Control of cardiovascular variability during undisturbed wake-sleep behavior in hypocretin-deficient mice

The central neural mechanisms underlying differences in cardiovascular variability between wakefulness, non-rapid-eye-movement sleep (NREMS), and rapid-eye-movement sleep (REMS) remain poorly understood. These mechanisms may involve hypocretin (HCRT)/orexin signaling. HCRT signaling is linked to wake-sleep states, involved in central autonomic control, and impaired in narcoleptic patients. Thus, we investigated whether HCRT signaling plays a role in controlling cardiovascular variability during spontaneous behavior in HCRT-deficient mice. HCRT-ataxin3 transgenic mice lacking HCRT neurons (TG), knockout mice lacking HCRT peptides (KO), and wild-type controls (WT) were instrumented with electrodes for sleep recordings and a telemetric blood pressure transducer. Fluctuations of systolic blood pressure (SBP) and heart period (HP) during undisturbed wake-sleep behavior were analyzed with the sequence technique, cross-correlation functions, and coherent averaging of SBP surges. During NREMS, all mice had lower SBP variability, greater baroreflex contribution to HP control at low frequencies, and greater amplitude of the central autonomic and baroreflex changes in HP associated with SBP surges than during wakefulness. During REMS, all mice had higher SBP variability and depressed central autonomic and baroreflex HP controls relative to NREMS. HP variability during REMS was higher than during NREMS in WT only. TG and KO also had lower amplitude of the cardiac baroreflex response to SBP surges during REMS than WT. These results indicate that chronic lack of HCRT signaling may cause subtle alterations in the control of HP during spontaneous behavior. Conversely, the integrity of HCRT signaling is not necessary for the occurrence of physiological sleep-dependent changes in SBP variability.     

State-dependent effects of light-dark cycle on somatosensory and visual cortex EEG in rats

Somatosensory (SSctx) and visual cortex (Vctx) EEG were evaluated in rats under a 12:12-h light-dark (LD) cycle and under constant light (LL) or constant dark (DD) in each sleep or wake state. Under LD conditions during light period, relative Vctx EEG slow-wave activity (SWA) was higher than that of the SSctx, whereas during dark period, relative Vctx EEG SWA was lower than in the SSctx. These effects were state specific, occurring only during non-rapid eye movement sleep (NREMS). Under LL conditions, the duration of REMS and NREMS during the period that would have been dark if the LD cycle had continued (subjective dark period) was greater than under LD conditions. DD conditions had little effect on the duration of NREMS and REMS. SSctx and Vctx EEG SWA were suppressed by LL during the subjective dark period; however, the degree of Vctx SWA suppression was smaller than that of the SSctx. DD conditions during the subjective light period enhanced SSctx SWA, whereas Vctx SWA was suppressed. Under LL conditions during the subjective dark period, Vctx EEG power was higher than that of the SSctx across a broad frequency range during NREMS, REMS, and wakefulness. During DD, SSctx EEG power during NREMS was higher than that of the Vctx in the delta wave band, whereas SSctx power during REMS and wakefulness was higher than that of the Vctx in frequencies higher than 8 Hz. We concluded that the SSctx and Vctx EEGs are differentially affected by light during subsequent sleep. Results provide support for the notion that regional sleep intensity is dependent on prior regional afferent input.     

Systemic injection of a nitric oxide synthase inhibitor suppresses sleep responses to sleep deprivation in rats

We hypothesized that nitric oxide (NO) may play a role in homeostatic sleep regulation. To test this hypothesis, we studied the sleep deprivation (SD)-induced homeostatic sleep responses after intraperitoneal administration of an NO synthase inhibitor, Nomega-nitro-L-arginine methyl ester (L-NAME, a cumulative dose of 100 mg/kg). Amounts and intensity of sleep were increased in response to 8 h of SD in control rats (n = 8). Sleep amounts remained above baseline for 16 h after SD followed by a negative rebound. Rapid eye movement sleep (REMS) and non-REMS (NREMS) intensities were elevated for 16 and 4 h, respectively. L-NAME treatment (n = 8) suppressed the rebound increases in NREMS amount and intensity. REMS rebound was attenuated by L-NAME in the first dark period after SD; however, a second rebound appeared in the subsequent dark period. REMS intensity did not increase after SD in L-NAME-injected rats. The finding that the NO synthase inhibitor suppressed rebound increases in NREMS suggests that NO may play a role as a signaling molecule in homeostatic regulation of NREMS.     

