CD40 activation induces NREM sleep and modulates genes associated with sleep homeostasis.

The T-cell derived cytokine CD40 ligand is overexpressed in patients with autoimmune diseases. Through activation of its receptor, CD40 ligand leads to a tumor necrosis factor (TNF) receptor 1 (TNFR1) dependent impairment of locomotor activity in mice. Here we report that this effect is explained through a promotion of sleep, which was specific to non-rapid eye movement (NREM) sleep while REM sleep was suppressed. The increase in NREM sleep was accompanied by a decrease in EEG delta power during NREM sleep and by a decrease in the expression of transcripts in the cerebral cortex known to be associated with homeostatic sleep drive, such as Homer1a, Early growth response 2, Neuronal pentraxin 2, and Fos-like antigen 2. The effect of CD40 activation was mimicked by peripheral TNF injection and prevented by the TNF blocker etanercept. Our study indicates that sleep-wake dysregulation in autoimmune diseases may result from CD40 induced TNF:TNFR1 mediated alterations of molecular pathways, which regulate sleep-wake behavior.

CD40 activation induces NREM sleep and modulates genes associated with sleep homeostasis

The T-cell derived cytokine CD40 ligand is overexpressed in patients with autoimmune diseases. Through activation of its receptor, CD40 ligand leads to a tumor necrosis factor (TNF) receptor 1 (TNFR1) dependent impairment of locomotor activity in mice. Here we report that this effect is explained through a promotion of sleep, which was specific to non-rapid eye movement (NREM) sleep while REM sleep was suppressed. The increase in NREM sleep was accompanied by a decrease in EEG delta power during NREM sleep and by a decrease in the expression of transcripts in the cerebral cortex known to be associated with homeostatic sleep drive, such as Homer1a, Early growth response 2, Neuronal pentraxin 2, and Fos-like antigen 2. The effect of CD40 activation was mimicked by peripheral TNF injection and prevented by the TNF blocker etanercept. Our study indicates that sleep-wake dysregulation in autoimmune diseases may result from CD40 induced TNF:TNFR1 mediated alterations of molecular pathways, which regulate sleep-wake behavior.

Lucid dreaming during NREM sleep: Two case reports

Lucid dreams – dreams in which the dreamer is aware that is dreaming – most frequently occur during REM sleep, yet there is some evidence suggesting that lucid dreaming can occur during NREM sleep as well. By conducting a sleep laboratory study on lucid dreams, we found two possible instances of lucidity during NREM sleep which are reported here. While lucid dreaming during NREM sleep seems to be much rarer and more difficult to achieve, it appears to be possible and is most likely to occur during N1 sleep, somewhat less likely during N2 sleep and yet to be observed during N3 sleep. Future studies should explore induction methods, underlying neural mechanisms and perceptual/dream content differences between REM and NREM lucid dreams. Furthermore, a consensus agreement is needed to define what is meant by lucid dreaming and create a vocabulary that is helpful in clarifying variable psychophysiological states that can support self-reflective awareness.

Effects of sleep fragmentation on the arousability to resistive loading in NREM and REM sleep in normal men

STUDY OBJECTIVE: In healthy subjects, arousability to inspiratory resistive loading is greater during rapid eye movement (REM) sleep compared with non-REM (NREM) sleep but is poorest in REM sleep in patients with sleep apnea. We therefore examined the hypothesis that sleep fragmentation impairs arousability, especially from REM sleep. DESIGN: Two blocks of 3 polysomnographies (separated by at least 1 week) were performed randomly. An inspiratory-loaded night followed either 2 undisturbed control nights (LN(C)) or 2 acoustically fragmented nights (LN(F)) SETTING: Sleep laboratory. PARTICIPANTS: Sixteen healthy men aged 20 to 29 years. INTERVENTIONS: In both loaded nights, an inspiratory resistive load was added via a valved facemask every 2 minutes during sleep and turned off either when arousal occurred or after 2 minutes. MEASUREMENTS AND RESULTS: During LN(F), arousability remained significantly greater in REM sleep (71% aroused within 2 minutes) compared with stage 2 (29%) or stage 3/4 (16%) sleep. After sleep fragmentation, arousability was decreased in stage 2 sleep (LN(F): 29%; LN(C): 38%; p < .05) and low in early REM sleep, increasing across the night (p < .01). In stage 3/4 sleep, neither an attenuation nor a change across the night was seen after sleep fragmentation. CONCLUSIONS: Mild sleep fragmentation is already sufficient to attenuate arousability in stage 2 sleep and to decrease arousability in early, compared with late, REM sleep. This means that sleep fragmentation affects the arousal response to increasing resistance and that the effects are different in stage 2 and REM sleep. The biologic reason for this increase in the arousal response in REM sleep across the night is not clear.

