Agomelatine restores a physiological response to stress in the aged rat.

Short duration immobilization stress (IS) in younger rats is followed by a sleep rebound involving slow wave sleep (SWS) and, more particularly, rapid eye movement (REM) sleep. This rebound, expressing the ability of the brain to confront a stress challenge, is now accepted as a marker of the homeostasis. In older rats (24-25 months), however, an IS of 1h is not followed by a sleep rebound. To determine whether this impairment is reversible, we analyzed the effects of the antidepressant agomelatine, on stress-related sleep rebound in older animals. Older and younger (3-5 months) rats were equipped with electroencephalographic (EEG) and electromyographic (EMG) electrodes and polygraphic recordings were achieved under basal conditions with a digitized set-up. Older rats were pretreated with agomelatine (40mg/kg/day) for 3 days, with IS applied on the third day, whereas younger rats were only subjected to IS. Polygraphic recordings achieved under basal conditions confirmed the conventional impairments of the sleep/wake architecture in older animals, including decreased delta power, shortened REM sleep bouts, and modified sleep/wake circadian rhythms. Older rats pretreated with agomelatine for 3 days showed a reversal of the deficit observed in the beta-1, but not in the delta, EEG power band. Application of an IS to older rats after agomelatine pretreatment resulted in a REM sleep rebound in response to stress. These findings indicate that agomelatine, by improving beta-1 EEG power band and by inducing stress-related sleep rebound in older animals, contributes to the homeostasis maintenance.

Cortical and pontine variations occurring in the voltammetric no signal throughout the sleep-wake cycle in the rat.

Voltammetric measurements of nitric oxide (NO) were performed either in the frontal cortex (Cx) or in the nucleus raphe dorsalis (nRD) of rats equipped for polygraphic recordings. In the frontal cortex, the 650 mV signal related to NO exhibited its highest height during the waking state (W) and decreased slightly during slow-wave sleep (SWS) and even more during paradoxical sleep (PS). In the nRD, opposite variations were observed, i.e. the signal tended towards an increase during SWS and raised more consistently during PS versus W. Recordings performed either in the Cx or the nRD, throughout the light (12-h) and dark (12-h) periods, exhibited opposite nycthemeral changes, i.e. the signal height was higher in the Cx and lower in the nRD during the dark period and conversely for the light one. Paracrine and synaptic mechanisms taking place within the pons and, at least partly, also reflected in the Cx need to be further investigated.

Waking selective neurons in the posterior hypothalamus and their response to histamine H3-receptor ligands: an electrophysiological study in freely moving cats.

Neurons which discharge selectively during waking (waking selective) have been found in the tuberomamillary nucleus (TM) and adjacent areas of the posterior hypothalamus. Although they share some electrophysiological properties with aminergic neurons, there is no direct evidence that they are histaminergic. We have recorded from posterior hypothalamic neurons during the sleep-wake cycle in freely moving cats, and investigated the effects on waking selective neurons of specific ligands of histaminergic H3-receptors, which autoregulate the activity of histaminergic neurons. Two types of neurons were seen. Waking selective neurons, termed "waking-on (W-on)," were located exclusively within the TM and adjacent areas, and discharged at a low regular rate during waking (1.71-2.97 Hz), decreased firing during light slow wave sleep (SWS), became silent during deep SWS and paradoxical sleep (PS) and resumed their activity on, or a few seconds before, awakening. "Waking-related" neurons, located in an area dorsal to the TM, displayed a similar, although less regular, low rate of firing (1.74-5.41 Hz) and a similar discharge profile during the sleep-wake cycle; however, unlike "W-on" neurons, they did not completely stop firing during deep SWS and PS. Intramuscular (i.m.) injection of ciproxifan (an H3-receptor antagonist, 1mg/kg), significantly increased the discharge rate of W-on neurons and induced c-fos expression in histamine-immunoreactive neurons, whereas i.m. injection of imetit (an H3-receptor agonist, 1mg/kg) or microinjection of alpha-methylhistamine (another H3-receptor agonist, 0.025-0.1 microg/0.2 microl) in the vicinity of these cells significantly decreased their discharge rate. Moreover, the effect of the antagonist was reversed by the agonists and vice versa. In contrast, "waking-related" neurons were unaffected by these H3-receptor ligands. These data provide evidence for the histaminergic nature of "W-on" neurons and their role in cortical desynchronization during waking, and highlight the heterogeneity of posterior hypothalamic neuronal populations, which might serve different functions during the wakefulness.

