Maturation of sleep homeostasis in developing rats: a role for preoptic area neurons

The present study evaluated the hypothesis that developmental changes in hypothalamic sleep-regulatory neuronal circuits contribute to the maturation of sleep homeostasis in rats during the fourth postnatal week. In a longitudinal study, we quantified electrographic measures of sleep during baseline and in response to sleep deprivation (SD) on postnatal days 21/29 (P21/29) and P22/30 (experiment 1). During 24-h baseline recordings on P21, total sleep time (TST) during the light and dark phases did not differ significantly. On P29, TST during the light phase was significantly higher than during the dark phase. Mean duration of non-rapid-eye-movement (NREM) sleep bouts was significantly longer on P29 vs. P21, indicating improved sleep consolidation. On both P22 and P30, rats exhibited increased NREM sleep amounts and NREM electroencephalogram delta power during recovery sleep (RS) compared with baseline. Increased NREM sleep bout length during RS was observed only on P30. In experiment 2, we quantified activity of GABAergic neurons in median preoptic nucleus (MnPN) and ventrolateral preoptic area (VLPO) during SD and RS in separate groups of P22 and P30 rats using c-Fos and glutamic acid decarboxylase (GAD) immunohistochemistry. In P22 rats, numbers of Fos(+)GAD(+) neurons in VLPO did not differ among experimental conditions. In P30 rats, Fos(+)GAD(+) counts in VLPO were elevated during RS. MnPN neuronal activity was state-dependent in P22 rats, but Fos(+)GAD(+) cell counts were higher in P30 rats. These findings support the hypothesis that functional emergence of preoptic sleep-regulatory neurons contributes to the maturation of sleep homeostasis in the developing rat brain.     

Sleep-promoting functions of the hypothalamic median preoptic nucleus: inhibition of arousal systems

Recent work supports the hypotheses developed by von Economo and Nauta and elaborated by Sallanon et al. that the POA contains a sleep-promoting output that opposes wake-promoting neuronal groups in the PH. The POA gives rise to descending pathways that terminate within wake-promoting populations in pLH, PH and midbrain. Current evidence suggests that this output originates in POA sleep-active GABAergic neurons. This output also seems to convey the signals of homeostatic drive. Disynaptic projections from the SCN to both MnPN and VLPO were recently identified. These may regulate the circadian control of sleep propensity. The hypothesis that the descending projections from POA sleep-active neurons to sites of arousal-related neurons originates in GABAergic neurons must be confirmed. Also to be further clarified is the anatomical distribution of putative sleep-active GABAergic neurons within the POA. Segregated groups have been found in the MnPN and VLPO, but unit recording studies of sleep-active neurons, lesion studies and local neurochemical application studies all indicate that sleep-active neurons may be found diffusely in the POA and adjacent areas. The MnPN has been shown previously to be involved in water balance and blood pressure regulation and to be responsive to hyperthermia. Our studies suggest that this nucleus also contains sleep-active, putative sleep-promoting neurons. However, interactions between sleep control and physiological variables must be considered. In particular, the details of neuronal basis of the coupling of warm-sensitive neurons in MnPN to the POA hypnogenic output has not been explored. It is also worth noting that both the VLPO and MnPN lie close to the ventricular and subarachnoid surface and are punctuated by radial arterioles. The possibility that the sleep-regulatory functions of these sites is coupled to physiological signals conveyed through epithelial cells has been suggested for the actions of PGD2 but has yet to be explored in detail for other putative hypnogens.     

What are the mechanisms activating the sleep-active neurons located in the preoptic area?

The purpose of this review is to outline the mechanisms responsible for the induction and maintenance of slow-wave sleep (SWS, also named non–rapid eye movement or non-REM sleep). The latest hypothesis on the mechanisms by which cortical activity switches from an activated state during waking to a synchronised state during SWS is presented. It is proposed that the activated cortical state during waking is induced by the activity of multiple waking systems, including the serotonergic, noradrenergic, cholinergic and hypocretin systems located at different subcortical levels. In contrast, the neurons inducing SWS are mainly localized in the ventrolateral preoptic (VLPO) and median preoptic nuclei. These neurons use the inhibitory neurotransmitter gamma-aminobutyric acid (GABA). The notion that the switch from waking to SWS is due to the inhibition of the waking systems by the VLPO sleep-active neurons is introduced. At the onset of sleep, the sleep neurons are activated by the circadian clock localized in the suprachiasmatic nucleus and a powerful hypnogenic factor, adenosine, which progressively accumulates in the brain during waking.

