated nocturnal blood pressure in humans^{105, 107} and animals¹⁰⁸ alike. However, the mechanisms underlying non-dipping status are inconsistent between animal and human studies.^{105, 108} Administration of prazosin, an α₁-receptor antagonist, alleviates blunted blood pressure dipping patterns in orexin-knockout mice,¹⁰⁸ hinting at a role for augmented peripheral sympathetic nerve activity and thus increased peripheral vasoconstriction as a key determinant underlying the impaired blood pressure dipping pattern. However, this hypothesized sympathetic augmentation is not observed in humans.¹⁰⁵ It is notable that Alvente et al.¹⁰⁸ also observed a normalized blood pressure dipping pattern after application of atenolol, a β₁-adrenergic antagonist in orexin knockout mice, which reduces adrenergic influence at the level of the heart. This finding suggests that while reductions in orexin levels may lead to reduced sympathetic outflow to regional arteries, including muscle sympathetic nerves, augmented regional sympathetic outflow to other organs such as the heart may be responsible for maintained elevations in nocturnal blood pressure. However, assessment of cardiac sympathetic activity in humans with narcolepsy via cardiac scintigraphy does not support this hypothesis.¹⁰⁹ These discrepant findings of nocturnal sympathetic function and elevated blood pressure in NT1 patients infer that the nocturnal cardiovascular dysfunction is secondary to disturbed sleep, not necessarily orexin deficiency. Nocturnal arousals are associated with elevations in sympathetic outflow and blood pressure.^{110, 111} Importantly, the assessment by Donadio et al.¹⁰⁵ only sampled MSNA during periods of undisturbed sleep, potentially limiting the extension of their findings to durations of sleep characterized by frequent arousals commonly associated with narcolepsy.¹¹² Grimaldi et al.¹⁰⁷ have shown that fluctuations in nocturnal blood pressure in narcolepsy are temporally related to sleep disruption caused by arousals, periodic limb movements, etc., indicating that the apparent contrary increases in nocturnal blood pressure in orexin-deficient humans may be more closely associated with sleep disruption, and not orexin-deficiency alone (Figure 3).

Figure 3:

Nocturnal blood pressure fluctuations in a representative individual with narcolepsy (A) and a healthy control (B). Nocturnal fluctuations in systolic blood pressure (SBP) in individuals with narcolepsy are temporally related to changes in wake/sleep state. NC, narcolepsy with cataplexy; SBP, systolic blood pressure; C, control subject. Obtained with permission from Grimaldi et al.¹⁰⁷

Based on these findings, it is plausible that increased sympathetic and cardiovascular reactivity to frequent arousals may lead to the augmented nocturnal blood pressure in NT1.¹⁰⁷ Conversely, during the daytime when orexin neurons should be active,^{56, 57} the absence of these neurons in narcoleptic patients results in reduced sympathetic activity.¹¹ This is further supported by a blunted morning blood pressure surge in narcoleptic patients,¹⁰⁷ given the well-characterized relationship between augmented sympathetic reactivity and elevated blood pressure surge in the morning.¹¹³

It is worth noting, however, that a recent study examining heart rate reactivity to nocturnal arousal and sleep disruption reported that NT1 patients with low levels of CSF orexin-A exhibited blunted reactivity.¹¹⁴ Given the findings that orexin neuronal activation precedes arousal,⁵⁶ and in turn facilitates shifts from sleep to wakefulness,^{70, 71} deficient orexin neural activity may blunt the cardiovascular response to sleep disruption through inadequate orexin signaling. While blunted cardiovascular reactivity to nocturnal arousal may not support the augmented nocturnal blood pressure observed in NT1, an increased sleep fragmentation may lead to frequent arousals, whereby cardiovascular parameters remain elevated throughout the night¹⁰⁷ despite acutely depressed reactivity to singular arousal events.¹¹⁴