Polyunsaturated fatty acids in the blood of spontaneously or induced muricidal male Wistar rats.

Serum levels of several n-6 and n-3 polyunsaturated fatty acids were compared in male Wistar muricidal (Mu) and non-Mu rats. The Mu behavior was either spontaneous or induced by long-term isolation, feeding with a starch-enriched polyunsaturated fatty acid diet (PUFA+S), water restriction, or adrenalectomy (ADX). Arachidonic acid (ARA) levels were lower in diet-induced (PUFA+S) Mu rats than in their non-Mu controls. Total n-6 fatty acid levels were also lower in spontaneously Mu rats than in spontaneously non-Mu rats. Docosahexaenoic acid (DHA) and total n-3 fatty acids levels were lower in rats with isolation-induced Mu behavior. The n-3/n-6 ratio was higher in spontaneously Mu rats than in spontaneously non-Mu rats. The changes in ARA levels were greater than those in DHA levels, possibly due to the higher blood-brain barrier passage of arachidonic acid. The results were analyzed in the light of recent results showing a role of PUFAs in human and animal behavior.

Effect of noradrenergic denervation of medial prefrontal cortex and dentate gyrus on recovery after sleep deprivation in the rat.

The noradrenergic-locus coeruleus (LC) system has a regulatory influence on forebrain neuronal networks. We have previously shown that the amygdala is strongly implicated in the mechanism of rebound seen after a 10 h sleep deprivation (SD). In the present study, our objective was to determine whether the medial prefrontal cortex and dentate gyrus (DG) which receive an important innervation from the LC, play a role in the rebound mechanisms. We found that microinjection of the specific noradrenergic neurotoxin, N-(2-chloroethyl)-N-ethyl-2-bromobenzylamine, into these regions had no effect on the increase in paradoxical sleep duration seen after SD, suggesting that noradrenergic (NA) innervation of the prefrontal cortex and DG are not involved in sleep rebound regulation.

Sleep and epilepsy: A key role for nitric oxide?

PURPOSE: It has been suggested that nitric oxide (NO) is involved in sleep mechanisms and in the pathophysiology of epilepsy. Data are, however, controversial because it is not clear whether NO facilitates sleep or waking, or whether it exerts pro-or antiepileptic influences. METHODS: The question was considered through NO voltammetric measurements and electroencephalographic recordings performed in GAERS rats (Genetic Absence Epilepsy Rat from Strasbourg): an experimental model of "petit-mal" human disease. Regulatory processes of sleep and epilepsy were studied after administration of a NO synthase inhibitor [l-arginine-p-nitroanilide (l-ANA) 100 mg/kg i.p.], a NO donor (SIN-1 100 ng/2 microl i.c.v.), and the antiepileptic drugs used in clinic [valproate (VPA 200 mg/kg i.p.) and ethosuximide (ESM 100 mg/kg i.p.)]. RESULTS: In GAERS rats, spontaneous circadian organizations of spike-wave discharges and paradoxical sleep (PS) occur in an opposite way; spontaneous NO concentrations are higher during seizures than during wakefulness, slow-wave sleep, and PS, respectively. l-ANA induces a disappearance of NO peak, an epileptic induction, and a loss of PS while SIN-1 induces opposite effects. Antiepileptic effects of VPA and ESM are associated with a PS increase and a significant release of NO. CONCLUSIONS: These results indicate that NO could be, in GAERS rats, a central piece in the reciprocal inhibitory mechanisms regulating the induction of PS and spike-wave discharges. NO could prevent absence epilepsy and act as an antiepileptic substance in facilitating PS. Antiepileptic efficiency of VPA and ESM may work through their ability to release NO. A track for a new treatment of petit-mal disease in children can be envisioned.

Effect of noradrenergic denervation of the amygdala upon recovery after sleep deprivation in the rat.