     

Preoptic hypothalamic warming suppresses laryngeal dilator activity during sleep

Upper airway dilator activity during sleep appears to be diminished under conditions of enhanced sleep propensity, such as after sleep deprivation, leading to worsening of obstructive sleep apnea (OSA). Non-rapid eye movement (NREM) sleep propensity originates in sleep-active neurons of the preoptic area (POA) of the hypothalamus and is facilitated by activation of POA warm-sensitive neurons (WSNs). We hypothesized that activation of WSNs by local POA warming would inhibit activity of the posterior cricoarytenoid (PCA) muscle, an airway dilator, during NREM sleep. In chronically prepared unrestrained cats, the PCA exhibited inspiratory bursts in approximate synchrony with inspiratory diaphragmatic activity during waking, NREM, and REM. Integrated inspiratory PCA activity (IA), peak activity (PA), and the lead time (LT) of the onset of inspiratory activity in PCA relative to diaphragm were significantly reduced in NREM sleep and further reduced during REM sleep compared with waking. Mild bilateral local POA warming (0.5-1.2 degrees C) significantly reduced IA, PA, and LT during NREM sleep compared with a prewarming NREM baseline. In some animals, effects of POA warming on PCA activity were found during waking or REM. Because POA WSN activity is increased during spontaneous NREM sleep and regulates sleep propensity, we hypothesize that this activation contributes to reduction of airway dilator activity in patients with OSA.     

Rhythms in Fos expression in brain areas related to the sleep-wake cycle in the diurnal Arvicanthis niloticus

Most mammals show daily rhythms in sleep and wakefulness controlled by the primary circadian pacemaker, the suprachiasmatic nucleus (SCN). Regardless of whether a species is diurnal or nocturnal, neural activity in the SCN and expression of the immediate-early gene product Fos increases during the light phase of the cycle. This study investigated daily patterns of Fos expression in brain areas outside the SCN in the diurnal rodent Arvicanthis niloticus. We specifically focused on regions related to sleep and arousal in animals kept on a 12:12-h light-dark cycle and killed at 1 and 5 h after both lights-on and lights-off. The ventrolateral preoptic area (VLPO), which contained cells immunopositive for galanin, showed a rhythm in Fos expression with a peak at zeitgeber time (ZT) 17 (with lights-on at ZT 0). Fos expression in the paraventricular thalamic nucleus (PVT) increased during the morning (ZT 1) but not the evening activity peak of these animals. No rhythm in Fos expression was found in the centromedial thalamic nucleus (CMT), but Fos expression in the CMT and PVT was positively correlated. A rhythm in Fos expression in the ventral tuberomammillary nucleus (VTM) was 180 degrees out of phase with the rhythm in the VLPO. Furthermore, Fos production in histamine-immunoreactive neurons of the VTM cells increased at the light-dark transitions when A. niloticus show peaks of activity. The difference in the timing of the sleep-wake cycle in diurnal and nocturnal mammals may be due to changes in the daily pattern of activity in brain regions important in sleep and wakefulness such as the VLPO and the VTM.     

