Non-rapid Eye Movement Sleep Regulation Research Articles

Efferent pathways from the suprachiasmatic nucleus to the horizontal limbs of diagonal band promote NREM sleep during the dark phase in mice

The regulation of circadian rhythms and the sleep–wake states involves in multiple neural circuits. The suprachiasmatic nucleus (SCN) is a circadian pacemaker that controls the rhythmic oscillation of mammalian behaviors. The basal forebrain (BF) is a critical brain region of sleep–wake regulation, which is the downstream of the SCN. Retrograde tracing of cholera toxin subunit B showed a direct projection from the SCN to the horizontal limbs of diagonal band (HDB), a subregion of the BF. However, the underlying function of the SCN–HDB pathway remains poorly understood. Herein, activation of this pathway significantly increased non–rapid eye movement (NREM) sleep during the dark phase by using optogenetic recordings. Moreover, activation of this pathway significantly induced NREM sleep during the dark phase for first 4 h by using chemogenetic methods. Taken together, these findings reveal that the SCN–HDB pathway participates in NREM sleep regulation and provides direct evidence of a novel SCN-related pathway involved in sleep–wake states regulation.

Dihydropyridine calcium blockers do not interfere with non-rapid eye movement sleep

Non-rapid eye movement (NREM) sleep is tightly homeostatically regulated and essential for survival. In the electroencephalogram (EEG), oscillations in the delta (0.5-4 Hz) range are prominent during NREM sleep. These delta oscillations are, to date, the best indicator for homeostatic sleep regulation; they are increased after prolonged waking and fade during NREM sleep. The precise mechanisms underlying sleep homeostasis and the generation of EEG delta oscillations are still being investigated. Activity-dependent neuronal calcium influx has been hypothesized to play an important role in generating delta oscillations and might be involved in downstream signaling that mediates sleep function. Dihydropyridine blockers of L-type voltage-gated calcium channels (VGCCs) are in wide clinical use to treat hypertension and other cardiovascular disorders and are readily blood-brain-barrier penetrant. We therefore, wanted to investigate their potential effects on EEG delta oscillation and homeostatic NREM sleep regulation in freely behaving mice. In vivo two-photon imaging of cortical neurons showed larger spontaneous calcium transients in NREM sleep compared to waking. Application of the dihydropyridine calcium blocker nicardipine significantly reduced cortical calcium transients without affecting the generation of delta oscillations. Nicardipine also did not affect EEG delta oscillations over 24 h following application. The time spent in NREM sleep and NREM episode duration was also not affected. Thus, acute block of calcium entry through L-type VGCCs does not interfere with EEG delta oscillations or their homeostatic regulation, despite prior evidence from calcium channel knockout mice.

Does the adenosine deaminase (ADA) gene confer risk of sleepwalking?

Sleepwalking is a common non-rapid eye movement (NREM) parasomnia and a significant cause of sleep-related injuries. While evidence suggest that the occurrence of this condition is partly determined by genetic factors, its pattern of inheritance remains unclear, and few molecular studies have been conducted. One promising candidate is the adenosine deaminase (ADA) gene. Adenosine and the ADA enzyme play an important role in the homeostatic regulation of NREM sleep. In a single sleepwalking family, genome-wide analysis identified a locus on chromosome 20, where ADA lies. In this study, we examined if variants in the ADA gene were associated with sleepwalking. In total, 251 sleepwalking patients were clinically assessed, and DNA samples were compared to those from 94 unaffected controls. Next-generation sequencing of the whole ADA gene was performed. Bio-informatic analysis enabled the identification of variants and assessed variants enrichment in our cohort compared to controls. We detected 25 different coding and non-coding variants, of which 22 were found among sleepwalkers. None were enriched in the sleepwalking population. However, many missense variants were predicted as likely pathogenic by at least two in silico prediction algorithms. This study involves the largest sleepwalking cohort in which the role of a susceptibility gene was investigated. Our results did not reveal an association between ADA gene and sleepwalking, thus ruling out the possibility of ADA as a major genetic factor for this condition. Future work is needed to identify susceptibility genes.

