Wakefulness In Rats Research Articles

Spontaneous sleep and homeostatic sleep regulation in ghrelin knockout mice

Ghrelin is well known for its feeding and growth hormone-releasing actions. It may also be involved in sleep regulation; intracerebroventricular administration and hypothalamic microinjections of ghrelin stimulate wakefulness in rats. Hypothalamic ghrelin, together with neuropeptide Y and orexin form a food intake-regulatory circuit. We hypothesized that this circuit also promotes arousal. To further investigate the role of ghrelin in the regulation of sleep-wakefulness, we characterized spontaneous and homeostatic sleep regulation in ghrelin knockout (KO) and wild-type (WT) mice. Both groups of mice exhibited similar diurnal rhythms with more sleep and less wakefulness during the light period. In ghrelin KO mice, spontaneous wakefulness and rapid-eye-movement sleep (REMS) were slightly elevated, and non-rapid-eye-movement sleep (NREMS) was reduced. KO mice had more fragmented NREMS than WT mice, as indicated by the shorter and greater number of NREMS episodes. Six hours of sleep deprivation induced rebound increases in NREMS and REMS and biphasic changes in electroencephalographic slow-wave activity (EEG SWA) in both genotypes. Ghrelin KO mice recovered from NREMS and REMS loss faster, and the delayed reduction in EEG SWA, occurring after sleep loss-enhanced increases in EEG SWA, was shorter-lasting compared with WT mice. These findings suggest that the basic sleep-wake regulatory mechanisms in ghrelin KO mice are not impaired and they are able to mount adequate rebound sleep in response to a homeostatic challenge. It is possible that redundancy in the arousal systems of the brain or activation of compensatory mechanisms during development allow for normal sleep-wake regulation in ghrelin KO mice.

Modafinil enhances extracellular levels of dopamine in the nucleus accumbens and increases wakefulness in rats

Modafinil (MOD) is a wakefulness-promoting drug that improves the alertness levels in narcolepsy; however, the molecular mechanism of action remains to be elucidated. We found that after a single icv injection of MOD (10 μg/5 μl) the extracellular levels of dopamine (DA) and l-DOPA collected from the nucleus accumbens were increased and decreased, respectively. Separately, the icv administration of MOD (10 μg/5 μl) to rats enhanced wakefulness (W) whereas diminished sleep during 4 h. Lastly, the alertness induced by MOD was partially antagonized by the sleep-inducing endocannabinoid anandamide (ANA). We conclude that MOD enhances the extracellular levels of DA, promotes W and its effects on sleep are partially blocked by ANA.

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.

Obestatin alters sleep in rats

Obestatin is a recently identified peptide derived from the ghrelin gene. Ghrelin stimulates food intake whereas obestatin has an opposite effect in rats. Previous experiments in our laboratory revealed that ghrelin also induces wakefulness in rats. The aim of the present experiments was to study the effect of obestatin on sleep. Rats received intraperitoneal ( n = 7; 16 or 64 μg/kg) or intracerebroventricular (i.c.v.; n = 11) injection of pyrogen-free isotonic NaCl or obestatin (1, 4 and 16 μg in a volume of 4 μl) at dark onset. Sleep-wake activity was recorded for 23 h. I.c.v. administration of 16 μg obestatin induced a significant increase (∼58%) in the time spent in non-rapid-eye-movement sleep (NREMS) in the first hour after the injection. This resulted from an increase in the number of NREMS episodes and shortened sleep latency. Electroencephalographic (EEG) slow-wave activity (0.5–4 Hz) was reduced by obestatin treatment. The initial increase in NREMS time was followed by a decrease in both NREMS and REMS in the second hour after the injection. Peripheral injection of obestatin did not induce significant changes in sleep in any post-injection hours. Results suggest that the sleep-promoting effect of centrally administered obestatin may be part of the behavioral manifestation of satiety elicited by the peptide. Current results confirm the finding that two regulatory peptides derived from the same gene have opposite actions in the same species.

Central administration of neuropeptide Y induces wakefulness in rats

Neuropeptide Y (NPY) is a well-characterized neuromodulator in the central nervous system, primarily implicated in the regulation of feeding. NPY, orexins, and ghrelin form a hypothalamic food intake regulatory circuit. Orexin and ghrelin are also implicated in sleep-wake regulation. In the present experiments, we studied the sleep-modulating effects of central administration of NPY in rats. Rats received intracerebroventricular injection of physiological saline or three different doses of NPY (0.4, 2, and 10 microg in a volume of 4 microl) at light onset. Another group of rats received bilateral microinjection of saline or 2 microg NPY in the lateral hypothalamus in a volume of 0.2 microl. Sleep-wake activity and motor activity were recorded for 23 h. Food intake after the control and treatment injections was also measured on separate days. Intracerebroventricular and lateral hypothalamic administration of NPY suppressed non-rapid-eye-movement sleep and rapid-eye-movement sleep in rats during the first hour after the injection and also induced changes in electroencephalogram delta power spectra. NPY stimulated food intake in the first hour after both routes of administration. Data are consistent with the hypothesis that NPY has a role in the integration of feeding, metabolism, and sleep regulation.