Autonomic activity during human sleep as a function of time and sleep stage

While there is a developing understanding of the influence of sleep on cardiovascular autonomic activity in humans, there remain unresolved issues. In particular, the effect of time within the sleep period, independent of sleep stage, has not been investigated. Further, the influence of sleep on central sympathetic nervous system (SNS) activity is uncertain because results using the major method applicable to humans, the low frequency (LF) component of heart rate Variability (HRV), have been contradictory, and because the method itself is open to criticism. Sleep and cardiac activity were measured in 14 young healthy subjects on three nights. Data was analysed in 2-min epochs. All epochs meeting specified criteria were identified, beginning 2 h before, until 7 h after, sleep onset. Epoch values were allocated to 30-min bins and during sleep were also classified into stage 2, slow wave sleep (SWS) and rapid eye movement (REM) sleep. The measures of cardiac activity were heart irate (HR), blood pressure (BP), high frequency (HF) and LF components of HRV and pre-ejection period (PEP). During non-rapid eye movement (NREM) sleep autonomic balance shifted from sympathetic to parasympathetic dominance, although this appeared to be more because of a shift in parasympathetic nervous system (PNS) activity. Autonomic balance during REM was in general similar to wakefulness. For BP and the HF and LF components the change occurred abruptly at sleep onset and was then constant over time within each stage of sleep, indicating that any change in autonomic balance over the sleep period is a consequence of the changing distribution of sleep stages. Two variables, HR and PEP, did show time effects reflecting a circadian influence over HR and perhaps time asleep affecting PEP. While both the LF component and PEP showed changes consistent with reduced sympathetic tone during sleep, their pattern of change over time differed.

Ecology and neurophysiology of sleep in two wild sloth species

Study Objectives: Interspecific variation in sleep measured in captivity correlates with various physiological and environmental factors, including estimates of predation risk in the wild. However, it remains unclear whether prior comparative studies have been confounded by the captive recording environment. Herein we examine the impact of predation pressure on sleep in sloths living in the wild. Design: Comparison of two closely related sloth species, one exposed to predation and one free from predation. Setting: Panamanian mainland rainforest (predators present) and island mangrove (predators absent). Participants: Mainland (Bradypus variegatus, 5 males and 4 females) and island (Bradypus pygmaeus, 6 males) sloths. Interventions: None. Measurements and Results: EEG and EMG activity were recorded using a miniature data logger. Although both species spent between 9 and 10 hours per day sleeping, the mainland sloths showed a preference for sleeping at night, whereas island sloths showed no preference for sleeping during the day or night. EEG activity during NREM sleep showed lower low-frequency power, and increased spindle and higher frequency power in island sloths when compared to mainland sloths. Conclusions: In sloths sleeping in the wild, predation pressure influenced the timing of sleep, but not the amount of time spent asleep. The preference for sleeping at night in mainland sloths may be a strategy to avoid detection by nocturnal cats. The pronounced differences in the NREM sleep EEG spectrum remain unexplained, but might be related to genetic or environmental factors.

Sleep EEG changes after middle cerebral artery infarcts in mice: different effects of striatal and cortical lesions.

STUDY OBJECTIVES: Hemispheric stroke in humans is associated with sleep-wake disturbances and sleep electroencephalogram (EEG) changes. The correlation between these changes and stroke extent remains unclear. In the absence of experimental data, we assessed sleep EEG changes after focal cerebral ischemia of different extensions in mice. DESIGN: Following electrode implantation and baseline sleep-wake EEG recordings, mice were submitted to sham surgery (control group), 30 minutes of intraluminal middle cerebral artery (MCA) occlusion (striatal stroke), or distal MCA electrocoagulation (cortical stroke). One and 12 days after stroke, sleep-wake EEG recordings were repeated. The EEG recorded from the healthy hemisphere was analyzed visually and automatically (fast Fourier analysis) according to established criteria. MEASUREMENTS AND RESULTS: Striatal stroke induced an increase in non-rapid eye movement (NREM) sleep and a reduction of rapid eye movement sleep. These changes were detectable both during the light and the dark phase at day 1 and persisted until day 12 after stroke. Cortical stroke induced a less-marked increase in NREM sleep, which was present only at day 1 and during the dark phase. In cortical stroke, the increase in NREM sleep was associated in the wake EEG power spectra, with an increase in the theta and a reduction in the beta activity. CONCLUSION: Cortical and striatal stroke lead to different sleep-wake EEG changes in mice, which probably reflect variable effects on sleep-promoting and wakefulness-maintaining neuronal networks.