There has been limited research assessing autonomic control in populations with narcolepsy type 2 (NT2), though this is likely due to its etiology being less understood than that of NT1, making diagnosis and further assessment challenging. NT2 is phenotypically similar to NT1, although it is not associated with additional cataplexy. While NT2 has been associated with partial loss of orexin neurons within the hypothalamus,¹¹⁵ CSF orexin levels are more often within normal ranges when compared to narcolepsy with cataplexy.^{116, 117} The similarities in symptomology, yet differing levels of orexin deficiency between NT1 and NT2, make comparative analysis of autonomic dysfunction between the two disease subtypes an applicable avenue of future research to delineate the unique roles of orexin dysfunction versus sleep impairment on sympathetic control in humans.

In summary, while research within NT1 patients supports a role for orexin in maintaining wake basal sympathetic tone, discrepant findings exist regarding its role in nocturnal autonomic control, making it difficult to definitively determine the nocturnal role of orexin on sympathetic outflow. Future work assessing the diurnal impacts of orexin-deficiency on sympathetic regulation of the vasculature are warranted to disentangle the complex mechanisms underlying cardiovascular health concerns in individuals with narcolepsy.

Insomnia

In contrast with narcolepsy, insomnia has been suggested to result from excessive orexin neural activity.⁷⁸ In a large sample of 228 humans with chronic insomnia, significant elevations in plasma orexin-A levels were reported when compared to controls.¹¹⁸ Orexin-A is highly lipophilic,¹¹⁹ allowing its access across the blood brain barrier and subsequent monitoring in the bloodstream. Further, plasma orexin-A levels were exacerbated both in the duration and self-reported severity of the disorder,¹¹⁸ outlining a key role for excessive orexin neural activity as an underlying factor of chronic insomnia.

While assessment of sympathetic activity in humans with chronic insomnia has been a variable of interest in numerous studies, the findings are often controversial and inconsistent with one another (for review¹²⁰), although this may be primarily due to differing methodologies utilized to assess and/or estimate sympathetic activity. While early assessment of sympathetic activity in chronic insomnia reported elevated plasma norepinephrine levels,¹²¹ a recent study by Grimaldi et al.¹²² reported the opposite. These findings are beneficial given existing scientific gaps, but the use of plasma catecholamine sampling is subject to numerous limitations¹²³ that limit the interpretability of the data.

To date, only one study has directly assessed sympathetic neural activity via microneurography in participants with chronic insomnia.¹⁰ In a cross-sectional analysis of 12 individuals with diagnosed chronic insomnia and 12 healthy controls, Carter et al.¹⁰ reported that baseline sympathetic outflow in insomnia participants did not differ compared to controls, although sympathetic baroreflex sensitivity was reduced. Tang et al.¹¹⁸ reported that orexin concentrations differ based upon disease severity and the course of the disorder. It is possible that lack of differences in baseline sympathetic outflow reported by Carter et al.¹⁰ were due, in part, to differing levels of insomnia symptom severity and time-course, thus differing levels of central orexin neural activation. Evidence in support of this notion can be observed in both rats and rabbits, whereby dose-dependent increases in peripheral sympathetic activity were observed following central administration of orexin peptides.^{86, 87}