Homeostatic regulation of NREM sleep, but not REM sleep, in Australian magpies.

We explore non-rapid eye movement (NREM) and rapid eye movement (REM) sleep homeostasis in Australian magpies (Cracticus tibicen tyrannica). We predicted that magpies would recover lost sleep by spending more time in NREM and REM sleep, and by engaging in more intense NREM sleep as indicated by increased slow-wave activity (SWA). Continuous 72-h recordings of EEG, EMG, and tri-axial accelerometry, along with EEG spectral analyses, were performed on wild-caught Australian magpies housed in indoor aviaries. Australian magpies were subjected to two protocols of night-time sleep deprivation: full 12-h night (n = 8) and first 6-h half of the night (n = 5), which were preceded by a 36-h baseline recording and followed by a 24-h recovery period. Australian magpies recovered from lost NREM sleep by sleeping more, with increased NREM sleep consolidation, and increased SWA during recovery sleep. Following 12-h of night-time sleep loss, magpies also showed reduced SWA the following night after napping more during the recovery day. Surprisingly, the magpies did not recover any lost REM sleep. Only NREM sleep is homeostatically regulated in Australian magpies with the level of SWA reflecting prior sleep/wake history. The significance of emerging patterns on the apparent absence of REM sleep homeostasis, now observed in multiple species, remains unclear.

GABA neurons in the ventral tegmental area regulate non-rapid eye movement sleep in mice

Sleep/wakefulness cycle is regulated by coordinated interactions between sleep- and wakefulness-regulating neural circuitry. However, the detailed mechanism is far from understood. Here, we found that glutamic acid decarboxylase 67-positive GABAergic neurons in the ventral tegmental area (VTAGad67+) are a key regulator of non-rapid eye movement (NREM) sleep in mice. VTAGad67+ project to multiple brain areas implicated in sleep/wakefulness regulation such as the lateral hypothalamus (LH). Chemogenetic activation of VTAGad67+ promoted NREM sleep with higher delta power whereas optogenetic inhibition of these induced prompt arousal from NREM sleep, even under highly somnolescent conditions, but not from REM sleep. VTAGad67+ showed the highest activity in NREM sleep and the lowest activity in REM sleep. Moreover, VTAGad67+ directly innervated and inhibited wake-promoting orexin/hypocretin neurons by releasing GABA. As such, optogenetic activation of VTAGad67+ terminals in the LH promoted NREM sleep. Taken together, we revealed that VTAGad67+ play an important role in the regulation of NREM sleep.

Increased interictal spike activity associated with transient slow wave trains during non-rapid eye movement sleep

Non-rapid eye movement (NREM) sleep is characterized by recurring transient events (Cyclic Alternating Patterns, CAP), some of which consist of increased slow wave activity (A1 subtype). Such transient slow-wave events may play an important role in NREM sleep regulation and are known to facilitate epileptiform activity. In our study we investigated the relationship between interictal spike activity and Cyclic Alternating Patterns in three epileptic patients, using simultaneous scalp electroencephalogram (EEG) recording and intracranial electrocorticography. A significant increase of interictal activity was found during CAP A1 subtypes. A positive correlation between scalp EEG delta power and spike activity was found only in CAP A1 subtypes, but not during other events. These results show that transient (but not subcontinuous) delta activity has a facilitating effect on epileptoform activity, also suggesting a functional dissociation between morphologically similar delta activities.

Effects of i.c.v. administration of interleukin-1 on sleep and body temperature of interleukin-6-deficient mice

