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Implantable optical technologies provide measurements of cerebral hemodynamic activity from freely behaving animals without movement constraint or anesthesia. In order to study state-dependent neural evoked responses and the consequential hemodynamic response, we simultaneously measured EEG and scattered light changes in chronically implanted rats. Recordings took place under freely behaving conditions, allowing us to compare the evoked responses across wake, sleep, and anesthetized states. The largest evoked electrical and optical responses occurred during quiet sleep compared to wake and REM sleep while isoflurane anesthesia showed a large, late burst of electrical activity synchronized to the stimulus, but an earlier optical response.

Understanding neurovascular coupling is a prerequisite for the interpretation of results obtained from modern neuroimaging techniques. This study investigated the hemodynamic and neural responses in rat somatosensory cortex elicited by 16 seconds electrical whisker stimuli. Hemodynamics were measured by optical imaging spectroscopy and neural activity by multichannel electrophysiology. Previous studies have suggested that the whisker-evoked hemodynamic response contains two mechanisms, a transient ‘backwards' dilation of the middle cerebral artery, followed by an increase in blood volume localized to the site of neural activity. To distinguish between the mechanisms responsible for these aspects of the response, we presented whisker stimuli during normocapnia (‘control'), and during a high level of hypercapnia. Hypercapnia was used to ‘predilate' arteries and thus possibly ‘inhibit' aspects of the response related to the ‘early' mechanism. Indeed, hemodynamic data suggested that the transient stimulus-evoked response was absent under hypercapnia. However, evoked neural responses were also altered during hypercapnia and convolution of the neural responses from both the normocapnic and hypercapnic conditions with a canonical impulse response function, suggested that neurovascular coupling was similar in both conditions. Although data did not clearly dissociate early and late vascular responses, they suggest that the neurovascular coupling relationship is neurogenic in origin.

We recorded the electroencephalographic (EEG) responses evoked by transcranial magnetic stimulation (TMS) during the first rapid eye movement (REM) sleep episode of the night and we compared them with the responses obtained during previous wakefulness and NREM sleep. Confirming previous findings, upon falling into NREM sleep, cortical activations became more local and stereotypical, indicating a significant impairment of the intracortical dialogue. During REM sleep, a state in which subjects regain consciousness but are almost paralyzed, TMS triggered more widespread and differentiated patterns of cortical activation, that were similar to the ones observed in wakefulness. Similarly, TMS/hd-EEG may be used to probe the internal dialogue of the thalamocortical system in brain injured patients that are unable to move and communicate.

Objective

It has been hypothesized that slow wave activity, a well established measure of sleep homeostasis that increases after waking and decreases after sleep, may reflect changes in cortical synaptic strength. If so, the amplitude of sensory evoked responses should also vary as a function of time awake and asleep in a way that reflects sleep homeostasis.

Methods

Using 256-channel, high-density electroencephalography (EEG) in 12 subjects, auditory evoked potentials (AEP) and spontaneous waking data were collected during wakefulness before and after sleep.

Results

The amplitudes of the N1 and P2 waves of the AEP were reduced after a night of sleep. In addition, the decline in N1 amplitude correlated with low-frequency EEG power during non-rapid eye movement sleep and spontaneous wakefulness, both homeostatically regulated measures of sleep need.

Conclusion

The decline in AEP amplitude after a night of sleep may reflect a homeostatic reduction in synaptic strength.

Significance

These findings provide further evidence for a connection between synaptic plasticity and sleep homeostasis.

Cortical evoked response potentials (ERPs) display a rich set of waveforms that are both context and state dependent. However, the mechanisms that underlie state dependent ERP patterns are unclear. Determining those mechanisms through analysis of single trial ERP waveform signatures may provide insight into the regulation of cortical column state and the roles that sleep plays in cortical function. We implanted rats with EEG and EMG electrodes to record ERPs and to assess sleep/wake states continuously during 1-2s random auditory clicks. Individual cortical auditory ERPs were sorted into one of eight behavioral states, and fell into three categories based on amplitude and latency characteristics. ERPs within waking and rapid eye movement (REM) sleep were predominately low amplitude and short latency. Approximately 50% of ERPs during light quiet sleep (QS1 and QS2) exhibited low amplitude, short latency responses, and the remaining ERPs had high amplitude, long latency responses. This distribution was characteristic of EEG fluctuations during low frequency delta waves. Significantly more individual ERPs showed very low amplitudes during deep quiet sleep (QS3 and QS4), resulting in a lower average ERP. These results support the hypothesis that evoked response amplitudes and waveform patterns follow specific EEG patterns. Since evoked response characteristics distribute differently across states, they could aid our understanding of sleep mechanisms through state related and local neural signaling.