We previously showed that the noradrenergic locus coeruleus (NA-LC) was involved in the regulatory mechanisms of the paradoxical sleep rebound following a 10 h sleep deprivation by using a systemic injection of a specific neurotoxin, N-(2-chloroethyl)-N-ethyl-2-bromobenzylamine (DSP-4). Given that rebound mechanisms are mainly located in the forebrain, we planned to study the role of the forebrain structures receiving LC afferences. In this study we evaluated the involvement of noradrenergic afferences to the central nucleus of the amygdala in the sleep rebound by DSP-4 microinjections into the central nucleus of the rat amygdala. The results showed that during the first recovery day, the paradoxical sleep rebound is lower in DSP-4 treated rats (-67.28%). These findings indicate that the amygdala, through its NA afferents, contributes to the sleep rebound mechanisms.

Influence of a 1-h immobilization stress on sleep and CLIP (ACTH(18-39)) brain contents in adrenalectomized rats.

Basal sleep amounts in adrenalectomized rats (AdX), as compared to intact animals, exhibit a significant increase in slow-wave sleep (SWS), a tendency towards an increase in paradoxical sleep (PS), and circadian rhythms (SWS and PS) flattened in amplitude. An immobilization stress (IS) of 1 h, imposed on AdX rats at the beginning of the dark period, is accompanied by an intense polygraphic waking. Just after the IS, SWS amount become significantly higher than in control rats (+44%/11 h of darkness) whereas significant increases of PS occur only 5-10 h after the IS (+24%/11 h of darkness). A specific radioimmunoassay for CLIP (corticotropin-like intermediate lobe peptide or ACTH(18-39)) was performed in biopsies taken either from the nucleus raphe dorsalis (nRD) or the arcuate nucleus (AN). In the nRD, just after the IS, phosphorylated CLIP (Ph-CLIP) concentration exhibits a decreasing tendency, but 4 h later, it increases significantly (+22%, p<0.05). In the AN, Ph-CLIP concentration remains unchanged after the IS as well as 4 h later. These results differ from those previously reported in intact animals also submitted to a 1-h IS, that is, a SWS rebound less marked (+27%/11 h of darkness), a PS rebound more important starting immediately after the IS (+46%/11 h of darkness) and a significant increase in Ph-CLIP occurring just after the end of the restraint. In conclusion, data obtained after a restraint stress either in AdX or in control rats point out the dependence of the PS rebound on the nRD Ph-CLIP concentration.

Hypoprolactinemic rats under conditions of constant darkness or constant light. Effects on the sleep-wake cycle, cerebral temperature and sulfatoxymelatonin levels.

In genetic hypoprolactinemic rats under light-dark (LD) conditions, the circadian rhythms of slow-wave (SWS) and paradoxical (PS) sleep display an alteration of their phase relationship. The aim of our study was to investigate the effects of constant darkness (DD) or constant light (LL) on the daily distribution and amounts of sleep-wake stages, cerebral temperature and concentrations of the urinary melatonin metabolite, 6-sulfatoxymelatonin, in prolactin-deficient rats. After 3 weeks of DD, the SWS period was 24 h 8+/-6 min and the acrophase occurred at 15:44+/-1:35, while for PS, the period was more stable than during LD (24 h 10+/-8 min vs. 24 h 55+/-43 min) and the acrophase occurred at 16:44+/-1:54. Under LL conditions, circadian sleep rhythms persisted during the first 3 days, then completely disappeared during the third week, to be replaced by ultradian rhythms (period of 4-6 h). Time-series analysis showed that the two sleep states became synchronous as early as the second day under constant conditions. The total amount of PS was increased under both conditions (LL and DD) at the expense of duration of waking. Under LD and constant conditions, the pattern of changes in cerebral temperature was similar to that for wakefulness (W). Sulfatoxymelatonin was rhythmically secreted under both LD and DD conditions, whereas, under LL conditions, its rhythm was abolished. The results show that, in IPL rats in the absence of a zeitgeber, the PS and SWS rhythms recover a synchronous phase relationship and PS amounts are increased.

Noradrenaline neurotoxin DSP-4 effects on sleep and brain temperature in the rat.