Cytokines in brain contribute to the regulation of physiological processes and complex behavior, including sleep. The cytokines that have been most extensively studied with respect to sleep are interleukin (IL)-1β, tumor necrosis factor (TNF)-α, and IL-6. Administration of these cytokines into laboratory animals, or in some cases into healthy human volunteers, increases the amount of time spent in non-rapid eye movement (NREM) sleep. Although antagonizing the IL-1 or TNF systems reduces the amount of time laboratory animals spend in NREM sleep, interactions among these three cytokine systems as they pertain to the regulation of physiological NREM sleep are not well understood. To further elucidate mechanisms in brain by which IL-1β, TNFα, and/or IL-6 contribute to NREM sleep regulation, we injected recombinant murine interleukin-1β (muIL-1β) into C57BL/6J mice and into IL-6-deficient mice (IL-6 knockout, KO). IL-6 KO (B6.129S6- Il6 tm1Kopf ; n=13) and C57BL/6J mice ( n=14) were implanted with telemeters to record the electroencephalogram (EEG) and core body temperature, as well as with indwelling guide cannulae targeted to one of the lateral ventricles. After recovery and habituation, mice were injected intracerebroventricularly just prior to dark onset on different days with either 0.5 μl vehicle (pyrogen-free saline; PFS) or with 0.5 μl PFS containing one of four doses of muIL-1β (2.5 ng, 5 ng, 10 ng, 50 ng). No mouse received more than two doses of muIL-1β, and administration of muIL-1β doses was counter-balanced to eliminate potential order effects. Sleep–wake behavior was determined for 24 h after injections. i.c.v. administration of muIL-1β increased in NREM sleep of both mouse strains in a dose-related fashion, but the maximal increase was of greater magnitude in C57Bl/6J mice. muIL-1β induced fever in C57Bl/6J mice but not in IL-6 KO mice. Collectively, these data demonstrate IL-6 is necessary for IL-1 to induce fever, but IL-6 is not necessary for IL-1 to alter NREM sleep.

Growth hormone-releasing hormone: cerebral cortical sleep-related EEG actions and expression

Growth hormone-releasing hormone (GHRH), its receptor (GHRHR), and other members of the somatotropic axis are involved in non-rapid eye movement sleep (NREMS) regulation. Previously, studies established the involvement of hypothalamic GHRHergic mechanisms in NREMS regulation, but cerebral cortical GHRH mechanisms in sleep regulation remained uninvestigated. Here, we show that unilateral application of low doses of GHRH to the surface of the rat somatosensory cortex ipsilaterally decreased EEG delta wave power, while higher doses enhanced delta power. These actions of GHRH on EEG delta wave power occurred during NREMS but not during rapid eye movement sleep. Further, the cortical forms of GHRH and GHRHR were identical to those found in the hypothalamus and pituitary, respectively. Cortical GHRHR mRNA and protein levels did not vary across the day-night cycle, whereas cortical GHRH mRNA increased with sleep deprivation. These results suggest that cortical GHRH and GHRHR have a role in the regulation of localized EEG delta power that is state dependent, as well as in their more classic hypothalamic role in NREMS regulation.

Prostaglandin D2 in Sleep-Wake Regulation: Recent Progress and Perspectives

Prostaglandin (PG) D2 is one of the most active endogenous sleep-promoting substances, which induces physiological sleep in rodents, primates, and most probably in humans as well. In this update article, we review recent experimental results concerning the molecular mechanisms underlying sleep-wake regulation by PGD2, the link between the humoral regulation by the PGD2 system, and the neural network involved in the promotion of non-rapid eye movement (NREM) sleep and the abnormality of NREM sleep regulation found in gene-manipulated mice for PGD synthase.

Effect of Lesions of the Ventrolateral Preoptic Nucleus on NREM and REM Sleep

Neurons in the ventrolateral preoptic nucleus (VLPO) in rats show c-fos activation after sleep and provide GABAergic innervation of the major monoamine arousal systems, suggesting that they may be a necessary part of the brain circuitry that produces sleep. We examined the effects on sleep behavior in rats of cell-specific damage to the VLPO by microinjection of ibotenic acid. Severe lesions of the central cell cluster of the VLPO ( approximately 80-90% cell loss bilaterally) caused a 60-70% decrease in delta power and a 50-60% decrease in nonrapid-eye-movement (NREM) sleep time (p < 0.001). The number of remaining Fos-immunoreactive neurons in the VLPO cell cluster was linearly related to NREM sleep time (r = 0.77; p < 0.001) and total electroencephalogram delta power (r = 0. 79; p < 0.001) but not to rapid-eye-movement (REM) sleep (r = 0.35; p > 0.10). Lesions in the region containing scattered VLPO neurons medial or dorsal to the cell cluster caused smaller changes in NREM sleep time (24.5 or 15%, respectively) but were more closely associated with loss of REM sleep (r = 0.74; p < 0.01). The insomnia caused by bilateral VLPO lesions persisted for at least 3 weeks. Lesions of the VLPO caused no change in mean body temperature or its circadian variation; after small lesions of the ventromedial preoptic nucleus, body temperature showed normal circadian variation but a wider temperature range, and sleep behavior was not affected. These experiments delineate distinct preoptic sites with primary effects on the regulation of NREM sleep, REM sleep, and body temperature.