Essential Roles of GABA Transporter-1 in Controlling Rapid Eye Movement Sleep and in Increased Slow Wave Activity after Sleep Deprivation

GABA is the major inhibitory neurotransmitter in the mammalian central nervous system that has been strongly implicated in the regulation of sleep. GABA transporter subtype 1 (GAT1) constructs high affinity reuptake sites for GABA and regulates GABAergic transmission in the brain. However, the role of GAT1 in sleep-wake regulation remains elusive. In the current study, we characterized the spontaneous sleep-wake cycle and responses to sleep deprivation in GAT1 knock-out (KO) mice. GAT1 KO mice exhibited dominant theta-activity and a remarkable reduction of EEG power in low frequencies across all vigilance stages. Under baseline conditions, spontaneous rapid eye movement (REM) sleep of KO mice was elevated both during the light and dark periods, and non-REM (NREM) sleep was reduced during the light period only. KO mice also showed more state transitions from NREM to REM sleep and from REM sleep to wakefulness, as well as more number of REM and NREM sleep bouts than WT mice. During the dark period, KO mice exhibited more REM sleep bouts only. Six hours of sleep deprivation induced rebound increases in NREM and REM sleep in both genotypes. However, slow wave activity, the intensity component of NREM sleep was briefly elevated in WT mice but remained completely unchanged in KO mice, compared with their respective baselines. These results indicate that GAT1 plays a critical role in the regulation of REM sleep and homeostasis of NREM sleep.     

MicroRNA 132 alters sleep and varies with time in brain

MicroRNA (miRNA) levels in brain are altered by sleep deprivation; however, the direct effects of any miRNA on sleep have not heretofore been described. We report herein that intracerebroventricular application of a miRNA-132 mimetic (preMIR-132) decreased duration of non-rapid-eye-movement sleep (NREMS) while simultaneously increasing duration of rapid eye movement sleep (REMS) during the light phase. Further, preMIR-132 decreased electroencephalographic (EEG) slow-wave activity (SWA) during NREMS, an index of sleep intensity. In separate experiments unilateral supracortical application of preMIR-132 ipsilaterally decreased EEG SWA during NREMS but did not alter global sleep duration. In addition, after ventricular or supracortical injections of preMIR-132, the mimetic-induced effects were state specific, occurring only during NREMS. After local supracortical injections of the mimetic, cortical miRNA-132 levels were higher at the time sleep-related EEG effects were manifest. We also report that spontaneous cortical levels of miRNA-132 were lower at the end of the sleep-dominant light period compared with at the end of the dark period in rats. Results suggest that miRNAs play a regulatory role in sleep and provide a new tool for investigating sleep regulation.     

Loss of circadian organization of sleep and wakefulness during hibernation

We investigated circadian and homeostatic regulation of nonrapid eye movement (NREM) sleep in golden-mantled ground squirrels during euthermic intervals between torpor bouts. Slow-wave activity (SWA; 1-4 Hz) and sigma activity (10-15 Hz) represent the two dominant electroencephalographic (EEG) frequency components of NREM sleep. EEG sigma activity has a strong circadian component in addition to a sleep homeostatic component, whereas SWA mainly reflects sleep homeostasis [Dijk DJ and Czeisler CA. J Neurosci 15: 3526-3538, 1995; Dijk DJ, Shanahan TL, Duffy JF, Ronda JM, and Czeisler CA. J Physiol (Lond) 505: 851-858, 1997]. Animals maintained under constant conditions continued to display circadian rhythms in both sigma activity and brain temperature throughout euthermic intervals, whereas sleep and wakefulness showed no circadian organization. Instead, sleep and wakefulness were distributed according to a 6-h ultradian rhythm. SWA, NREM sleep bout length, and sigma activity responded homeostatically to the ultradian sleep-wake pattern. We suggest that the loss of sleep-wake consolidation in ground squirrels during the hibernation season may be related to the greatly decreased locomotor activity during the hibernation season and may be necessary for maintenance of multiday torpor bouts characteristic of hibernating species.     