Effect of expiratory positive airway pressure on sleep disordered breathing.

STUDY OBJECTIVES: We sought to determine the effect of expiratory positive airway pressure on end expiratory lung volume (EELV) and sleep disordered breathing in obstructive sleep apnea patients. DESIGN: Observational physiology study PARTICIPANTS: We studied 10 OSA patients during sleep wearing a facial mask. We recorded 1 hour of NREM sleep without treatment (baseline) and 1 hour with 10 cm H2O EPAP in random order, while measuring EELV and breathing pattern. RESULTS: The mean EELV change between baseline and EPAP was only 13.3 mL (range 2-25 mL). Expiratory time was significantly increased with EPAP compared to baseline 2.64 +/- 0.54 vs 2.16 +/- 0.64 sec (P = 0.002). Total respiratory time was longer with EPAP than at baseline 4.44 +/- 1.47 sec vs 3.73 +/- 0.88 sec (P = 0.3), and minute ventilation was lower with EPAP vs baseline 7.9 +/- 4.17 L/min vs 9.05 +/- 2.85 L/min (P = 0.3). For baseline (no treatment) and EPAP respectively, the mean apnea+hypopnea index (AHI) was 62.6 +/- 28.7 and 56.8 +/- 30.3 events per hour (P = 0.4). CONCLUSION: In OSA patients during sleep, the application of 10 cm H2O EPAP led to prolongation of expiratory time with only marginal increases in FRC. These findings suggest important mechanisms exist to avoid hyperinflation during sleep.

The Ca(V)3.3 calcium channel is the major sleep spindle pacemaker in thalamus.

Low-threshold (T-type) Ca(2+) channels encoded by the Ca(V)3 genes endow neurons with oscillatory properties that underlie slow waves characteristic of the non-rapid eye movement (NREM) sleep EEG. Three Ca(V)3 channel subtypes are expressed in the thalamocortical (TC) system, but their respective roles for the sleep EEG are unclear. Ca(V)3.3 protein is expressed abundantly in the nucleus reticularis thalami (nRt), an essential oscillatory burst generator. We report the characterization of a transgenic Ca(V)3.3(-/-) mouse line and demonstrate that Ca(V)3.3 channels are indispensable for nRt function and for sleep spindles, a hallmark of natural sleep. The absence of Ca(V)3.3 channels prevented oscillatory bursting in the low-frequency (4-10 Hz) range in nRt cells but spared tonic discharge. In contrast, adjacent TC neurons expressing Ca(V)3.1 channels retained low-threshold bursts. Nevertheless, the generation of synchronized thalamic network oscillations underlying sleep-spindle waves was weakened markedly because of the reduced inhibition of TC neurons via nRt cells. T currents in Ca(V)3.3(-/-) mice were <30% compared with those in WT mice, and the remaining current, carried by Ca(V)3.2 channels, generated dendritic [Ca(2+)](i) signals insufficient to provoke oscillatory bursting that arises from interplay with Ca(2+)-dependent small conductance-type 2 K(+) channels. Finally, naturally sleeping Ca(V)3.3(-/-) mice showed a selective reduction in the power density of the σ frequency band (10-12 Hz) at transitions from NREM to REM sleep, with other EEG waves remaining unaltered. Together, these data identify a central role for Ca(V)3.3 channels in the rhythmogenic properties of the sleep-spindle generator and provide a molecular target to elucidate the roles of sleep spindles for brain function and development.