Substantial evidence suggests that brain regions that have been disproportionately used during waking will require a greater intensity and/or duration of subsequent sleep. For example, rats use their whiskers in the dark and their eyes during the light which manifests as a greater magnitude of electroencephalogram (EEG) slow wave activity in the somatosensory and visual cortex during sleep in the corresponding light and dark periods respectively. The parsimonious interpretation of such findings is that sleep is distributed across local brain regions and is use-dependent. The fundamental properties of sleep can also be experimentally defined locally at the level of small neural assemblies such as cortical columns. In this view, sleep is orchestrated, but not fundamentally driven, by central mechanisms. We explore two physiological markers of local, use-dependent sleep, namely, an electrical marker apparent as a change in the size and shape of an electrical evoked response, and a metabolic marker evident as an evoked change in blood volume and oxygenation delivered to activated tissue. Both markers, applied to cortical columns, provide a means to investigate physiological mechanisms for the distributed homeostatic regulation of sleep, and may yield new insights into the consequences of sleep loss and sleep pathologies on waking brain function.

Norepinephrine (NE)-containing locus ceruleus (LC) has been known to participate in the regulation of the sleep-wake cycle according to the differential firing rate. The aim of this study was to know the change of extracellular NE level in the rat amygdala, which are reciprocally connected with LC, during sleep-wakefulness. Extracellular NE levels in the rat amygdala were investigated during different stages of the sleep-waking cycle using in vivo microdialysis and polygraphic recording. Dialysates were collected every 5 min and correlated with the results of polygraphic recording. The content of NE was measured by high-performance liquid chromatography with electrochemical detection. NE level was the highest in active waking (AW) and, when compared to AW, NE level was progressively lower in quiet waking (QW; 86%), quiet sleep (QS; 72%), and active sleep (AS or REM sleep; 61%). This result suggests that the rat amygdala also participates in the regulation of the sleep-wake cycle according to the differential NE release.

The function of the brain activity that defines slow wave sleep (SWS) and rapid eye movement (REM) sleep in mammals is unknown. During SWS, the level of electroencephalogram slow wave activity (SWA or 0.5–4.5 Hz power density) increases and decreases as a function of prior time spent awake and asleep, respectively. Such dynamics occur in response to waking brain use, as SWA increases locally in brain regions used more extensively during prior wakefulness. Thus, SWA is thought to reflect homeostatically regulated processes potentially tied to maintaining optimal brain functioning. Interestingly, birds also engage in SWS and REM sleep, a similarity that arose via convergent evolution, as sleeping reptiles and amphibians do not show similar brain activity. Although birds deprived of sleep show global increases in SWA during subsequent sleep, it is unclear whether avian sleep is likewise regulated locally. Here, we provide, to our knowledge, the first electrophysiological evidence for local sleep homeostasis in the avian brain. After staying awake watching David Attenborough's The Life of Birds with only one eye, SWA and the slope of slow waves (a purported marker of synaptic strength) increased only in the hyperpallium—a primary visual processing region—neurologically connected to the stimulated eye. Asymmetries were specific to the hyperpallium, as the non-visual mesopallium showed a symmetric increase in SWA and wave slope. Thus, hypotheses for the function of mammalian SWS that rely on local sleep homeostasis may apply also to birds.