Growth hormone-releasing hormone and corticotropin-releasing hormone enhance non-rapid-eye-movement sleep after sleep deprivation

The neuropeptides growth hormone (GH)-releasing hormone (GHRH) and corticotropin-releasing hormone (CRH) regulate sleep and nocturnal hormone secretion in a reciprocal fashion, at least in males. GHRH promotes sleep and GH and inhibits hypothalamo-pituitary-adrenocortical (HPA) hormones. CRH exerts opposite effects. In women, a sexual dimorphism was found because GHRH impairs sleep and stimulates HPA hormones. Sleep deprivation (SD) is the most powerful stimulus for inducing sleep. Studies in rodents show a key role of GHRH in sleep promotion after SD. The effects of GHRH and CRH on sleep-endocrine activity during the recovery night after SD are unknown. We compared sleep EEG, GH, and cortisol secretion between nights before and after 40 h of SD in 48 normal women and men aged 19-67 yr. During the recovery night, GHRH, CRH, or placebo were injected repetitively. After placebo during the recovery night, non-rapid-eye-movement sleep (NREMS) and rapid-eye-movement sleep (REMS) increased and wakefulness decreased compared with the baseline night. After GHRH, the increase of NREMS and the decrease of wakefulness were more distinct than after placebo. Also, after CRH, NREMS increased higher than after placebo, and a positive correlation was found between age and the baseline-related increase of slow-wave sleep. REMS increased after placebo and after GHRH, but not after CRH. EEG spectral analysis showed increases in the lower frequencies and decreases in the higher frequencies during NREMS after each of the treatments. Cortisol and GH did not differ between baseline and recovery nights after placebo. After GHRH, GH increased and cortisol decreased. Cortisol increased after CRH. No sex differences were found in these changes. Our data suggest that GHRH and CRH augment NREMS promotion after SD. Marked differences appear to exist in peptidergic sleep regulation between spontaneous and recovery sleep.     

Day- and nighttime injection of a nitric oxide synthase inhibitor elicits opposite sleep responses in rats

Previous studies suggest that nitric oxide (NO) may play a role in sleep regulation, particularly in the homeostatic process. The present studies were undertaken to compare the sleep effects of injecting a NO synthase (NOS) inhibitor when homeostatic sleep pressure is naturally highest (light onset) or when it is at its nadir (dark onset) in rats. Sleep, electroencephalogram delta-wave activity during nonrapid eye movement sleep (NREMS), also known as slow-wave activity (SWA), and brain temperature responses to three doses of the NOS inhibitor N(omega)-nitro-L-arginine methyl ester (L-NAME ; 5, 50, and 100 mg/kg) injected intraperitoneally at light or dark onset were examined in rats (n = 6 to 8). The effects of 5 mg/kg L-NAME were determined in both normal and vagotomized (VX) rats. Light onset administration of 50 mg/kg L-NAME decreased NREMS amounts and suppressed SWA and increased rapid eye movement sleep (REMS) amounts. At dark onset, L-NAME injection also dose dependently suppressed SWA; however, unlike light onset injections, both NREMS and REMS amounts were increased after all three doses. Sleep responses to 5 mg/kg L-NAME were not different in control and VX rats, suggesting that the sleep effects of L-NAME are not mediated through the activation of sensory vagal mechanisms. The present findings suggest that timing of the injection is a major determinant of the sleep responses observed after systemic L-NAME injection in rats.     