Molecular mechanisms of sleep-wake regulation: a role of prostaglandin D2

Prostaglandin (PG) D2 is a major prostanoid in the brains of rats and other mammals, including humans. When PGD synthase (PGDS), the enzyme that produces PGD2 in the brain, was inhibited by the intracerebroventricular infusion of its selective inhibitors, i.e. tetravalent selenium compounds, the amount of sleep decreased both time and dose dependently. The amount of sleep of transgenic mice, in which the human PGDS gene had been incorporated, increased several fold under appropriate conditions. These data indicate that PGDS is a key enzyme in sleep regulation. In situ hybridization, immunoperoxidase staining and direct enzyme activity determination of tissue samples revealed that PGDS is hardly detectable in the brain parenchyma but is localized in the membrane systems surrounding the brain, namely, the arachnoid membrane and choroid plexus, from which it is secreted into the cerebrospinal fluid (CSF) to become beta-trace, a major protein component of the CSF. PGD2 exerts its somnogenic activity by binding to PGD2 receptors exclusively localized at the ventrorostral surface of the basal forebrain. When PGD2 was infused into the subarachnoid space below the rostral basal forebrain, striking expression of proto-oncogene Fos immunoreactivity (FosIR) was observed in the ventrolateral preoptic area (VLPO), a putative sleep centre, concurrent with sleep induction. Fos expression in the VLPO was positively correlated with the preceding amount of sleep and negatively correlated with Fos expression in the tuberomammillary nucleus (TMN), a putative wake centre. These observations suggest that PGD2 may induce sleep via leptomeningeal PGD2 receptors with subsequent activation of the VLPO neurons and downregulation of the wake neurons in the TMN area. Adenosine may be involved in the signal transduction associated with PGD2.     

Effects on sleep of microdialysis of adenosine A1 and A2a receptor analogs into the lateral preoptic area of rats

Evidence suggests that adenosine (AD) is an endogenous sleep factor. The hypnogenic action of AD is mediated through its inhibitory A1 and excitatory A2A receptors. Although AD is thought to be predominantly active in the wake-active region of the basal forebrain (BF), a hypnogenic action of AD has been demonstrated in several other brain areas, including the preoptic area. We hypothesized that in lateral preoptic area (LPOA), a region with an abundance of sleep-active neurons, AD acting via A1 receptors would induce waking by inhibition of sleep-active neurons and that AD acting via A2A receptors would promote sleep by stimulating the sleep-active neurons. To this end, we studied the effects on sleep of an AD transport inhibitor, nitrobenzyl-thio-inosine (NBTI) and A1 and A2A receptor agonists/antagonists by microdialyzing them into the LPOA. The results showed that, in the sleep-promoting area of LPOA: 1) A1 receptor stimulation or inhibition of AD transport by NBTI induced waking and 2) A2A receptor stimulation induced sleep. We also confirmed that NBTI administration in the wake promoting area of the BF increased sleep. The effects of AD could be mediated either directly or indirectly via interaction with other neurotransmitter systems. These observations support a hypothesis that AD mediated effects on sleep-wake cycles are site and receptor dependent.     

Compensatory sleep response to 12 h wakefulness in young and old rats

There is a pronounced decline in sleep with age. Diminished output from the circadian oscillator, the suprachiasmatic nucleus, might play a role, because there is a decrease in the amplitude of the day-night sleep rhythm in the elderly. However, sleep is also regulated by homeostatic mechanisms that build sleep drive during wakefulness, and a decline in these mechanisms could also decrease sleep. Because this question has never been addressed in old animals, the present study examined the effects of 12 h wakefulness on compensatory sleep response in young (3.5 mo) and old (21.5 mo) Sprague-Dawley and F344 rats. Old rats in both strains had a diminished compensatory increase in slow-wave sleep (SWS) after 12 h of wakefulness (0700-1900, light-on period) compared with the young rats. In contrast, compensatory REM sleep rebound was unaffected by age. To assess whether the reduced SWS rebound in old rats might result from loss of neurons implicated in sleep generation, we counted the number of c-Fos immunoreactive (c-Fos-ir) cells in the ventral lateral preoptic (VLPO) area and found no differences between young and old rats. These findings indicate that old rats, similar to elderly humans, demonstrate less sleep after prolonged wakefulness. The findings also indicate that although old rats have a decline in sleep, this cannot be attributed to loss of VLPO neurons implicated in sleep.     

Estradiol replacement enhances sleep deprivation-induced c-Fos immunoreactivity in forebrain arousal regions of ovariectomized rats