N-(2-chloroethyl)-N-ethyl-2-bromobenzylamine (DSP-4) has a selective degenerative effect on noradrenergic fibers originating from locus coeruleus (LC) neurons. In the present study, we studied its effect on vigilance states and brain temperature by continuous recordings for periods of 1-5 days and 2-4 weeks following DSP-4 treatment. On the first day, paradoxical sleep duration was significantly decreased (-67%, P < 0.05), slow-wave sleep (SWS) duration increased (+16%, P < 0.05) up to 48 h after DSP-4 treatment (+8%, P < 0.05) and the wake period decreased (-8%, P < 0.05). The vigilance states returned to control values 4-5 days later. The brain temperature was decreased during the first night (-2 degrees C) and then recovered the control values. Two and 4 weeks after DSP-4 treatment, paradoxical sleep was still decreased (-18% and -23%, respectively, P < 0.05), while SWS was significantly increased only at night during the fourth week (+23%, P < 0.05). These results therefore provide evidence for a differential involvement of the noradrenergic LC system in sleep mechanisms depending on the light-dark cycle. Different hypotheses are proposed.

A key role for the caudoventral pontine tegmentum in the simultaneous generation of eye saccades in bursts and associated ponto-geniculo-occipital waves during paradoxical sleep in the cat.

Ponto-geniculo-occipital waves and rapid eye movements (eye saccades) are two prominent phasic events of paradoxical sleep which occur in conjunction. Although they have been studied intensively, the neuronal link between these two events is still poorly understood. On the basis of our previous results, combining brainstem transections and carbachol microinjections, we postulated that the oculomotor and ponto-geniculo-occipital systems do not work in series, but in parallel, and that the caudoventral pontine tegmentum might represent a structure controlling and/or co-ordinating the simultaneous production of the two phenomena. This hypothesis was further supported by the demonstration that, during paradoxical sleep, the instantaneous velocity of eye saccades in bursts is higher than that of isolated ones which, in turn, are more rapid than waking saccades. This acceleration of eye saccades in bursts also seems to be under the cholinergic control of the caudoventral pontine tegmentum. In order to test the hypothesis that this area may be a prime mover leading to the simultaneous appearance of these two phasic events as a whole, we investigated, in the present study, the effects of pharmacological stimulation (with carbachol) and inhibition (with atropine) of the caudoventral pontine tegmentum on the production and the characteristics of eye saccades and ponto-geniculo-occipital waves. Cats' eye movements were recorded using the technique of the scleral search coil in a magnetic field, together with sleep-waking parameters. We found that: (i) unilateral microinjections of carbachol (0.4 microg) induced, during waking, a majority of long bursts of ponto-geniculo-occipital waves (i.e. bursts containing at least five waves) which had intra-burst intervals similar to natural ones (48-259 ms) and decreased the frequency of isolated ponto-geniculo-occipital waves; (ii) unilateral microinjections of atropine (2.4 microg) strongly decreased, during paradoxical sleep, the frequency (number/min) of eye saccades in bursts directed contralaterally to the side of the injection (by 48-54%) and reduced the velocity of these saccades to that of isolated eye saccades. Atropine also significantly reduced the frequency (by 60%) of all types of bursts of ponto-geniculo-occipital waves, with a maximal effect (80% reduction) on long bursts of ponto-geniculo-occipital waves, while it increased the frequency of isolated ponto-geniculo-occipital waves. However, atropine did not change the value of intra-burst intervals. These findings support the hypothesis that eye saccades in bursts and associated ponto-geniculo-occipital waves are generated as a whole by a common structure and that this structure is at least partly defined by the caudoventral pontine tegmentum.

Influence of a 1 h immobilization stress on sleep states and corticotropin-like intermediate lobe peptide (CLIP or ACTH18-39, Ph-ACTH18-39) brain contents in the rat.

A 1 h immobilization stress (IS) was imposed to rats at the beginning of the dark period, i.e., when the animals start to be active. The IS was accompanied by an intense polygraphic waking and followed, over 12 h of the dark period, by a significant rebound of slow-wave sleep (SWS, +17%) and paradoxical sleep (PS, +57%). In order to estimate the IS-related changes in the endogenous concentrations of corticotropin-like intermediate lobe peptide (CLIP, ACTH18-39) and related compounds, a specific radioimmunoassay (RIA) was used. Assays performed in cerebral biopsies taken from arcuate (AN) and raphe dorsalis (nRD) nuclei led to the obtention of 2 main immunoreactive peaks, corresponding to CLIP and its phosphorylated form Ph-CLIP. Just after end of the IS and within the nRD. Ph-CLIP immunoreactivity increased by about 95%. Four hours later, i.e., when PS rebound was maximal, a 37% increase in Ph-CLIP immunoreactivity was measured in the AN. These observations have never been described before. In the blood, at the end of the restraint, CLIP/ACTH1-39 total immunoreactivity was increased by 330%. It returned to baseline level 4 h later. Blood concentration of corticosterone was also increased by 56% at the end of the IS and was close to baseline level 4 h later. Data reported here indicate that the IS first triggers an increase in Ph-CLIP within the nRD. Since the nRD contains sleep permissive components, this increase might be determinant for the SWS and PS rebound induction. The changes observed in the blood as regards CLIP/ACTH1-39 total immunoreactivity and corticosterone concentration testify to the efficacy of the IS and are part of the conventional picture accompanying such a situation. Finally, the increase in Ph-CLIP, occurring in the AN 4 h after the end of the restraint, might be part of the restorative processes necessary to compensate the stress overshoot.