Elevated body temperature during sleep in orexin knockout mice

Core body temperature (Tb) is influenced by many physiological factors, including behavioral state, locomotor activity, and biological rhythms. To determine the relative roles of these factors, we examined Tb in orexin knockout (KO) mice, which have a narcolepsy-like phenotype with severe sleep-wake fragmentation. Because orexin is released during wakefulness and is thought to promote heat production, we hypothesized that orexin KO mice would have lower Tb while awake. Surprisingly, Tb was the same in orexin KO mice and wild-type (WT) littermates during sustained wakefulness. Orexin KO mice had normal diurnal variations in Tb, but the ultradian rhythms of Tb, locomotor activity, and wakefulness were markedly reduced. During the first 15 min of spontaneous sleep, the Tb of WT mice decreased by 1.0 degrees C, but Tb in orexin KO mice decreased only 0.4 degrees C. Even during intense recovery sleep after 8 h of sleep deprivation, the Tb of orexin KO mice remained 0.7 degrees C higher than in WT mice. This blunted fall in Tb during sleep may be due to inadequate activation of heat loss mechanisms or sustained activity in heat-generating systems. These observations reveal an unexpected role for orexin in thermoregulation. In addition, because heat loss is an essential aspect of sleep, the blunted fall in Tb of orexin KO mice may provide an explanation for the fragmented sleep of narcolepsy.     

Sleep in mice with nonfunctional growth hormone-releasing hormone receptors

The role of the somatotropic axis in sleep regulation was studied by using the lit/lit mouse with nonfunctional growth hormone (GH)-releasing hormone (GHRH) receptors (GHRH-Rs) and control heterozygous C57BL/6J mice, which have a normal phenotype. During the light period, the lit/lit mice displayed significantly less spontaneous rapid eye movement sleep (REMS) and non-REMS (NREMS) than the controls. Intraperitoneal injection of GHRH (50 microg/kg) failed to promote sleep in the lit/lit mice, whereas it enhanced NREMS in the heterozygous mice. Subcutaneous infusion of GH replacement stimulated weight gain, increased the concentration of plasma insulin-like growth factor-1 (IGF-1), and normalized REMS, but failed to restore normal NREMS in the lit/lit mice. The NREMS response to a 4-h sleep deprivation was attenuated in the lit/lit mice. In control mice, intraperitoneal injection of ghrelin (400 microg/kg) elicited GH secretion and promoted NREMS, and intraperitoneal administration of the somatostatin analog octretotide (Oct, 200 microg/kg) inhibited sleep. In contrast, these responses were missing in the lit/lit mice. The results suggest that GH promotes REMS whereas GHRH stimulates NREMS via central GHRH-Rs and that GHRH is involved in the mediation of the sleep effects of ghrelin and somatostatin.     

Central but not systemic administration of ghrelin induces wakefulness in mice

Ghrelin is a brain-gut peptide hormone widely known for its orexigenic and growth hormone-releasing activities. Findings from our and other laboratories indicate a role of ghrelin in sleep regulation. The effects of exogenous ghrelin on sleep-wake activity in mice are, however, unknown. The aim of the present study was to determine the sleep-modulating effects of ghrelin after central and systemic administrations in mice. Sleep-wake activity after intracerebroventricular (i.c.v.) administration of 0.2, 1 and 5 μg ghrelin and intraperitoneal injections of 40, 100, and 400 μg/kg ghrelin prior to light onset were determined in C57BL/6 mice. In addition, body temperature, motor activity and 1-hour food intake was measured after the systemic injections. Sleep effects of systemic ghrelin (40 and 400 μg/kg) injected before dark onset were also determined. I.c.v. injection of ghrelin increased wakefulness and suppressed non-rapid-eye-movement sleep and electroencephalographic slow-wave activity in the first hour after injections. Rapid-eye-movement sleep was decreased for 2-4 hours after each dose of ghrelin. Sytemic administration of ghrelin did not induce changes in sleep-wake activity in mice at dark or light onset. Motor activity and body temperature remained unaltered and food intake was significantly increased after systemic injections of ghrelin given prior the light period. These findings indicate that the activation of central, but not peripheral, ghrelin-sensitive mechanisms elicits arousal in mice. The results are consistent with the hypothesis that the activation of the hypothalamic neuronal circuit formed by ghrelin, orexin, and neuropeptide Y neurons triggers behavioral sequence characterized by increased wakefulness, motor activity and feeding in nocturnal rodents.     