Participation of histaminergic H 1 and noradrenergic α 1 receptors in orexin A-induced wakefulness in rats

The participation of histaminergic H 1 and noradrenergic α 1 receptors in orexin A-induced wakefulness was studied by examining the sleep–wakefulness cycle in rats. Intracerebroventricular infusion of orexin A (1 nmol) caused an increase in the wakefulness state, while non-rapid eye movement sleep (NREM sleep) and rapid eye movement sleep (REM sleep) states were decreased. Prazosin (150 nmol) showed no significant antagonistic effect on the orexin A-induced increase in the wakefulness state and decrease in NREM and REM sleep. On the contrary, pyrilamine (150 nmol) was effective in antagonizing orexin A-induced increase in wakefulness and decrease in NREM sleep. When prazosin (150 nmol) and pyrilamine (150 nmol) were simultaneously perfused into the lateral ventricle, an almost complete antagonistic effect was observed with the increase in the wakefulness state and decrease in NREM sleep. Orexin A (1 nmol) caused a significant decrease in the histamine contents of the cortex, hippocampus and hypothalamus, whereas noradrenaline contents were decreased only in the hypothalamus. From these results, we concluded that the arousal effect induced by orexin A occurs through histaminergic H 1 and noradrenergic α 1 receptors, although participation of the H 1 receptor was more important than the α 1 receptor.

ROLE OF HYPOCRETIN-1,2 (OREXIN A AND B) IN PAIN PERCEPTION

Hypocretins/orexins are primary excitatory neuropeptides located exclusively in neurons of the lateral hypothalamic area, which send projections to most monoaminergic nuclei. It has been reported that i.c.v. injection of hypocretin 1 (orexin A) enhances wakefulness in rats and mice. The present work was carried out to examine the roles of hypocretins in nociception in mice. The presence of robust projections from the hypothalamus to laminae I and II of the spinal cord strongly suggests the role of hypocretins in the nociceptive pathways. Localization of the fibers of hypocretin-containing neurons in the hypothalamus, locus coeruleus, thalamus and periaqueductal gray is also consistent with their central roles in sensory processing. To investigate the role of these hypothalamic peptides, C57BL/6 mice were administered with hypocretin 1 and 2 (i.c.v.) and intrathecally (I.T.), and examined for pain thresholds using three kinds of nociceptive tasks. These included assays for thermal (hot-plate, tail-flick, paw-withdrawal), mechanical (tail-pressure), chemical (formalin, capsaicin and abdominal stretch) nociceptions and behavioral responses. The I.C.V. and I.T. administration of hypocretins produced morphine-like anti-nociceptive effects in mice. The anti-nociceptive effects of hypocretins 1 (orexin A) were more remarkable than those of hypocretins 2 (orexin B). The effects of hypocretin 1 were completely blocked by 3-dipropyl-8-cyclopentylxanthine (DPCPX), a nonselective adenosine receptor antagonist but not by naloxone. No motor impairments were observed with both of the compounds at the doses studied. The present findings suggested that the hypocretin-containing neurons in the hypothalamus have a potential role in the modulation of nociceptive transmission.

Prostaglandin E2Activates the Histaminergic System via the EP4Receptor to Induce Wakefulness in Rats

Prostaglandin (PG)E2 promotes the wakeful state when administered into the posterior hypothalamus, in which the histaminergic tuberomammillary nucleus (TMN) is located. To explore the neurotransmitter mechanisms responsible for PGE2-induced wakefulness in rats, we examined the effect of PGE2 on the activity of the histaminergic system and the involvement of PGE2 receptor subtypes in the response. PGE2 perfusion in the TMN at doses of 100, 200, and 400 pmol/min for 2 hr significantly increased histamine release from the medial preoptic area and frontal cortex in a dose-dependent manner, as measured by in vivo microdialysis. Among the agonists of the four distinct subtypes of PGE2 receptors (EP1-4) tested, only the EP4 receptor agonist (ONO-AE1-329) mimicked the excitatory effect of PGE2 on histamine release from both the medial preoptic area and frontal cortex. Perfusion of either PGE2 or the EP4 agonist into the TMN at a dose of 200 pmol/min for 1 hr increased histidine decarboxylase activity, histidine decarboxylase mRNA level, and histamine content in the hypothalamus. In situ hybridization revealed that EP4 receptor mRNA was expressed in histidine decarboxylase-immunoreactive neurons of the TMN region. Furthermore, EP4 agonist perfusion into the TMN induced wakefulness. These findings indicate that PGE2 induces wakefulness through activation of the histaminergic system via EP4 receptors.