Role of the locus coeruleus in the sleep rebound following two different sleep deprivation methods in the rat.

The aim of the present study was to assess the involvement of the locus coeruleus in the paradoxical sleep rebound following sleep deprivation in the rat. Animals were sleep-deprived for 10 h before, and after, specific N-(2-chloroethyl)-N-ethyl-2-bromobenzylamine (DSP-4) lesioning of the noradrenergic-locus coeruleus system. Sleep deprivation was produced using either an instrumental (water tank) or pharmacological (methylamphetamine) method. After lesioning, the rats submitted to the instrumental method showed a significant decrease in the paradoxical and slow-wave sleep rebounds (-54% and -78%, respectively), while animals receiving metamphetamine did not. Our results suggest that the noradrenergic system of the locus coeruleus is a relevant component of the sleep rebound mechanisms. However, the extent of involvement is dependent on the sleep deprivation method used.

Effects of tianeptine, sertraline and clomipramine on brain serotonin metabolism: a voltammetric approach in the rat.

Tianeptine is a substance enhancing the serotonir uptake while sertraline and clomipramine inhibit it. By means of 5-hydroxyin-doleacetic acid (5-HIAA) voltammetric measurements, this study investigated their influence on serotonin metabolism which depends mainly upon the activity of monoamine oxidase type A. After tianeptine injection the 5-HIAA signal increased by about 60%. This effect was maintained when the animals were pre-treated with MDL 72145 (an inhibitor of monoamine oxidase type B) but reduced when clorgyline (an inhibitor of monoamine oxidase type A) was administered after tianeptine. Administration of sertraline or clomipramine reduced the 5-HIAA signal by about 30-50%, whether the animals were pre-treated with MDL 72145 or not. It is to be concluded that tianeptine, sertraline and clomipramine can regulate the 5-HT fraction present in the synaptic cleft, not only by acting at the level of the serotoninergic neurons, but also by favoring or reducing the access of the amine to monoamine oxidase type A which is synthesized within non-serotoninergic neurons and glial cells.

The caudo ventral pontine tegmentum is involved in the generation of high velocity eye saccades in bursts during paradoxical sleep in the cat.

Cat eye movements were recorded in the head restrained condition, with the technique of the scleral search coil in a magnetic field, and the maximum velocity/amplitude relationships were analyzed for saccades in the following conditions, (1) during waking (W); (2) during paradoxical sleep (PS); and (3) during W following carbachol microinjections in the medioventral part of the caudal pontine tegmentum. These findings indicate that (1) the neurophysiological mechanisms underlying carbachol induced events are similar to those acting during PS and that the caudal pontine tegmentum might be the generator of high velocity eye saccades in bursts accompanied by ponto geniculo occipital waves (PGOw) during PS, and (2) caudal pontine tegmentum neurons show 'state-dependent' responsiveness to cholinergic inputs, suggesting that a change in the synaptic inputs and/or the membrane properties of these neurons during PS may be responsible for the induction of saccadic eye movements in bursts and associated PGOw.

Sleep increase after immobilization stress: role of the noradrenergic locus coeruleus system in the rat.