Time-of-day modulation of homeostatic and allostatic sleep responses to chronic sleep restriction in rats

To study sleep responses to chronic sleep restriction (CSR) and time-of-day influences on these responses, we developed a rat model of CSR that takes into account the polyphasic sleep patterns in rats. Adult male rats underwent cycles of 3 h of sleep deprivation (SD) and 1 h of sleep opportunity (SO) continuously for 4 days, beginning at the onset of the 12-h light phase ("3/1" protocol). Electroencephalogram (EEG) and electromyogram (EMG) recordings were made before, during, and after CSR. During CSR, total sleep time was reduced by ～60% from baseline levels. Both rapid eye movement sleep (REMS) and non-rapid eye movement sleep (NREMS) during SO periods increased initially relative to baseline and remained elevated for the rest of the CSR period. In contrast, NREMS EEG delta power (a measure of sleep intensity) increased initially, but then declined gradually, in parallel with increases in high-frequency power in the NREMS EEG. The amplitude of daily rhythms in NREMS and REMS amounts was maintained during SO periods, whereas that of NREMS delta power was reduced. Compensatory responses during the 2-day post-CSR recovery period were either modest or negative and gated by time of day. NREMS, REMS, and EEG delta power lost during CSR were not recovered by the end of the second recovery day. Thus the "3/1" CSR protocol triggered both homeostatic responses (increased sleep amounts and intensity during SOs) and allostatic responses (gradual decline in sleep intensity during SOs and muted or negative post-CSR sleep recovery), and both responses were modulated by time of day.     

Age-related changes in the circadian and homeostatic regulation of human sleep

The reduction of electroencephalographic (EEG) slow-wave activity (SWA) (EEG power density between 0.75-4.5 Hz) and spindle frequency activity, together with an increase in involuntary awakenings during sleep, represent the hallmarks of human sleep alterations with age. It has been assumed that this decrease in non-rapid eye movement (NREM) sleep consolidation reflects an age-related attenuation of the sleep homeostatic drive. To test this hypothesis, we measured sleep EEG characteristics (i.e., SWA, sleep spindles) in healthy older volunteers in response to high (sleep deprivation protocol) and low sleep pressure (nap protocol) conditions. Despite the fact that the older volunteers had impaired sleep consolidation and reduced SWA levels, their relative SWA response to both high and low sleep pressure conditions was similar to that of younger persons. Only in frontal brain regions did we find an age-related diminished SWA response to high sleep pressure. On the other hand, we have clear evidence that the circadian regulation of sleep during the 40 h nap protocol was changed such that the circadian arousal signal in the evening was weaker in the older study participants. More sleep occurred during the wake maintenance zone, and subjective sleepiness ratings in the late afternoon and evening were higher than in younger participants. In addition, we found a diminished melatonin secretion and a reduced circadian modulation of REM sleep and spindle frequency-the latter was phase-advanced relative to the circadian melatonin profile. Therefore, we favor the hypothesis that age-related changes in sleep are due to weaker circadian regulation of sleep and wakefulness. Our data suggest that manipulations of the circadian timing system, rather than the sleep homeostat, may offer a potential strategy to alleviate age-related decrements in sleep and daytime alertness levels.     

Neurobiology of the sleep-wake cycle: sleep architecture, circadian regulation, and regulatory feedback

This mini-review article presents the remarkable progress that has been made in the past decade in our understanding of the neural circuitry underlying the regulation of sleep-wake states and circadian control of behaviors. Following a brief introduction to sleep architecture and physiology, the authors describe the neural circuitry and neurotransmitters that regulate sleep and cortical arousal (i.e., wakefulness). They next examine how sleep and wakefulness are regulated by mutual inhibition between sleep-and arousal-promoting circuitry and how this interaction functions analogously to an electronic "flip-flop" switch that ensures behavioral state stability. The authors then discuss the role of circadian and homeostatic processes in the consolidation of sleep, including the physiologic basis of homeostatic sleep drive (i.e., wake-dependent increase in sleep propensity) and the role of the SCN in the circadian regulation of sleep-wake cycles. Finally, they describe the hypothalamic circuitry for the integration of photic and nonphotic environmental time cues and how this integration allows organisms to sculpt patterns of rest-activity and sleep-wake cycles that are optimally adaptive.     