Although baseline MSNA was not different between insomnia and controls, MSNA reactivity to the cold pressor test was augmented in chronic insomnia¹⁰ (Figure 4). In animal models, activity of orexin neurons can differ based upon the type of stress.^{104, 124} Kayaba et al.¹⁰⁴ reported a significantly blunted cardiovascular and locomotor response to social stress in orexin knockout mice. Conversely, the cardiovascular response to noxious stimuli was not impacted by orexin neuronal depletion, suggesting that orexin-mediated cardiovascular reactivity primarily occurs in response to social stressors that require active vigilance of the environment.¹⁰⁴ These findings were supported by subsequent research reporting that pharmaceutical dual orexin receptor antagonism blunts the cardiovascular response to fear stress (i.e., re-exposure to previous foot-shock box), but not to restraint or cold stress.¹²⁴ These findings in animal models are not concordant with those in humans with chronic insomnia,¹⁰ perhaps in part due to the differing forms of cold stress.^{10, 124} In the study by Carter et al.,¹⁰ participants were required to submerge their hand voluntarily up to the wrist in cold ice water for 2 minutes, while the animals studied by Furlong et al.¹²⁴ were placed in a 4°C refrigerator. The differences in the modality of the cold stress utilized makes translation of the findings between the two studies challenging. Further, previous work has shown that acute total sleep deprivation leads to an augmented pain perception in healthy adults exposed to cold pressor test.¹²⁵ The chronic insomnia participants in the study by Carter et al.¹⁰ received, on average, just over 6 hours of sleep per night based on 2-week actigraphy wristwatch monitoring in their home environment. As such, the insomnia participants tested likely had concurrent chronic sleep restriction that may have influenced how the cold stress and its associated perceived pain.¹²⁵ The combined difficulties in the translatability of stressor methodologies between animal^{104, 124} and human¹⁰ studies, as well as the added perceptual/psychological influence on reactivity to cold stress in humans^{10, 125} makes assessment of orexin on sympathetic regulation at rest and in response to stress difficult to interpret. Future work is needed to assess stressor-specific sympathetic reactivity in individuals with chronic insomnia, ideally in tandem with assessment of plasma or CSF orexin concentrations.

Figure 4:

Muscle sympathetic nerve (MSNA) reactivity to the cold pressor test in healthy adults and individuals with chronic insomnia. Individuals with chronic insomnia showed augmented total MSNA reactivity to cold pressor stress (A). This is similarly depicted in the representative neurogram (B) showing healthy controls (B, Top) and individuals with chronic insomnia (B, bottom) during resting baseline and a 2-minute cold pressor test. The dotted lines (B) represent amplitude normalization to the largest sympathetic burst during baseline recording. MSNA, muscle sympathetic nerve activity. Obtained with permission from Carter et al.¹⁰

Dual Orexin Receptor Antagonism and Sympathetic Cardiovascular Control

Despite the difficulties regarding direct assessment of orexin neural activity in individuals with chronic insomnia, the recent utilization of dual orexin receptor antagonists for insomnia treatment^{80, 81} offer a unique opportunity to assess the role of orexin in sympathetic regulation. In hypertensive animal models, systemic dual orexin receptor antagonism reduced blood pressure.^{126, 127} Specifically, oral administration of almorexant in spontaneously hypertensive rats reduced daytime and nocturnal arterial pressure levels, and reduced norepinephrine sampled from cerebrospinal fluid and plasma.¹²⁷ Similarly, in genetically hypertensive BPH/2J mice, Jackson et al.¹²⁶ observed reduced blood pressure following intraperitoneal almorexant administration. Following almorexant administration, sympathetic ganglionic blockade only marginally reduced blood pressure in BPH/2J mice, indicating a significant sympathoinhibitory effect of pharmaceutical orexin receptor antagonism alone.¹²⁶ To date, few studies have assessed the cardiovascular and sympathetic consequences of orexin receptor antagonism in humans. In a study of treated hypertensive subjects with insomnia symptoms that persisted for at least one month, 2 weeks of treatment with Suvorexant did not improve ambulatory blood pressure compared to the placebo condition.¹²⁸

Patel et al.¹²⁹ examined the acute impact of low-dose SB-649868, a dual orexin-receptor antagonist, on neuroendocrine and sympathetic responsiveness to hypoglycemia in healthy young men. While hypoglycemia elicited an increase in circulating epinephrine and norepinephrine, orexin antagonism did not impact these responses.¹²⁹ However, in a population of psychiatric patients treated with Suvorexant, plasma norepinephrine levels tended to decrease following 8-weeks of treatment,¹³⁰ although plasma norepinephrine sampling is once again subject to numerous limitations in interpretability, and does not directly assess regional differentiation in sympathetic outflow.¹²³