Induction of Hippocampal Long-Term Potentiation during Waking Leads to Increased Extrahippocampalzif-268Expression during Ensuing Rapid-Eye-Movement Sleep

Rapid-eye-movement (REM) sleep plays a key role in the consolidation of memories acquired during waking (WK). The search for mechanisms underlying that role has revealed significant correlations in the patterns of neuronal firing, regional blood flow, and expression of the activity-dependent gene zif-268 between WK and subsequent REM sleep. Zif-268 integrates a major calcium signal transduction pathway and is implicated by several lines of evidence in activity-dependent synaptic plasticity. Here we report that the induction of hippocampal long-term potentiation (LTP) during WK in rats leads to an upregulation of zif-268 gene expression in extrahippocampal regions during subsequent REM sleep episodes. This upregulation occurs predominantly in the amygdala, entorhinal, and auditory cerebral cortices during the first REM sleep episodes after LTP induction and reaches somatosensory and motor cerebral cortices as REM sleep recurs. We also show that hippocampal inactivation during REM sleep blocks extrahippocampal zif-268 upregulation, indicating that cortical and amygdalar zif-268 expression during REM sleep is under hippocampal control. Thus, expression of an activity-dependent gene involved in synaptic plasticity propagates gradually from the hippocampus to extrahippocampal regions as REM sleep recurs. These findings suggest that a progressive disengagement of the hippocampus and engagement of the cerebral cortex and amygdala occurs during REM sleep. They are also consistent with the view that REM sleep constitutes a privileged window for hippocampus-driven cortical activation, which may play an instructive role in the communication of memory traces from the hippocampus to the cerebral cortex.

Sleep and the hypothalamo-pituitary-adrenocortical system.

The intention of this review is to summarize the current knowledge on the bidirectional interaction between sleep EEG and the secretion of corticotropin (ACTH) and cortisol. The administration of various hypothalamic-pituitary- adrenocortical (HPA) hormones and their antagonists exerts specific sleep-EEG changes in several species including humans. It is well documented that corticotropin releasing hormone (CRH) impairs sleep and enhances vigilance. In addition, it may promote REM sleep. Changes in the growth hormone-releasing hormone (GHRH):CRH ratio in favour of CRH appear to contribute to shallow sleep, elevated cortisol levels and blunted GH in depression and ageing. On the other hand, in women GHRH appears to exert CRH-like effects on sleep. Acute cortisol administration increases slow-wave sleep (SWS) and GH, probably due to feedback inhibition of CRH, and inhibits REM sleep. With the mixed glucocorticoid and progesterone receptor antagonist mifepriston sleep is disrupted. Subchronic administration of the glucocorticoid agonist methylprednisolone desinhibited REM sleep. A synergism of elevated CRH and cortisol activity may contribute to REM disinhibition during depression. Also ACTH and vasopressin modulate sleep specifically but their physiological role remains unclear. For example acute icv vasopressin enhances wakefulness in rats, whereas its long-term administration increases SWS in the elderly. In various studies the interaction of sleep EEG and HPA hormones has been investigated at the baseline, after manipulation of sleep-wake behaviour and after environmental changes. Most studies agree that the circadian pattern of cortisol is relatively independent from sleep and environmental influences. Some data suggest a major effect of light on cortisol secretion. Sleeping is widely associated with blunting and awakenings are linked with increases of HPA hormones.

Changes in sleep–wakefulness after 6-hydroxydopamine lesion of the preoptic area

This study was undertaken to assess the role of catecholamine fibers, terminating in the preoptic area, in regulating sleep–wakefulness in rats. Sleep–wakefulness was assessed on the basis of 24 h electroencephalogram, electromyogram and electro-oculogram recordings before and after destruction of catecholaminergic terminals at the medial preoptic area by bilateral intracerebral injection of 6-hydroxydopamine (8 μg in 0.2 μl). There was a mild reduction in sleep and increase in wakefulness after the lesion. The increase in active wakefulness observed after eight days of lesion persisted even on the 12th day. In spite of the reduction in sleep, the day–night sleep ratio was not affected by 6-hydroxydopamine lesion of the preoptic area. The results indicate that the noradrenergic fibers at the preoptic area have a hypnogenic role.