L-364,718, a cholecystokinin-A receptor antagonist, suppresses feeding-induced sleep in rats

Feeding induces increased sleep in several species, including rats. The aim of the study was to determine if CCK plays a role in sleep responses to feeding. We induced excess eating in rats by 4 days of starvation and studied the sleep responses to refeeding in control and CCK-A receptor antagonist-treated animals. Sleep was recorded on 2 baseline days when food was provided ad libitum. After the starvation period, sleep was recorded on 2 refeeding days when the control rats (n = 8) were injected with vehicle and the experimental animals (n = 8) received intraperitoneal injections of L-364,718 (500 microg/kg, on both refeeding days). In the control group, refeeding caused increases in rapid eye movement sleep (REMS) and non-REMS (NREMS) and decreases in NREMS intensity as indicated by the slow-wave activity (SWA) of the electroencephalogram. CCK-A receptor antagonist treatment completely prevented the SWA responses and delayed the NREMS responses to refeeding; REMS responses were not simply abolished, but the amount of REMS was below baseline after the antagonist treatment. These results suggest that endogenous CCK, acting on CCK-A receptors, may play a key role in eliciting postprandial sleep.     

Startle-evoked changes in diaphragmatic activity during wakefulness and sleep

Tonic inhibition of some respiratory muscles occurs as part of the generalized muscle atonia of rapid-eye-movement sleep (REMS). A second type of inhibition of the diaphragm during REMS, fractionations, consists of brief pauses in the diaphragmatic electromyogram (DIA EMG) in association with phasic events. Because motor inhibition can occur as part of the startle response, and the brain is highly activated during REMS, we hypothesized that the neural basis of the fractionations might be activation of a startle network. To test this hypothesis, tone bursts (100 dB, 20-ms duration at 15-s intervals) were applied to cats at a fixed inspiratory level in the DIA moving average during REMS, non-rapid-eye-movement sleep (NREMS), and wakefulness. Parallel sham studies (no tone applied) were obtained for each state. The response of the DIA EMG was averaged over 100 ms by using the tone pulse as a trigger, and the following parameters of the DIA EMG were measured: latency to peak and/or nadir, increment or decrement in activity, and duration of peak and/or nadir. After a tone, all five animals studied displayed a profound suppression of DIA activity during REMS (latency to nadir 42.4 +/- 10.0 ms, duration of suppression 35.9 +/- 17.6 ms). Similarly, DIA activity was suppressed in all cats during NREMS (latency to nadir 40.9 +/- 13.3 ms, duration 23.9 +/- 13.4 ms). An excitatory response was observed in only two cats during NREMS and wakefulness. The similarity of startle-induced DIA EMG pauses to spontaneous fractionations of DIA activity during REMS suggests that the latter result from activation of a central startle system.     

Sleep,activity, temperature and arousal responses of mice deficient for muscarinic receptor M2 or M4

AimsThe type 2 muscarinic receptor (M2R) differs from the other G-protein-coupled muscarinic receptor (type 4, or M4R) in tissue distribution and physiologic effects. We studied the impact of these receptors on sleep and arousal by using M2R and M4R knock-out (KO) mice.Main methodsM2R and M4R KO and genetically intact mice were compared in terms of normal patterns of sleep, responses to sleep loss, infectious challenge and acoustic startle, and acoustic prepulse inhibition of startle (PPI).Key findingsUnder basal conditions, M2R and M4R KO mice do not differ from the background strain or each other in the amount or diurnal pattern of sleep, locomotor activity, and body temperature. After enforced sleep loss, M2R KO mice, in contrast to the other two strains, show no rebound in slow-wave sleep (SWS) time, although their SWS is consolidated, and they show a greater rebound in time spent in REMS (rapid-eye-movement sleep) and REMS consolidation. During influenza infection, M2R KO mice, as compared with the other strains, show marked hypothermia and a less robust increase in SWS. During Candida albicans infection, M2R KO mice show a greater increase in SWS and a greater inflammatory response than do the other strains. M2R KO mice also show greater acoustic startle amplitude than does the background strain, although PPI was not different across the 3 strains over a range of stimulus intensities.SignificanceTaken together, these findings support different roles for M2R and M4R in the modulation of sleep and arousal during homeostatic challenge.     