Prostaglandin D synthase gene is involved in the regulation of non-rapid eye movement sleep.

To examine the function of prostaglandin (PG) D synthase (PGDS) gene, as well as endogenously produced PGD(2) in sleep regulation in vivo, we generated transgenic (TG) mice that overexpress human PGDS gene to study their sleep behavior. Although no difference was observed in the sleep/wake patterns between wild-type and TG mice, a striking time-dependent increase in non-rapid eye movement (NREM), but not in rapid eye movement (REM), sleep was observed in two independent lines of TG mice after stimulation by tail clipping. Concomitantly, the spontaneous locomotor activity of TG animals was drastically decreased in response to the tail clip. Induction of NREM sleep in TG mice was positively correlated with the PGD(2) production in the brain. Sleep, locomotion, and PGD(2) content were essentially unchanged in wild-type mice after tail clipping. The results with TG mice demonstrate the involvement of the PGDS gene in the regulation of NREM sleep.

Sleep after arousal from hibernation is not homeostatically regulated.

Electroencephalographic slow-wave activity (SWA) in non-rapid eye movement (NREM) sleep is directly related to prior sleep/wake history, with high levels of SWA following extended periods of wake. Therefore, SWA has been thought to reflect the level of accumulated sleep need. The discovery that euthermic intervals between hibernation bouts are spent primarily in sleep and that this sleep is characterized by high and monotonically declining SWA has led to speculation that sleep homeostasis may play a fundamental role in the regulation of the timing of bouts of hibernation and periodic arousals to euthermia. It was proposed that because the SWA profile seen after arousal from hibernation is strikingly similar to what is seen in nonhibernating mammals after extended periods of wakefulness, that hibernating mammals may arouse from hibernation with significant accumulated sleep need. This sleep need may accumulate during hibernation because the low brain temperatures during hibernation may not be compatible with sleep restorative processes. In the present study, golden-mantled ground squirrels were sleep deprived during the first 4 h of interbout euthermia by injection of caffeine (20 mg/kg ip). We predicted that if the SWA peaks after bouts of hibernation reflected a homeostatic response to an accumulated sleep need, sleep deprivation should simply have displaced and possibly augmented the SWA to subsequent recovery sleep. Instead we found that after caffeine-induced sleep deprivation of animals just aroused from hibernation, the anticipated high SWA typical of recovery sleep did not occur. Similar results were found in a study that induced sleep deprivation by gentle handling (19). These findings indicate that the SWA peak immediately after hibernation does not represent homeostatic regulation of NREM sleep, as it normally does after prolonged wakefulness during euthermia, but instead may reflect some other neurological process in the recovery of brain function from an extended period at low temperature.

Thermosensitive neurons of the diagonal band in rats: relation to wakefulness and non-rapid eye movement sleep

The thermosensitivity of the neurons in the diagonal band of Broca (DBB) was studied in 12 freely moving rats by determining responses to local cooling or warming with a water perfused thermode. Of 151 neurons studied, 37 (25%) neurons met the criterion for thermosensitivity including 17 warm-sensitive (WSNs) and 20 cold-sensitive neurons (CSNs). The spontaneous discharge rates of WSNs and CSNs were recorded through 1–3 sleep-waking cycles. The discharge of WSNs and CSNs during waking and non-rapid eye movement (NREM) sleep were different. Of 17 WSNs, 10 exhibited increased discharge rates during NREM sleep as compared with waking (NREM/Wake discharge ratio, ≥ 1.2). Of 20 CSNs, 14 discharged more slowly during NREM sleep as compared with waking (NREM/Wake discharge ratio, ≤ 0.8). In both WSNs and CSNs, changes in discharge rate preceded EEG changes at the waking-NREM transition. These results support a hypothesis that the activation of sleep-related WSNs and the deactivation of wake-related CSNs play a role in the onset and regulation of NREM sleep.