ON THE ROLE OF PERIOD-2 IN THE CIRCADIAN AND HOMEOSTATIC REGULATION OF SLEEP

Humans spend one third of their life sleeping, then we could raise the basic question: Why do we sleep? Despite the fact that we still don't fully understand its function, we made much progress in understanding at different levels how sleep is regulated. One model suggests that sleep is regulated by two processes: a homeostatic process that tracks the need for sleep and by a circadian rhythm that determines the preferred time-of-day sleep occurs. At the molecular level circadian rhythms are a property of interlocking transcriptional regula-tors referred to as clock genes. The heterodimeric transcription factors BMAL1::CLOCK/NPAS2 drive the transcription of many target genes including the clock genes Cryptochome1 (Cry1), Cry2, Period1 (Per1), and Per2. The encoded CRY/PER proteins are transcriptional inhibitors of BMAL1::CLOCK/NPAS2 thereby providing negative feedback to their own transcription. These genes seem, however, also involved in sleep homeostasis because the brain expression of clock genes, es-pecially that of Per2, increase as a function of time-spent-awake and because mice lacking clock genes display altered sleep homeostasis. The aim of first part of my doctoral work has been to advance our understanding the link that exists between sleep homeostasis and circadian rhythms investigating a possible mechanism by which sleep deprivation could alter clock gene expression by quantifying DNA-binding of the core-clock genes BMAL1, CLOCK and NPAS2 to their target chromatin loci including the E-box enhancers of the Per2 promoter. We made use of chromatin immunoprecipitation (ChIP) and quantitative poly-merase chain reaction (qPCR) to show that DNA-binding of CLOCK and BMAL1 to their target genes changes as a function of time-of-day in both liver and cerebral cortex. We then performed a 6h sleep deprivation (SD) and observed a significant decrease in DNA-binding of CLOCK and BMAL1 to Dbp. This is consistent with a decrease in Dbp mRNA levels after SD. The DNA-binding of NPAS2 and BMAL1 to Per2 was similarly decreased following SD. However, SD has been previously shown to in-crease Per2 expression in the cortex which seems paradoxical. Our results demonstrate that sleep-wake history can affect the molecular clock machinery directly at the level of the chromatin thereby altering the cortical expression of Dbp and Per2, and likely other targets. However, the precise dy-namic relationship between DNA-binding and mRNA expression, especially for Per2, remains elusive. The second aim of my doctoral work has been to perform an in depth characterization of cir-cadian rhythmicity, sleep architecture, analyze the response to SD in full null-Per2 knock-out (Per2-/-) mice, and Per1-/- mice, as well as their double knock-out offspring (Per1,2-/-) and littermate wildtype (Wt) mice. The techniques used include locomotor activity recording by passive infrared (PIR) sen-sors, EEG/EMG surgery, recording, and analysis, and cerebral cortex extraction and quantification of mRNA levels by qPCR. Under standard LD12:12 conditions, we found that wakefulness onset, as well as the time courses of clock gene expression in the brain and corticosterone plasma levels were ad-vanced by about 2h in Per2-/- mice compared to Wt mice. When released under constant dark condi-tions almost all Per2-/- mice (97%) became arrhythmic immediately. From these observations, we conclude that while Per2-/- mice seem to be able to anticipate dark onset, this does not result from a self-sustained circadian clock. Our results suggest instead that the earlier onset of activity results from a labile, not-self sustained 22h rhythm linked to light onset suggesting the existence of a light-driven rhythm. Analyses of sleep under LD12:12 conditions revealed that in both Per2-/- and Per1,2-/- mice the same sleep phenotypes are observed compared to Wt mice: increased NREM sleep frag-mentation and inability to adequately compensate the loss of NREM sleep. That suggests a possible role of PER2 in sleep consolidation and recovery.

Sleep function: current questions and new approaches.

The mammalian brain oscillates through three distinct global activity states: wakefulness, non-rapid eye movement (NREM) sleep and REM sleep. The regulation and function of these 'vigilance' or 'behavioural' states can be investigated over a broad range of temporal and spatial scales and at different levels of functional organization, i.e. from gene expression to memory, in single neurons, cortical columns or the whole brain and organism. We summarize some basic questions that have arisen from recent approaches in the quest for the functions of sleep. Whereas traditionally sleep was viewed to be regulated through top-down control mechanisms, recent approaches have emphasized that sleep is emerging locally and regulated in a use-dependent (homeostatic) manner. Traditional markers of sleep homeostasis, such as the electroencephalogram slow-wave activity, have been linked to changes in connectivity and plasticity in local neuronal networks. Thus waking experience-induced local network changes may be sensed by the sleep homeostatic process and used to mediate sleep-dependent events, benefiting network stabilization and memory consolidation. Although many questions remain unanswered, the available data suggest that sleep function will best be understood by an analysis which integrates sleep's many functional levels with its local homeostatic regulation.

Differential effects of sodium oxybate and baclofen on EEG, sleep, neurobehavioral performance, and memory.