The aim of this study was to elucidate physiological processes that are involved in the homeostatic regulation of REM sleep. Adult rats were chronically instrumented with sleep–wake recording electrodes. Following post-surgical recovery, rats were habituated extensively for freely moving polygraphic recording conditions. On the first experimental recording day (baseline day, BLD), polygraphic signs of undisturbed sleep–wake activities were recorded for 4 h (between 11:00 AM and 3:00 PM). During the second experimental recording day (REM sleep deprivation day, RDD), rats were selectively deprived of REM sleep for the first 2 h and then allowed to have normal sleep–wake for the following 2 h. The results demonstrated that during the first 2 h, compared to BLD, RDD recordings exhibited 87.80% less time in REM sleep and 16% more time in non-REM (NREM) sleep. The total percentages of wakefulness remained comparable between the BLD and RDD. During the RDD, the mean number of REM sleep episodes was much higher than in the BLD, indicating increased REM sleep drive. Electroencephalographic (EEG) power spectral analysis revealed that selective REM sleep deprivation increased delta power but decreased theta power during the residual REM sleep. During the last 2 h, after REM sleep deprivation, rats spent 51% more time in REM sleep compared to the BLD. Also during this period, the number of REM sleep episodes with the shortest (5–30 s) and longest (>120 s) duration increased during the RDD. These findings suggest that the REM sleep homeostatic process involves increased delta- and decreased theta-frequency wave activities in the cortical EEG.

We hypothesized that adenosine, acting via the A1 receptor, is a key factor in the homeostatic control of sleep. The increase in extracellular levels of adenosine during prolonged wakefulness is thought to facilitate the transition to sleep by reducing the discharge activity of wakefulness-promoting neurons in the basal forebrain. Adenosine A1 receptor control of the homeostatic regulation of sleep was tested by microdialysis perfusion of antisense oligonucleotides against the mRNA of the A1 receptor in the magnocellular cholinergic region of the basal forebrain of freely behaving rats. After microdialysis perfusion of A1 receptor antisense in the basal forebrain, spontaneous levels of sleep-wakefulness showed a significant reduction in non-rapid eye movement (REM) sleep with an increase in wakefulness. After 6 hr of sleep deprivation, the antisense-treated animals spent a significantly reduced amount of time in non-REM sleep, with postdeprivation recovery sleep hours 2−5 showing a reduction of ∼50−60%. There was an even greater postdeprivation reduction in delta power (60−75%) and a concomitant increase in wakefulness. All behavioral state changes returned to control (baseline) values after the cessation of antisense administration. Control experiments with microdialysis perfusion of nonsense (randomized antisense) oligonucleotides and with artificial CSF showed no effect during postdeprivation recovery sleep or spontaneously occurring behavioral states. Antisense to the A1 receptor suppressed A1 receptor immunoreactivity but did not show any neurotoxicity as visualized by Fluoro-Jade staining. These data support our hypothesis that adenosine, acting via the A1 receptor, in the basal forebrain is a key component in the homeostatic regulation of sleep.

The principal site that generates both REM sleep and wakefulness is located in the mesopontine reticular formation, whereas non-REM sleep (NREM) is primarily dependent upon the functioning of neurons that are located in the preoptic region of the hypothalamus. In the present study, we were interested in determining whether the occurrence of NREM might also depend on the activity of mesopontine structures, as has been shown for wakefulness and REM sleep.

Adult cats were maintained in one of the following states: quiet wakefulness (QW), alert wakefulness (AW), NREM, or REM sleep induced by microinjections of carbachol into the nucleus pontis oralis (REM-carbachol). Subsequently, they were euthanized and single labeling immunohistochemical studies were undertaken to determine state-dependent patterns of neuronal activity in the brainstem based upon the expression of the protein Fos. In addition, double labeling immunohistochemical studies were carried out to detect neurons that expressed Fos as well as choline acetyltransferase, tyrosine hydroxylase or GABA.

During NREM, only a few Fos immunoreactive cells were present in different regions of the brainstem; however, a discrete cluster of Fos+ neurons was observed in the caudolateral peribrachial region (CLPB). The number of the Fos+ neurons in the CLPB during NREM was significantly greater (67.9 ± 10.9, P < 0.0001) compared to QW (8.0 ± 6.7), AW (5.2 ± 4.2) or REM-carbachol (8.0 ± 4.7). In addition, there was a positive correlation (R = 0.93) between the time the animals spent in NREM and the number of Fos+ neurons in the CLPB. Fos-immunoreactive neurons in the CLPB were neither cholinergic nor catecholaminergic; however about 50% of these neurons were GABAergic.