miR-155 deletion modulates lipopolysaccharide-induced sleep in female mice

Immune signaling is known to regulate sleep. miR-155 is a microRNA that regulates immune responses. We hypothesized that miR-155 would alter sleep regulation. Thus, we investigated the potential effects of miR-155 deletion on sleep-wake behavior in adult female homozygous miR-155 knockout (miR-155^KO) mice and littermate controls (WT). Mice were implanted with biotelemetry units and EEG/EMG biopotentials were recorded continuously for three baseline days. miR-155^KO mice had decreased bouts of NREM and REM sleep compared with WT mice, but no differences were observed in the length of sleep bouts or total time spent in sleep-wake states. Locomotor activity and subcutaneous temperature did not differ between WT and miR-155^KO mice. Following baseline recordings, mice were sleep-deprived during the first six hours of the rest phase (light phase; ZT 0–6) followed by an 18 h recovery period. There were no differences between groups in sleep rebound (% sleep and NREM δ power) after sleep deprivation. Following recovery from sleep deprivation, mice were challenged with a somnogen (viz., lipopolysaccharide (LPS)) one hour prior to the initiation of the dark (active) phase. Biopotentials were continuously recorded for the following 24 h, and miR-155^KO mice displayed increased wakefulness and decreased NREM sleep during the dark phase following LPS injection. Additionally, miR-155^KO mice had reduced EEG slow-wave responses (0.5–4 Hz) compared to WT mice. Together, our findings indicate that miR-155 deletion attenuates the somnogenic and EEG delta-enhancing effects of LPS.

Abbreviations: ANOVA: analysis of variance; EEG: electroencephalogram; EMG: electromyogram; h: hour; IL-1: interleukin-1; IL-6: interleukin-6; IP: intra-peritoneal; LPS: lipopolysaccharide; miR/miRNA: microRNA; miR-155^KO: miR-155 knockout; NREM: non-rapid eye movement; REM: rapid eye movement; TNF: tumor necrosis factor; SWS: slow-wave sleep; WT: wild-type.     

Ghrelin microinjection into forebrain sites induces wakefulness and feeding in rats

Ghrelin, a gut-brain peptide, is best known for its role in the stimulation of feeding and growth hormone release. In the brain, orexin, neuropeptide Y (NPY), and ghrelin are parts of a food intake regulatory circuit. Orexin and NPY are also implicated in maintaining wakefulness. Previous experiments in our laboratory revealed that intracerebroventricular injections of ghrelin induce wakefulness in rats. To further elucidate the possible role of ghrelin in the regulation of arousal, we studied the effects of microinjections of ghrelin into hypothalamic sites, which are implicated in the regulation of feeding and sleep, such as the lateral hypothalamus (LH), medial preoptic area (MPA), and paraventricular nucleus (PVN) on sleep in rats. Sleep responses, motor activity, and food intake after central administration of 0.04, 0.2, or 1 mug (12, 60, or 300 pmol) ghrelin were recorded. Microinjections of ghrelin into the LH had strong wakefulness-promoting effects lasting for 2 h. Wakefulness was also stimulated by ghrelin injection into the MPA and PVN; the effects were confined to the first hour after the injection. Ghrelin's non-rapid-eye-movement sleep-suppressive effect was accompanied by attenuation in the electroencephalographic (EEG) slow-wave activity and changes in the EEG power spectrum. Food consumption was significantly stimulated after microinjections of ghrelin into each hypothalamic site. Together, these results are consistent with the hypothesis that forebrain ghrelinergic mechanisms play a role in the regulation of vigilance, possibly through activating the components of the food intake- and arousal-promoting network formed by orexin and NPY.