Chapter 22 Sleep regulation: relation to photoperiod, sleep duration, waking activity, and torpor

The homeostatic regulation of non-rapid eye-movement sleep (NREMS) consists of the changes in both duration and intensity, the latter being reflected by EEG slow-wave activity (SWA). In some species, the seasonal changes have a prominent effect on behavior and on physiological variables, and specific, seasonally recurring behaviors, such as breeding and torpor, are triggered by the photoperiod. During emergence from torpor, SWA rapidly increased to a level that significantly exceeded the baseline value. The increase of SWA and the spectral changes of the sleep EEG after emergence from torpor closely resembled the increase of SWA after 4-hour sleep deprivation. However, the effect of torpor was shorter lasting. To control for the metabolic exertion during the rise of body temperature, the emergence from torpor was promoted by heating the animal. The increase of SWA after this manipulation did not differ from the rise following the unaided termination of torpor. Taken together, these studies, as well as the hibernation experiments, strongly indicate that Process S builds up not only during waking but also in the course of other non-sleep states. They point to the exquisite role of sleep that alone can reduce the level of S.

Neuronal discharge of preoptic/anterior hypothalamic thermosensitive neurons: relation to NREM sleep.

Thermosensitive neurons of the preoptic/anterior hypothalamic area (POAH) have been implicated in the regulation of non-rapid eye movement (NREM) sleep. We attempted to identify those medial POAH thermosensitive neurons that may be involved in NREM sleep regulation. The thermosensitivity of medial POAH neurons was studied in five freely moving adult cats by local cooling or warming of the medial POAH with a water-perfused thermode. Of 308 neurons, 65 (21%) were classified as thermosensitive, including 31 (10%) warm-sensitive and 34 (11%) cold-sensitive neurons. The spontaneous discharge rates of 28 warm-sensitive, 34 cold-sensitive, and 115 randomly selected thermoinsensitive neurons were recorded through one to three sleep-waking cycles. Patterns of spontaneous activity for warm- and cold-sensitive neurons were different. Of 28 warm-sensitive neurons, 18 (64%) exhibited increased discharge rate during NREM sleep compared with waking (NREM/wake, > or = 1.2). This subpopulation of warm-sensitive neurons also exhibited significantly increased thermosensitivity when tested during NREM sleep. Of 34 cold-sensitive neurons, 25 (74%) discharged more slowly during NREM sleep compared with waking (NREM/wake, < or = 0.8). This subpopulation of cold-sensitive neurons exhibited decreased thermosensitivity during NREM sleep. These results are consistent with a hypothesis that the activation of sleep-related warm-sensitive neurons and the deactivation of wake-related cold-sensitive neurons may play a key role in the onset and regulation of NREM sleep.

Epilepsy and sleep

This review discusses several aspects of epilepsy and sleep. The level of wakefulness is controlled by transmitters such as acetylcholine, norepinephrine, serotonin, and histamine. These neurotransmitters are involved in modulatory neurotransmission of the ascending brain stem systems, which play an important role in controlling the sleep-wake cycle. Experimental evidence suggests that rapid eye movement sleep atonia is induced by increased endogenous acetylcholine release. Regarding sleep factors, recent data suggest that prolactin may stimulate rapid eye movement sleep and that growth hormone-releasing factor is involved in non-rapid eye movement sleep regulation. The neuropharmacologic features of several animal models of epilepsy have been investigated. Epileptiform discharges of genetic absence seizures in rats have been found to be suppressed by cholinergic drugs. The beta-carboline abecarnil has a strong antiepileptic effect on spike-wave discharges in rats that generate spontaneous spike-wave discharges, and it may be useful as an antiepileptic drug. Sleep epilepsy has been studied in a model of amygdala-kindled kittens. During the postkindling development, multifocal epilepsy with convulsions occurred that were distributed throughout the sleep-wake cycle; this finding agrees with the clinical literature. A typical feature of juvenile myoclonic epilepsy is the occurrence of seizures in a strict relationship to the sleep-wake cycle. There is now some neurophysiologic evidence that this syndrome also causes disturbance of sleep stability with increased arousal reactions. Nocturnal paroxysmal dystonia is not a homogeneous entity. Several clinical reports indicate that the short-lasting variant is most likely a form of frontal lobe epilepsy, but the nature of the longer-lasting variants is still obscure.