We conclude that a group of GABAergic and unidentified neurons in the CLPB are active during NREM and likely involved in the control of this behavioral state. These data open new avenues for the study of NREM, as well as for the explorations of interactions between these neurons that are activated during NREM, and cells of the adjacent pontine tegmentum that are involved in the generation of REM sleep.

Rationale: The ability of patients with central hypoventilation syndrome (CHS) to produce and process mechanoreceptor signals is unknown.

Objectives: Children with CHS hypoventilate during sleep, although they generally breathe adequately during wakefulness. Previous studies suggest that they have compromised central integration of afferent stimuli, rather than abnormal sensors or receptors. Cortical integration of afferent mechanical stimuli caused by respiratory loading or upper airway occlusion can be tested by measuring respiratory-related evoked potentials (RREPs). We hypothesized that patients with CHS would have blunted RREP during both wakefulness and sleep.

Methods: RREPs were produced with multiple upper airway occlusions and were obtained during wakefulness, stage 2, slow-wave, and REM sleep. Ten patients with CHS and 20 control subjects participated in the study, which took place at the Children's Hospital of Philadelphia. Each patient was age- and sex-matched to two control subjects. Wakefulness data were collected from 9 patients and 18 control subjects.

Measurements and Main Results: During wakefulness, patients demonstrated reduced Nf and P300 responses compared with control subjects. During non-REM sleep, patients demonstrated a reduced N350 response. In REM sleep, patients had a later P2 response.

Conclusions: CHS patients are able to produce cortical responses to mechanical load stimulation during both wakefulness and sleep; however, central integration of the afferent signal is disrupted during wakefulness, and responses during non-REM are damped relative to control subjects. The finding of differences between patients and control subjects during REM may be due to increased intrinsic excitatory inputs to the respiratory system in this state.

Averaged event-related potentials (ERPs) represent sensory and cognitive processing of stimuli during wakefulness independent of behavioural responses, and reflect the underlying state of the CNS during sleep. Components measured during wakefulness which are reflective of arousal state or the automatic switching of attention are sensitive to prior sleep disruption. Components reflecting active attentional influences during the waking state appear to be preserved in a rudimentary form during REM sleep, but in a way that highlights the differences in the neurochemical environment between wakefulness and REM sleep. Certain ERP components only appear within sleep. These begin to emerge at NREM sleep onset and may reflect inhibition of information processing and thus have utility as markers of the functional status of sleep preparatory mechanisms. These large amplitude NREM components represent synchronized burst firing of large number of cortical cells and are a reflection of the nervous system’s capacity to generate delta frequency EEG activity. As such they are useful in assessing the overall integrity of the nervous system in populations not showing substantial amounts of SWS as measured using traditional criteria. While requiring care in their interpretation, ERPs nonetheless provide a rich tool to investigators interested in probing the nervous system to evaluate daytime functioning in the face of sleep disruption, the ability of the sleeping nervous system to monitor the external environment, and the ability of the nervous system to respond to stimuli in a manner consistent with the initiation or maintenance of sleep.

Studies have shown that sleep recovery following different protocols of forced waking varies according to the level of stress inherent to each method. Sleep deprivation activates the hypothalamic-pituitary-adrenal axis and increased corticotropin-releasing hormone (CRH) impairs sleep. The purpose of the present study was to evaluate how manipulations of the CRH system during the sleep deprivation period interferes with subsequent sleep rebound. Throughout 96 hours of sleep deprivation, separate groups of rats were treated i.c.v. with vehicle, CRH or with alphahelical CRH9−41, a CRH receptor blocker, twice/day, at 07:00 h and 19:00 h. Both treatments impaired sleep homeostasis, especially in regards to length of rapid eye movement sleep (REM) and theta/delta ratio and induced a later decrease in NREM and REM sleep and increased waking bouts. These changes suggest that activation of the CRH system impact negatively on the homeostatic sleep response to prolonged forced waking. These results indicate that indeed, activation of the HPA axis—at least at the hypothalamic level—is capable to reduce the sleep rebound induced by sleep deprivation.