There is presently a lack of studies assessing the impact of orexin antagonism on subsequent sympathetic regulation in humans. Of the studies that have been conducted in humans,^128–130 differing concentrations and length of medication, as well as the lack of direct sympathetic recordings, make conclusions difficult. Further, differing populations and comorbidities, including hypertension¹²⁸ and psychiatric disorders,¹³⁰ add ambiguity. The presence of dual orexin receptor antagonists that have shown efficacy in clinical trials^{80, 81} provide a unique opportunity to assess direct sympathetic recordings in healthy and disordered populations to fully elucidate the impacts of orexin receptor antagonism on high fidelity markers of peripheral sympathetic activity.

Conclusions and Future Directions

While orexin producing neurons are locally produced within the lateral hypothalamic area, widespread projections to numerous brain regions allow them to have an important role in orchestrating complex physiological processes. Orexin axonal projections exhibit dense innervation in brain regions responsible for sleep/wakefulness, as well as central sympathetic regulatory brain regions, suggesting a potential mechanistic link between sleep disruption and sympathetic dysregulation. Studies performed in animals have documented a clear role for orexin in the regulation of sleep and wakefulness, as well as sympathetic outflow to the periphery in healthy and diseased models. The translation of these findings into humans has been challenging due to numerous methodological and ethical considerations. Assessment of direct sympathetic recordings in response to acute intranasal orexin administration demonstrated a moderate sympathomimetic effect of orexin, corresponding to findings observed in animals.⁹⁷ Studies assessing autonomic control in populations with sleep disorders, including narcolepsy and insomnia, have supported a role for deficient or excessive orexin neural activity as a mechanism underlying the autonomic impairments observed in these populations.^{10, 11, 107}

Despite the important conclusions taken from studies to date, additional work remains to determine how orexin mediates sleep and sympathetic control in humans. Direct measurements of sympathetic neural activity via microneurography are necessary to assess how acute and chronic treatment with oral dual orexin receptor antagonists impact sympathetic regulation and sleep parameters in healthy and disordered populations. How these treatments impact daytime versus nocturnal cardiovascular control will help to elucidate the impact of orexin versus secondary mechanisms, such as sleep fragmentation, on sympathetic regulation in humans. This is particularly important because sleep fragmentation alone has been associated with sympathoexcitation.¹⁰³ Second, attentiveness to stressors employed in laboratory studies appears warranted. In animal models, cardiovascular responses differ in response to social versus physical stress following orexin knockdown,^{104, 124} whereas in humans, individuals with chronic insomnia and presumed augmented orexin activity¹¹⁸ exhibit hyperreactivity to cold stress.¹⁰ Additional studies assessing different stressor types in humans is necessary. Third, attention to symptom severity in human sleep-disordered populations is warranted. Sympathetic activity is related to CSF orexin levels in narcolepsy,¹¹ and plasma orexin levels correspond to the severity of insomnia symptomology,¹⁰ indicating an association between present orexin levels and physiological dysfunction. Finally, age and sex need to be accounted for in research moving forward. Previous research from has shown that older, post-menopausal women are more susceptible to sympathoexcitation following sleep deprivation,⁷ supporting a greater association between poor sleep and hypertension in women compared to men.¹ Orexin antagonists were recently shown to reduce subjective vasomotor symptoms in older women,¹³¹ suggesting that treatment with orexin receptor antagonists may improve sleep quality surrounding the menopausal transition and into menopause. How these treatments impact subsequent cardiovascular and sympathetic function have yet to be determined. Further studies assessing the impact of orexin on sleep and sympathetic function may lead to a more robust understanding of mechanisms underlying the relationship between poor sleep and sympathetic dysfunction, and may lead to increased preventative care measures and improved the cardiovascular outlook for individuals impacted by chronic sleep disturbance.