Rolipram, an Antidepressant That Increases the Availability of cAMP, Transiently Enhances Wakefulness in Rats

A study was carried out on the effects on sleep of rolipram, an antidepressant that increases the availability of cAMP by inhibiting a phosphodiesterase isoenzyme. Rats were treated with rolipram (0.1 or 1 mg/kg) twice a day (at light and dark onset) for 11 days, after a chronic period of injection of physiological saline for habituation purposes. The sleep–wake activity was recorded for 12 h following the injection at light onset on the baseline day (physiological saline), on rolipram days 1, 5, and 11, and also on day 12, when physiological saline was injected again (withdrawal day). The high (1 mg/kg) dose of rolipram enhanced wakefulness (W) in postinjection h 1 on day 1 of rolipram treatment. After administration of 0.1 mg/kg rolipram, only a tendency to an increase in W was noted. The promotion of W might be attributed, at least in part, to an increased release of noradrenaline due to a cAMP-mediated stimulation of tyrosine hydroxylase.

Sleep and waking during acute histamine hs agonist BP 2.94 or hs antagonist carboperamide (MR 16155) administration in rats

The present study evaluated the effects of histamine H3 receptor agonist BP 2.94 or H3 receptor antagonist carboperamide (MR 16155) given by oral route on sleep and waking in rats surgically prepared for long-term recordings. BP 2.94 produced a significant increase of slow-wave sleep (SWS) that was related to slight decreases of waking, light sleep, and REM sleep. Carboperamide significantly increased waking and decreased SWS and REM sleep. Pretreatment with carboperamide prevented the effect of BP 2.94 on SWS. It is suggested that the effects of BP 2.94 or carboperamide on sleep and waking could depend on changes in the availability of histamine at the postsynaptic H1 receptor. Alternatively, activation or blockade of the H3 heteroreceptors found in the central catecholamine, indolamine, and acetylcholine nerve endings could inhibit or increase the release of noradrenaline, serotonin, dopamine, and acetylcholine. This would secondarily result in changes of sleep variables.

O-13-1 - Sleep and waking during acute histamine H 3-agonist BP 2.94 or H 3-antagonist carboperamide (MR 16155) administration in rats

The present study evaluated the effects of histamine H3 receptor agonist BP 2.94 or H3 receptor antagonist carboperamide (MR 16155) given by oral route on sleep and waking in rats surgically prepared for long-term recordings. BP 2.94 produced a significant increase of slow-wave sleep (SWS) that was related to slight decreases of waking, light sleep, and REM sleep. Carboperamide significantly increased waking and decreased SWS and REM sleep. Pretreatment with carboperamide prevented the effect of BP 2.94 on SWS. It is suggested that the effects of BP 2.94 or carboperamide on sleep and waking could depend on changes in the availability of histamine at the postsynaptic H1 receptor. Alternatively, activation or blockade of the H3 heteroreceptors found in the central catecholamine, indolamine, and acetylcholine nerve endings could inhibit or increase the release of noradrenaline, serotonin, dopamine, and acetylcholine. This would secondarily result in changes of sleep variables.

Lesion of descending 5-HT pathways increases zimeldine-induced waking in rats

Sleep, waking, and EEG power spectra were investigated in rats with spinal 5,6-dihydroxytryptamine (5,6-DHT) lesions, following 20 mg/kg zimeldine or vehicle IP injections. 5,6-DHT selectively lesioned the descending serotonergic pathways. Lesion alone did not change sleep and waking stages compared to baseline, except for a reduction in REM sleep. Consistent with earlier findings, zimeldine in nonlesioned rats increased waking the first 2 h of recording. Zimeldine treatment in lesioned rats gave a significant additional 50% increase in waking the first 2 h and a corresponding decrease in total slow wave sleep, suggesting a potentiation of these effects. Zimeldine gave no significant changes in waking EEG power spectral density. Lesion gave a tendency to reduction between 4.0 and 15.5 Hz compared with baseline, and between 10.0 and 16.5 compared to the independent control group. In both comparisons, the combined treatment strengthened this effect, again suggesting a potentiating effect of lesion. In sleep, zimeldine reduced power over the whole spectrum (0.5–20.0 Hz), less in the lower frequencies than in the higher frequencies.