Sleep enhances memories, particularly emotional memories. As such, it has been suggested that sleep deprivation may reduce post-traumatic stress disorder. This presumes that emotional memory consolidation is paralleled by a reduction in emotional reactivity, an association that has not yet been examined. In the present experiment, we utilized an incidental memory task in humans and obtained valence and arousal ratings during two sessions separated either by 12 hours of daytime wake or 12 hours including overnight sleep. Recognition accuracy was greater following sleep relative to wake for both negative and neutral pictures. While emotional reactivity to negative pictures was greatly reduced over wake, the negative emotional response was relatively preserved over sleep. Moreover, protection of emotional reactivity was associated with greater time in REM sleep. Recognition accuracy, however, was not associated with REM. Thus, we provide the first evidence that sleep enhances emotional memory while preserving emotional reactivity.

Background

Upper airway collapse does not occur during wake in obstructive sleep apnea patients. This points to wake-related compensatory mechanisms, and possibly to a modified corticomotor control of upper airway dilator muscles. The objectives of the study were to characterize the responsiveness of the genioglossus to transcranial magnetic stimulation during respiratory and non-respiratory facilitatory maneuvers in obstructive sleep apnea patients, and to compare it to the responsiveness of the diaphragm, with reference to normal controls.

Methods

Motor evoked potentials of the genioglossus and of the diaphragm, with the corresponding motor thresholds, were recorded in response to transcranial magnetic stimulation applied during expiration, inspiration and during maximal tongue protraction in 13 sleep apnea patients and 8 normal controls.

Main Results

In the sleep apnea patients: 1) combined genioglossus and diaphragm responses occurred more frequently than in controls (P < 0.0001); 2) the amplitude of the genioglossus response increased during inspiratory maneuvers (not observed in controls); 3) the latency of the genioglossus response decreased during tongue protraction (not observed in controls). A significant negative correlation was found between the latency of the genioglossus response and the apnea-hypopnea index; 4) the difference in diaphragm and genioglossus cortico-motor responses during tongue protraction and inspiratory loading differed between sleep apnea and controls.

Conclusion

Sleep apnea patients and control subjects differ in the response pattern of the genioglossus and of the diaphragm to facilitatory maneuvers, some of the differences being related to the frequency of sleep-related events.

Understanding the interaction between the nervous system and cerebral vasculature is fundamental to forming a complete picture of the neurophysiology of sleep and its role in maintaining physiological homeostasis. However, the intrinsic hemodynamics of slow-wave sleep (SWS) are still poorly known. We carried out 30 all-night sleep measurements with combined near-infrared spectroscopy (NIRS) and polysomnography to investigate spontaneous hemodynamic behavior in SWS compared to light (LS) and rapid-eye-movement sleep (REM). In particular, we concentrated on slow oscillations (3–150 mHz) in oxy- and deoxyhemoglobin concentrations, heart rate, arterial oxygen saturation, and the pulsation amplitude of the photoplethysmographic signal. We also analyzed the behavior of these variables during sleep stage transitions. The results indicate that slow spontaneous cortical and systemic hemodynamic activity is reduced in SWS compared to LS, REM, and wakefulness. This behavior may be explained by neuronal synchronization observed in electrophysiological studies of SWS and a reduction in autonomic nervous system activity. Also, sleep stage transitions are asymmetric, so that the SWS-to-LS and LS-to-REM transitions, which are associated with an increase in the complexity of cortical electrophysiological activity, are characterized by more dramatic hemodynamic changes than the opposite transitions. Thus, it appears that while the onset of SWS and termination of REM occur only as gradual processes over time, the termination of SWS and onset of REM may be triggered more abruptly by a particular physiological event or condition. The results suggest that scalp hemodynamic changes should be considered alongside cortical hemodynamic changes in NIRS sleep studies to assess the interaction between the autonomic and central nervous systems.

Background

Sleep is regulated by both a circadian and a homeostatic process. The homeostatic process reflects the duration of prior wakefulness: the longer one stays awake, the longer and/or more intense is subsequent sleep. In mammals, the best marker of the homeostatic sleep drive is slow wave activity (SWA), the electroencephalographic (EEG) power spectrum in the 0.5–4 Hz frequency range during non-rapid eye movement (NREM) sleep. In mammals, NREM sleep SWA is high at sleep onset, when sleep pressure is high, and decreases progressively to reach low levels in late sleep. Moreover, SWA increases further with sleep deprivation, when sleep also becomes less fragmented (the duration of sleep episodes increases, and the number of brief awakenings decreases). Although avian and mammalian sleep share several features, the evidence of a clear homeostatic response to sleep loss has been conflicting in the few avian species studied so far. The aim of the current study was therefore to ascertain whether established markers of sleep homeostasis in mammals are also present in the white-crowned sparrow (Zonotrichia leucophrys gambelii), a migratory songbird of the order Passeriformes. To accomplish this goal, we investigated amount of sleep, sleep time course, and measures of sleep intensity in 6 birds during baseline sleep and during recovery sleep following 6 hours of sleep deprivation.