In a preliminary study we showed that the sleep rebound occurring after sleep deprivation is decreased in rats treated with N-(2-chloroethyl)-N-ethyl-2-bromobenzylamine (DSP-4), a neurotoxic agent specific for the noradrenergic cells of the locus coeruleus (LC). Sleep deprivation methods not only involve sleep loss, but also stress, which per se may induce an increase in sleep duration. Extensive research showed that the locus coeruleus is involved in stress. To evaluate the participation of LC in this mechanism, the effect of DSP-4 treatment was studied on sleep duration following a short intense stress in the absence of sleep loss. The results showed that the augmentation of sleep after 1 h of immobilization stress is lower in DSP-4-treated rats (slow-wave sleep duration, -24%; paradoxical sleep duration, -52%). These findings suggest that the increase in sleep induced by such a stressor is mediated, at least in part, by the noradrenergic LC.

Corpus spongiosum penis pressure and perineal muscle activity during reflexive erections in the rat.

This experiment represents the first simultaneous analysis of corpus spongiosum of the penis (CSP) pressure and perineal muscle activity during reflex-induced penile erections in the awake rat and provides new data on the coordination of muscular and vascular events in erection physiology. Nine male Sprague-Dawley rats were implanted with pressure transducers under pentobarbital anesthesia (60 mg/kg) for use in a new technique of chronic erection recording involving simultaneous CSP pressure monitoring and electromyography (EMG) of the ischiocavernosus (IC) and bulbospongiosus (BS) muscles. The association of these physiological measurements with visually scored ex copula erectile events was investigated. Reflexive erectile events were systematically associated with an increase in baseline CSP pressure. Glans erections were associated with dramatic suprasystolic CSP pressure peaks concurrent with BS muscle bursts. Indeed, measures of CSP pressure and BS EMG activity were found to vary significantly with the intensity of glans erection. Flips of the penile body, however, involved small CSP pressure increases and IC bursts. These data demonstrate for the first time that both CSP pressures and perineal muscle activity vary significantly with the intensity of penile erection.

Effects of an acute immobilization stress upon proopiomelanocortin (POMC) mRNA levels in the mediobasal hypothalamus: a quantitative in situ hybridization study.

The aim of this study was to examine by quantitative in situ hybridization the effects of an acute stress on the expression of the POMC gene in the mediobasal hypothalamus (MBH) of the rat. In control animals, the highest levels of POMC mRNA were observed in the posterior periventricular region of the MBH. Lower levels were found in the anterior and posterior arcuate nucleus. At the end of a one hour immobilization, a small decrease (-8%) was observed in the periventricular region only. Four hours after the end of immobilization, increases in POMC mRNA levels were detected in the anterior part (7%), in the posterior part (25%) and in the periventricular region (13%) of the MBH. These results suggest that MBH POMC-derived peptides might be an important component in the central response to stress.

Ontogenesis of nocturnal organization of sleep spindles: a longitudinal study during the first 6 months of life.

Ontogenesis of sleep spindles was studied on overnight longitudinal recordings in 12 full-term infants at 1.5-3-4.5 and 6 months of life. Six parameters (density, duration, frequency, amplitude, asymmetry and asynchrony) were analyzed during both slow wave sleep (SII and delta) and during 5 periods of the night. Results show a significant increase of most parameters between 1.5 and 3 months of age. All spindle patterns developed quite rapidly during the first 3 months of infancy, possibly reflecting developmental changes in thalamo-cortical structures and maturation of the physiological system that produces spindles. The density of 12-14 Hz spindle frequency was higher in stage II when compared to stage delta, as in adults. Our data confirm previous reports on spindle ontogenesis and give a more complete aspect of this ontogenesis in relation to sleep development. Three months of age appeared to be a turning point in maturational processes and might reflect changes in central nervous system activity and behavior which take place during that period. Sleep spindle evolution seems to be an accurate reflection of the slow wave sleep (SWS) development, and our results are discussed in terms of the developmental aspect of SWS production and characterization of sleep stages in young infants. Concordance between quantitative aspects and nocturnal organization leads us to consider that the individualization of slow wave sleep (SWS) in infants occurs from 4.5 months of life.

Benzodiazepine binding sites and their modulators in hippocampus of violent suicide victims.

Benzodiazepine binding sites were studied by autoradiography in several hippocampic layers in brains of drug-free violent suicide victims (hanging) and matched controls. Kd was increased in suicides, and when brain sections from control subjects were incubated in the bath fluid that had previously served to incubate sections from suicides, Kd was increased in the same way. Results are discussed in terms of possible modulators of benzodiazepine binding sites, mainly tissue GABA and amino acid concentrations.