STUDY OBJECTIVES: Sodium oxybate (SO) is a GABA(B) agonist used to treat the sleep disorder narcolepsy. SO was shown to increase slow wave sleep (SWS) and EEG delta power (0.75-4.5 Hz), both indexes of NREM sleep (NREMS) intensity and depth, suggesting that SO enhances recuperative function of NREM. We investigated whether SO induces physiological deep sleep. DESIGN: SO was administered before an afternoon nap or before the subsequent experimental night in 13 healthy volunteers. The effects of SO were compared to baclofen (BAC), another GABA(B) receptor agonist, to assess the role of GABA(B) receptors in the SO response. MEASUREMENTS AND RESULTS: As expected, a nap significantly decreased sleep need and intensity the subsequent night. Both drugs reversed this nap effect on the subsequent night by decreasing sleep latency and increasing total sleep time, SWS during the first NREMS episode, and EEG delta and theta (0.75-7.25 Hz) power during NREMS. The SO-induced increase in EEG delta and theta power was, however, not specific to NREMS and was also observed during REM sleep (REMS) and wakefulness. Moreover, the high levels of delta power during a nap following SO administration did not affect delta power the following night. SO and BAC taken before the nap did not improve subsequent psychomotor performance and subjective alertness, or memory consolidation. Finally, SO and BAC strongly promoted the appearance of sleep onset REM periods. CONCLUSIONS: The SO-induced EEG slow waves seem not to be functionally similar to physiological slow waves. Our findings also suggest a role for GABA(B) receptors in REMS generation. CITATION: Vienne J; Lecciso G; Constantinescu I; Schwartz S; Franken P; Heinzer R; Tafti M. Differential effects of sodium oxybate and baclofen on EEG, sleep, neurobehavioral performance, and memory. SLEEP 2012;35(8):1071-1084.

Stimulus-induced, sleep-bound, focal seizures: a case report.

STUDY OBJECTIVES: In nocturnal frontal lobe epilepsy (NFLE), seizures occur almost exclusively during NREM sleep. Why precisely these seizures are sleep-bound remains unknown. Studies of patients with nonlesional familial forms of NFLE have suggested the arousal system may play a major role in their pathogenesis. We report the case of a patient with pharmaco-resistant, probably cryptogenic form of non-familial NFLE and strictly sleep-bound seizures that could be elicited by alerting stimuli and were associated with ictal bilateral thalamic and right orbital-insular hyperperfusion on SPECT imaging. DESIGN: Case report. SETTING: University Hospital Zurich. PATIENTS OR PARTICIPANTS: One patient with pharmaco-resistant epilepsy. CONCLUSION: This case shows that the arousal system plays a fundamental role also in cryptogenic non-familial forms of NFLE.

Altered Sleep Homeostasis in Rev-erbα Knockout Mice.

STUDY OBJECTIVES: The nuclear receptor REV-ERBα is a potent, constitutive transcriptional repressor critical for the regulation of key circadian and metabolic genes. Recently, REV-ERBα's involvement in learning, neurogenesis, mood, and dopamine turnover was demonstrated suggesting a specific role in central nervous system functioning. We have previously shown that the brain expression of several core clock genes, including Rev-erbα, is modulated by sleep loss. We here test the consequences of a loss of REV-ERBα on the homeostatic regulation of sleep. METHODS: EEG/EMG signals were recorded in Rev-erbα knockout (KO) mice and their wild type (WT) littermates during baseline, sleep deprivation, and recovery. Cortical gene expression measurements after sleep deprivation were contrasted to baseline. RESULTS: Although baseline sleep/wake duration was remarkably similar, KO mice showed an advance of the sleep/wake distribution relative to the light-dark cycle. After sleep onset in baseline and after sleep deprivation, both EEG delta power (1-4 Hz) and sleep consolidation were reduced in KO mice indicating a slower increase of homeostatic sleep need during wakefulness. This slower increase might relate to the smaller increase in theta and gamma power observed in the waking EEG prior to sleep onset under both conditions. Indeed, the increased theta activity during wakefulness predicted delta power in subsequent NREM sleep. Lack of Rev-erbα increased Bmal1, Npas2, Clock, and Fabp7 expression, confirming the direct regulation of these genes by REV-ERBα also in the brain. CONCLUSIONS: Our results add further proof to the notion that clock genes are involved in sleep homeostasis. Because accumulating evidence directly links REV-ERBα to dopamine signaling the altered homeostatic regulation of sleep reported here are discussed in that context.