Results

Continuous (24 hours) EEG and video recordings were used to measure baseline sleep and recovery sleep following short-term sleep deprivation. Sleep stages were scored visually based on 4-sec epochs. EEG power spectra (0.5–25 Hz) were calculated on consecutive 4-sec epochs. Four vigilance states were reliably distinguished based on behavior, visual inspection of the EEG, and spectral EEG analysis: Wakefulness (W), Drowsiness (D), slow wave sleep (SWS) and rapid-eye movement (REM) sleep. During baseline, SWA during D, SWS, and NREM sleep (defined as D and SWS combined) was highest at the beginning of the major sleep period and declined thereafter. Moreover, peak SWA in both SWS and NREM sleep increased significantly immediately following sleep deprivation relative to baseline.

Conclusion

As in mammals, sleep deprivation in the white-crowned sparrow increases the intensity of sleep as measured by SWA.

Objective

To determine if resistance to weight gain is associated with alterations in sleep/wake states and orexin receptor gene expression.

Design

Three-month old obesity susceptible Sprague-Dawley (SD) and obesity resistant (OR) rats were fed standard rodent chow. Sleep/wake cycle was measured by radiotelemetry and orexin receptor profiles in sleep/wake regulatory areas of the brain were quantified by quantitative RT-PCR.

Subjects

Adult male obesity susceptible SD and selectively-bred OR rats.

Measurements

Body weight, food intake, energy efficiency, percent time spent in active wake, quiet wake, slow-wave sleep (SWS), rapid eye movement (REM) sleep, number and mean duration of sleep/wake episodes, number of stage transitions, SWS sleep delta power and orexin receptor mRNA levels were measured.

Results

Obesity resistant rats weighed significantly less and had lower energy efficiency than SD rats. Food intake was not different between SD and OR rats. Time spent in quiet wake was similar between groups, and therefore active wake and quiet wake were combined and are referred to as ‘wakefulness’. Obesity resistant rats spent significantly more time in wakefulness and less time in SWS compared to SD rats during the 24 h recording period. Relative to SD rats, OR rats had significantly fewer sleep/wake episodes and the duration of the episodes were prolonged, indicating less fragmented sleep. Further, OR rats had fewer transitions between sleep stages, which indicates that OR rats were behaviorally more stable and had more consolidated sleep than obesity susceptible SD rats. Obesity resistant rats exhibited lower delta power during SWS sleep, indicating a lower sleep drive. Our results demonstrated greater orexin receptor gene expression in sleep regulatory brain areas in OR rats.

Conclusion

These results demonstrate that prolonged wakefulness, better sleep quality, lower sleep drive and greater orexin signaling may confer protection against obesity.

Previous studies have supported the hypothesis that macromolecular synthesis occurs in the brain during sleep as a response to prior waking activities and that prostaglandin D2 (PGD2) is an endogenous sleep substance whose effects are dependent on adenosine A2a receptor-mediated signaling. We compared gene expression in the cerebral cortex, basal forebrain and hypothalamus during PGD2-induced and adenosinergically-induced sleep to results from our previously-published study of recovery sleep (RS) after sleep deprivation (SD). Immediate early gene (IEG) expression in the cortex during sleep induced by PGD2- or by the selective adenosine A2a agonist CGS21680 showed limited similarity to that observed during RS while, in the basal forebrain and hypothalamus, widespread activation of IEGs not seen during RS occurred. In all three brain regions, PGD2 and CGS21680 reduced the expression of arc, a transcript whose expression is elevated during SD. Using GeneChips®, the majority of genes induced by either PGD2 or CGS21680 were induced by both, suggesting activation of the same pathways. However, gene expression induced in the brain after PGD2 or CGS21680 treatment was distinct from that described during RS after SD and apparently involves glial cell gene activation and signaling pathways in neural-immune interactions.