To understand how female sex hormones influence homeostatic mechanisms of sleep, we studied the effects of estradiol (E(2)) replacement on c-Fos immunoreactivity in sleep/wake-regulatory brain areas after sleep deprivation (SD) in ovariectomized rats. Adult rats were ovariectomized and implanted subcutaneously with capsules containing 17beta-E(2) (10.5 microg; to mimic diestrous E(2) levels) or oil. After 2 wk, animals with E(2) capsules received a single subcutaneous injection of 17beta-E(2) (10 microg/kg; to achieve proestrous E(2) levels) or oil; control animals with oil capsules received an oil injection. Twenty-four hours later, animals were either left undisturbed or sleep deprived by "gentle handling" for 6 h during the early light phase, and killed. E(2) treatment increased serum E(2) levels and uterus weights dose dependently, while attenuating body weight gain. Regardless of hormonal conditions, SD increased c-Fos immunoreactivity in all four arousal-promoting areas and four limbic and neuroendocrine nuclei studied, whereas it decreased c-Fos labeling in the sleep-promoting ventrolateral preoptic nucleus (VLPO). Low and high E(2) treatments enhanced the SD-induced c-Fos immunoreactivity in the laterodorsal subnucleus of the bed nucleus of stria terminalis and the tuberomammillary nucleus, and in orexin-containing hypothalamic neurons, with no effect on the basal forebrain and locus coeruleus. The high E(2) treatment decreased c-Fos labeling in the VLPO under nondeprived conditions. These results indicate that E(2) replacement modulates SD-induced or spontaneous c-Fos expression in sleep/wake-regulatory and limbic forebrain nuclei. These modulatory effects of E(2) replacement on neuronal activity may be, in part, responsible for E(2)'s influence on sleep/wake behavior.     

Quantitative physiologically based modeling of subjective fatigue during sleep deprivation

A quantitative physiologically based model of the sleep-wake switch is used to predict variations in subjective fatigue-related measures during total sleep deprivation. The model includes the mutual inhibition of the sleep-active neurons in the hypothalamic ventrolateral preoptic area (VLPO) and the wake-active monoaminergic brainstem populations (MA), as well as circadian and homeostatic drives. We simulate sleep deprivation by introducing a drive to the MA, which we call wake effort, to maintain the system in a wakeful state. Physiologically this drive is proposed to be afferent from the cortex or the orexin group of the lateral hypothalamus. It is hypothesized that the need to exert this effort to maintain wakefulness at high homeostatic sleep pressure correlates with subjective fatigue levels. The model's output indeed exhibits good agreement with existing clinical time series of subjective fatigue-related measures, supporting this hypothesis. Subjective fatigue, adrenaline, and body temperature variations during two 72 h sleep deprivation protocols are reproduced by the model. By distinguishing a motivation-dependent orexinergic contribution to the wake-effort drive, the model can be extended to interpret variation in performance levels during sleep deprivation in a way that is qualitatively consistent with existing, clinically derived results. The example of sleep deprivation thus demonstrates the ability of physiologically based sleep modeling to predict psychological measures from the underlying physiological interactions that produce them.     

Central pathway for spontaneous and prostaglandin E2-evoked cutaneous vasoconstriction

A reduction of heat loss to the environment through increased cutaneous vasoconstrictor (CVC) sympathetic outflow contributes to elevated body temperature during fever. We determined the role of neurons in the dorsomedial hypothalamus (DMH) in increases in CVC sympathetic tone evoked by PGE2 into the preoptic area (POA) in chloralose/urethane-anesthetized rats. The frequency of axonal action potentials of CVC sympathetic ganglion cells recorded from the surface of the tail artery was increased by 1.8 Hz following nanoinjections of bicuculline (50 pmol) into the DMH. PGE2 nanoinjection into the POA elicited a similar excitation of tail CVC neurons (+2.1 Hz). Subsequent to PGE2 into the POA, muscimol (400 pmol/side) into the DMH did not alter the activity of tail CVC neurons. Inhibition of neurons in the rostral raphé pallidus (rRPa) eliminated the spontaneous discharge of tail CVC neurons but only reduced the PGE2-evoked activity. Residual activity was abolished by subsequent muscimol into the rostral ventrolateral medulla. Transections through the neuraxis caudal to the POA increased the activity of tail CVC neurons, which were sustained through transections caudal to DMH. We conclude that while activation of neurons in the DMH is sufficient to activate tail CVC neurons, it is not necessary for their PGE2-evoked activity. These results support a CVC component of increased core temperature elicited by PGE2 in POA that arises from relief of a tonic inhibition from neurons in POA of CVC sympathetic premotor neurons in rRPa and is dependent on the excitation of CVC premotor neurons from a site caudal to DMH.     