The 5-HT 1A antagonist (−)-alprenolol fails to modify sleep or zimeldine-induced sleep-waking effects in rats

Sleep and waking in rats were studied for 8 h following administration of a selective 5-hydroxytryptamine (5-HT) reuptake inhibitor (zimeldine), a putative 5-HT 1A antagonist {L(−)-alprenolol hydrogene tartrate monohydrate [(−)-alprenolol]} and a combination of (−)-alprenolol and zimeldine. Consistent with earlier findings, zimeldine gave a biphasic effect on sleep and waking. Waking was increased during the first 3 h, followed by a small decrease. Deep slow-wave sleep (SWS-2) showed the opposite trend. An initial decrease in SWS-2 was followed by an increase after around 3 h. Rapid eye movement sleep was markedly suppressed and latencies to sleep increased after zimeldine. (−)-Alprenolol had no effects on the different sleep and waking stages or latencies to sleep. The 5-HT 1A antagonist also failed to modify the effects of zimeldine administration. The behavioral syndrome induced by a selective 5-HT 1A agonist [8-hydroxy-2-(di- n-propyl-amino)-tetralin (8-OH-DPAT)] was clearly antagonized by administration of (−)-alprenolol, indicating that (−)-alprenolol was an efficient 5-HT 1A blocker. The data indicate that the sleep-waking effects of zimeldine cannot easily be explained by stimulation of 5-HT 1A receptors.

Effects of zimeldine, a selective 5-HT reuptake inhibitor, combined with ritanserin, a selective 5-HT 2 antagonist, on waking and sleep stages in rats

Sleep and waking in rats were studied 8 h following administration of a selective 5-hydroxytryptamine (5-HT) reuptake inhibitor (zimeldine), a selective 5-HT 2 antagonist (ritanserin) and a combination of ritanserin and zimeldine. Consistent with earlier findings, zimeldine gave a biphasic effect on sleep and waking. Waking was increased the first 3 h, followed by an increase in deep slow wave sleep (SWS-2), maximal in hours 4 and 5. Ritanserin gave an increase in SWS-2 that was spread out over the recording period. Ritanserin + zimeldine also gave a biphasic effect as zimeldine did, and the initial increase in waking and the following increase in SWS-2 tended to be stronger. Thus, ritanserin did not block the initial waking effect seen after zimeldine administration, indicating that this waking effect was not due to 5-HT 2 stimulation. The increase in SWS-2 seemed to reflect an addition of the increases following the zimeldine and ritanserin alone conditions. This suggests that the increase in SWS-2 seen after 5-HT reuptake inhibition and 5-HT 2 blockade are independent phenomena. Zimeldine alone, ritanserin alone and the combination all gave a clear reduction of rapid eye movement sleep.

Ozone-induced bradycardia and arrhythmia and their relation to sleep—wakefulness in rats

Rats were chronically implanted with electrodes for EEG, EMG and ECG recordings and exposed to 0.1 ppm, 0.2 ppm ozone (O 3) or clean air as control for 5 consecutive days. Amounts of wakefulness (W), slow-wave sleep (SWS) and paradoxical sleep (PS) of the O 3-exposed rats were not statistically different from respective control values. Compared with control rats, heart rates of the O 3-exposed rats decreased and the number of bradyarrhythmic episodes increased with an increase in O 3 levels. The O 3-induced bradyarrhythmias occurred more prevalently in the order of W, SWS and PS state. Parasympathetic mechanisms were suggested on those cardiac abnormalities.

Analysis of the structure of sleep and waking in rats with a parkinsonian syndrome induced by 1-methyl-4-phenyl-1,2,3,6,-tetrahydropyridine and by oxotremorine

Analysis of the structure of sleep and waking in rats with a parkinsonian syndrome induced by 1-methyl-4-phenyl-1,2,3,6,-tetrahydropyridine and by oxotremorine

Dose-response effects of 8-cyclopropyltheophylline on sleep and wakefulness in rats.

The dose-response effects of the substituted xanthine 8-cyclopropyltheophylline (CPRT) on sleep and wakefulness (W) after intraperitoneal administration to rats were examined by means of simultaneous electroencephalographic (EEG) and electromyographic (EMG) recordings. Doses of 20 and 40 mg/kg CPRT increased W and decreased slow wave sleep (SWS) in rats, indicating CNS stimulant effects. The greatest CNS stimulation was produced by the lowest (20 mg/kg) dose of CPRT examined, which also increased the latency to SWS. In addition, the 20 mg/kg dose of CPRT also significantly decreased the amount of total sleep (TS), as compared to the vehicle group, during all time periods examined. In contrast, the 80 mg/kg dose of CPRT decreased W and increased both SWS and TS. However, this apparent hypnotic effect of the 80 mg/kg CPRT may be due to toxicity, since 80% of rats treated with this dose of the drug died within 48 h of injection.