Hypocretins (orexins) are hypothalamic neuropeptides that play a crucial role in regulating sleep/wake states and autonomic functions including parasympathetic cardiac activity. We have recently demonstrated stimulation of the lateral paragigantocellular nucleus (LPGi), the nucleus which is thought to play a role in rapid eye movement (REM) sleep control, activates an inhibitory pathway to preganglionic cardiac vagal neurons in the nucleus ambiguus (NA). In this study we test the hypothesis that hypocretin-1 modulates the inhibitory neurotransmission to cardiac vagal neurons evoked by stimulation of the LPGi using whole-cell patch-clamp recordings in an in vitro brain slice preparation from rats. Activation of hypocretin-1 receptors produced a dose-dependent and long-term facilitation of GABAergic postsynaptic currents evoked by electrical stimulation of the LPGi. Hypoxia/hypercapnia diminished LPGi-evoked GABAergic current in cardiac vagal neurons and this inhibition by hypoxia/hypercapnia was prevented by pre-application of hypocretin-1. The action of hypocretin-1 was blocked by the hypocretin-1 receptor antagonist SB-334867. Facilitation of LPGi-evoked GABAergic current in cardiac vagal neurons under both normal condition and during hypoxia/hypercapnia could be the mechanism by which hypocretin-1 affects parasympathetic cardiac function and heart rate during REM sleep. Furthermore, our findings indicate a new potential mechanism that might be involved in the cardiac arrhythmias, bradycardia, and sudden cardiac death that can occur during sleep.

Starvation, which is common in the wild, appears to initiate a genetic program that allows fruitflies to remain awake without the sleepiness and cognitive impairments that typically follow sleep deprivation.

Extended periods of waking result in physiological impairments in humans, rats, and flies. Sleep homeostasis, the increase in sleep observed following sleep loss, is believed to counter the negative effects of prolonged waking by restoring vital biological processes that are degraded during sleep deprivation. Sleep homeostasis, as with other behaviors, is influenced by both genes and environment. We report here that during periods of starvation, flies remain spontaneously awake but, in contrast to sleep deprivation, do not accrue any of the negative consequences of prolonged waking. Specifically, the homeostatic response and learning impairments that are a characteristic of sleep loss are not observed following prolonged waking induced by starvation. Recently, two genes, brummer (bmm) and Lipid storage droplet 2 (Lsd2), have been shown to modulate the response to starvation. bmm mutants have excess fat and are resistant to starvation, whereas Lsd2 mutants are lean and sensitive to starvation. Thus, we hypothesized that bmm and Lsd2 may play a role in sleep regulation. Indeed, bmm mutant flies display a large homeostatic response following sleep deprivation. In contrast, Lsd2 mutant flies, which phenocopy aspects of starvation as measured by low triglyceride stores, do not exhibit a homeostatic response following sleep loss. Importantly, Lsd2 mutant flies are not learning impaired after sleep deprivation. These results provide the first genetic evidence, to our knowledge, that lipid metabolism plays an important role in regulating the homeostatic response and can protect against neuronal impairments induced by prolonged waking.

Author Summary

It is well established in humans that sleep deficits lead to adverse outcomes, including cognitive impairments and an increased risk for obesity. Given the relationship between sleep and lipid stores, we hypothesized that metabolic pathways play a role in sleep regulation and contribute to deficits induced by sleep loss. Since starvation has a large impact on metabolic pathways and is an environmental condition that is encountered by animals living in the wild, we examined its effects on sleep in the fruit fly Drosophila melanogaster. Interestingly, when flies are starved they display an immediate increase in waking. However, in contrast to sleep deprivation, waking induced by starvation does not result in increased sleepiness or impairments in short-term memory. To identify the mechanisms underlying these processes, we evaluated mutants for genes that have been shown to alter an animal's response to starvation. Interestingly, brummer mutants, which are fat, show an exaggerated response to sleep loss. In contrast, mutants for Lipid storage droplet 2 are lean and are able to stay awake without becoming sleepy or showing signs of cognitive impairment. These results indicate that while sleep loss can alter lipids, lipid enzymes may, in turn, play a role in regulating sleep and influence the response to sleep deprivation.