Melanopsin as a Sleep Modulator: Circadian Gating of the Direct Effects of Light on Sleep and Altered Sleep Homeostasis in Opn4?/? Mice

Light influences sleep and alertness either indirectly through a well-characterized circadian pathway or directly through yet poorly understood mechanisms. Melanopsin (Opn4) is a retinal photopigment crucial for conveying nonvisual light information to the brain. Through extensive characterization of sleep and the electrocorticogram (ECoG) in melanopsin-deficient (Opn4^−/−) mice under various light–dark (LD) schedules, we assessed the role of melanopsin in mediating the effects of light on sleep and ECoG activity. In control mice, a light pulse given during the habitual dark period readily induced sleep, whereas a dark pulse given during the habitual light period induced waking with pronounced theta (7–10 Hz) and gamma (40–70 Hz) activity, the ECoG correlates of alertness. In contrast, light failed to induce sleep in Opn4^−/− mice, and the dark-pulse-induced increase in theta and gamma activity was delayed. A 24-h recording under a LD 1-h∶1-h schedule revealed that the failure to respond to light in Opn4^−/− mice was restricted to the subjective dark period. Light induced c-Fos immunoreactivity in the suprachiasmatic nuclei (SCN) and in sleep-active ventrolateral preoptic (VLPO) neurons was importantly reduced in Opn4^−/− mice, implicating both sleep-regulatory structures in the melanopsin-mediated effects of light. In addition to these acute light effects, Opn4^−/− mice slept 1 h less during the 12-h light period of a LD 12∶12 schedule owing to a lengthening of waking bouts. Despite this reduction in sleep time, ECoG delta power, a marker of sleep need, was decreased in Opn4^−/− mice for most of the (subjective) dark period. Delta power reached after a 6-h sleep deprivation was similarly reduced in Opn4^−/− mice. In mice, melanopsin''s contribution to the direct effects of light on sleep is limited to the dark or active period, suggesting that at this circadian phase, melanopsin compensates for circadian variations in the photo sensitivity of other light-encoding pathways such as rod and cones. Our study, furthermore, demonstrates that lack of melanopsin alters sleep homeostasis. These findings call for a reevaluation of the role of light on mammalian physiology and behavior.     

Melanopsin as a Sleep Modulator: Circadian Gating of the Direct Effects of Light on Sleep and Altered Sleep Homeostasis in Opn4−/− Mice

Light influences sleep and alertness either indirectly through a well-characterized circadian pathway or directly through yet poorly understood mechanisms. Melanopsin (Opn4) is a retinal photopigment crucial for conveying nonvisual light information to the brain. Through extensive characterization of sleep and the electrocorticogram (ECoG) in melanopsin-deficient (Opn4^−/−) mice under various light–dark (LD) schedules, we assessed the role of melanopsin in mediating the effects of light on sleep and ECoG activity. In control mice, a light pulse given during the habitual dark period readily induced sleep, whereas a dark pulse given during the habitual light period induced waking with pronounced theta (7–10 Hz) and gamma (40–70 Hz) activity, the ECoG correlates of alertness. In contrast, light failed to induce sleep in Opn4^−/− mice, and the dark-pulse-induced increase in theta and gamma activity was delayed. A 24-h recording under a LD 1-h1-h schedule revealed that the failure to respond to light in Opn4^−/− mice was restricted to the subjective dark period. Light induced c-Fos immunoreactivity in the suprachiasmatic nuclei (SCN) and in sleep-active ventrolateral preoptic (VLPO) neurons was importantly reduced in Opn4^−/− mice, implicating both sleep-regulatory structures in the melanopsin-mediated effects of light. In addition to these acute light effects, Opn4^−/− mice slept 1 h less during the 12-h light period of a LD 1212 schedule owing to a lengthening of waking bouts. Despite this reduction in sleep time, ECoG delta power, a marker of sleep need, was decreased in Opn4^−/− mice for most of the (subjective) dark period. Delta power reached after a 6-h sleep deprivation was similarly reduced in Opn4^−/− mice. In mice, melanopsin's contribution to the direct effects of light on sleep is limited to the dark or active period, suggesting that at this circadian phase, melanopsin compensates for circadian variations in the photo sensitivity of other light-encoding pathways such as rod and cones. Our study, furthermore, demonstrates that lack of melanopsin alters sleep homeostasis. These findings call for a reevaluation of the role of light on mammalian physiology and behavior.     