RP 62203, a 5-hydroxytryptamine2 antagonist, enhances deep NREM sleep in rats.

RP 62203, a naphtosultam derivative, is an antagonist at the 5-hydroxytryptamine2 (HT2) receptor. The sleep pattern of rats treated orally with RP 62203 was studied at doses ranging from 0.5 to 4 mg/kg. Following RP 62203 administration, the duration of deep nonrapid eye movement (NREM) sleep was found to increase at the expense of wakefulness in a dose-dependent manner from 0.5 mg/kg. The 5-HT2 receptor agonist DOI and the 5-HT1a receptor agonist 8 OH-DPAT induced a dose-related increase in wakefulness; treatment with RP 62203 reversed the enhancement of wakefulness produced by DOI but not that produced by 8 OH-DPAT. These data provide further evidence for the involvement of 5-HT2 receptors in the regulation of NREM sleep in rats. RP 62203 could therefore be of clinical interest in the management of sleep disorders, particularly those developing within a psychiatric context.

Effects of seganserin, a 5-HT 2 antagonist, and temazepam on human sleepsstages and EEG power spectra

The effects of seganserin, a specific 5HT 2 antagonist, on human sleep were assessed in two experiments and compared to the effects of temazepam and sleep deprivation. During daytime recovery sleep after sleep derivation, seganserin did not significantly enhance visually scored slow wave sleep (SWS, stages 3+4) or the EEG power density in the delta frequencies. Under these conditions temazepam reduced the power density in the delta and theta frequencies. During nightime sleep after a nap in the evening, seganserin caused an increase in SWS, a reduction in intermittent wakefulness, and an enhancement of the power density in the delta and theta frequencies during non-rapid eye movement (NREM) sleep. Temazepam induced a reduction in the power density in the delta and theta frequencies. It is concluded that the 5HT 2 antagonist, seganserin, can induce SWS. However, since the spectral results showed that the changes in the sleep EEG were not identical to those induced by sleep deprivation it seems premature to conclude that 5HT 2 receptors are primarily involved in NREM sleep regulation.

Ulcerative Colitis

<h3>Abstract</h3> The daily sleep/wakefulness cycle is regulated by coordinated interactions between sleep- and wakefulness-regulating neural circuitry. However, the detailed neural circuitry mediating sleep is far from understood. Here, we found that glutamic acid decarboxylase 67 (Gad67)-positive GABAergic neurons in the ventral tegmental area (VTA_Gad67+) are a key regulator of non-rapid eye movement (NREM) sleep in mice. VTA_Gad67+ neurons project to multiple brain areas implicated in sleep/wakefulness regulation such as the lateral hypothalamus (LH) and dorsal raphe nucleus. Chemogenetic activation of VTA_Gad67+ neurons promoted NREM sleep with higher delta power whereas optogenetic inhibition of these neurons induced prompt arousal from NREM sleep under highly somnolescent conditions, but not during REM sleep. <i>In vivo</i> fiber photometry recordings revealed that VTA_Gad67+ neurons showed the highest population activity in NREM sleep and the lowest activity in REM sleep. Acute brain slice electrophysiology combined with optogenetics revealed that VTA_Gad67+ neurons directly innervate and inhibit wake-promoting orexin/hypocretin neurons in the LH by releasing GABA. Taken together, we reveal that VTA_Gad67+ neurons play a crucial role in the regulation of NREM sleep.