Studies using drugs that increase or decrease GABAergic transmission suggest that GABA in the pontine reticular formation (PRF) promotes wakefulness and inhibits rapid eye movement (REM) sleep. Cholinergic transmission in the PRF promotes REM sleep, and levels of endogenous acetylcholine (ACh) in the PRF are significantly greater during REM sleep than during wakefulness or non-REM (NREM) sleep. No previous studies have determined whether levels of endogenous GABA in the PRF vary as a function of sleep and wakefulness. This study tested the hypothesis that GABA levels in cat PRF are greatest during wakefulness and lowest during REM sleep. Extracellular GABA levels were measured during wakefulness, NREM sleep, REM sleep, and the REM sleep-like state (REMNeo) caused by microinjecting neostigmine into the PRF. GABA levels varied significantly as a function of sleep and wakefulness, and decreased significantly below waking levels during REM sleep (−42%) and REMNeo (−63%). The decrease in GABA levels during NREM sleep (22% below waking levels) was not statistically significant. Compared to NREM sleep, GABA levels decreased significantly during REM sleep (−27%) and REMNeo (−52%). Comparisons of REM sleep and REMNeo revealed no differences in GABA levels or cortical EEG power. GABA levels did not vary significantly as a function of dialysis site within the PRF. The inverse relationship between changes in PRF levels of GABA and ACh during REM sleep indicates that low GABAergic tone combined with high cholinergic tone in the PRF contributes to the generation of REM sleep.

During slow-wave sleep and REM sleep, hippocampal place cells in the rat show replay of sequences previously observed during waking. We tested the hypothesis from computational modelling that the temporal structure of REM sleep replay could arise from an interplay of place cells with head direction cells in the postsubiculum. Physiological single-unit recording was performed simultaneously from five or more head direction or place by head direction cells in the postsubiculum during running on a circular track allowing sampling of a full range of head directions, and during sleep periods before and after running on the circular track. Data analysis compared the spiking activity during individual REM periods with waking as in previous analysis procedures for REM sleep. We also used a new procedure comparing groups of similar runs during waking with REM sleep periods. There was no consistent evidence for a statistically significant correlation of the temporal structure of spiking during REM sleep with spiking during waking running periods. Thus, the spiking activity of head direction cells during REM sleep does not show replay of head direction cell activity occurring during a previous waking period of running on the task. In addition, we compared the spiking of postsubiculum neurons during hippocampal sharp wave ripple events. We show that head direction cells are not activated during sharp wave ripples, while neurons responsive to place in the postsubiculum show reliable spiking at ripple events.

The perifornical–lateral hypothalamic area (PF/LH) contains neuronal groups playing an important role in control of waking and sleep. Among the brain regions that regulate behavioral states, one of the strongest sources of projections to the PF/LH is the median preoptic nucleus (MnPN) containing a sleep-active neuronal population. To evaluate the role of MnPN afferents in the control of PF/LH neuronal activity, we studied the responses of PF/LH cells to electrical stimulation or local chemical manipulation of the MnPN in freely moving rats. Single-pulse electrical stimulation evoked responses in 79% of recorded PF/LH neurons. No cells were activated antidromically. Direct and indirect transynaptic effects depended on sleep–wake discharge pattern of PF/LH cells. The majority of arousal-related neurons, that is, cells discharging at maximal rates during active waking (AW) or during AW and rapid eye movement (REM) sleep, exhibited exclusively or initially inhibitory responses to stimulation. Sleep-related neurons, the cells with elevated discharge during non-REM and REM sleep or selectively active in REM sleep, exhibited exclusively or initially excitatory responses. Activation of the MnPN via microdialytic application of L-glutamate or bicuculline resulted in reduced discharge of arousal-related and in excitation of sleep-related PF/LH neurons. Deactivation of the MnPN with muscimol caused opposite effects. The results indicate that the MnPN contains subset(s) of neurons, which exert inhibitory control over arousal-related and excitatory control over sleep-related PF/LH neurons. We hypothesize that MnPN sleep-active neuronal group has both inhibitory and excitatory outputs that participate in the inhibitory control of arousal-promoting PF/LH mechanisms.