Incorporation of caffeine into a quantitative model of fatigue and sleep

A recent physiologically based model of human sleep is extended to incorporate the effects of caffeine on sleep-wake timing and fatigue. The model includes the sleep-active neurons of the hypothalamic ventrolateral preoptic area (VLPO), the wake-active monoaminergic brainstem populations (MA), their interactions with cholinergic/orexinergic (ACh/Orx) input to MA, and circadian and homeostatic drives. We model two effects of caffeine on the brain due to competitive antagonism of adenosine (Ad): (i) a reduction in the homestatic drive and (ii) an increase in cholinergic activity. By comparing the model output to experimental data, constraints are determined on the parameters that describe the action of caffeine on the brain. In accord with experiment, the ranges of these parameters imply significant variability in caffeine sensitivity between individuals, with caffeine's effectiveness in reducing fatigue being highly dependent on an individual's tolerance, and past caffeine and sleep history. Although there are wide individual differences in caffeine sensitivity and thus in parameter values, once the model is calibrated for an individual it can be used to make quantitative predictions for that individual. A number of applications of the model are examined, using exemplar parameter values, including: (i) quantitative estimation of the sleep loss and the delay to sleep onset after taking caffeine for various doses and times; (ii) an analysis of the system's stable states showing that the wake state during sleep deprivation is stabilized after taking caffeine; and (iii) comparing model output successfully to experimental values of subjective fatigue reported in a total sleep deprivation study examining the reduction of fatigue with caffeine. This model provides a framework for quantitatively assessing optimal strategies for using caffeine, on an individual basis, to maintain performance during sleep deprivation.     

Changes in cyclic AMP concentration of the rat preoptic region as a result of long term exposure to cold

The cAMP concentration in the preoptic region was studied in rats during exposure to low ambient temperature (Ta: -10 degrees C) and after return to control Ta (22 degrees C). With respect to control cAMP concentration, changes were observed consisting of a decrease (delta cAMP: 4.19 /+- 0.15 pM/mg Pr; p < 0.001) at low Ta and an increase (delta cAMP: 1.40 /+- 0.13 pM/mg Pr; P < 0.05) after return to control Ta. In contrast, cortical cAMP concentration decreased both at low Ta (delta cAMP: 2.94 /+- 0.09 pM/mg Pr; P < 0.005) and after return to control Ta (delta cAMP: 3.21 /+- 0.09 pM/mg Pr; P < 0.001). such cAMP changes in the preoptic region may be related to different activation levels of thermoregulatory and sleep mechanisms.     

Contribution of REM sleep to Fos and FRA expression in the vestibular nuclei of rat leading to vestibular adaptation during the STS-90 Neurolab Mission

1. Electrophysical studies performed in ground-based experiments have shown that VN neurons respond to labyrinthine signals following stimulation of macular gravity receptors. Additional evidence indicates that VN neurons may also respond to extralabyrinthine signals of pontine origin, which occur during the PGO waves typical of REM sleep (Bizzi et al., 1964a, b; cf. also Pompeiano, 1967, 1970, 1974 for ref.). 2. In a previous study (Pompeiano et al., 2002) changes in Fos and FRA expression were used to identify the short-term (Fos) and the long-term (FRA) molecular changes which affect the VN neurons at different time points of the space flight. In particular, while Fos protein persists in the brain tissue only for a few hours (6-8 hrs) after its induction, FRA proteins, which can also be induced in the same experimental conditions, persist in the brain tissue for longer periods of time (i.e. from 12/24 hrs to days). 3. In order to relate the changes in gene expression which occurred in the VN during the space flight either to gravity changes or to REM sleep, we investigated in a recent study (Centini et al, 2006) the changes in Fos and FRA expression which occurred in different phases of the sleep-waking cycle, thus being indicative of the animal state. We could then compare the results obtained during the space lab Mission with those previously observed either in ground-based experiments during the physiological state of waking and slow-wave (SWS) or during neurochemically induced episodes of PS, as obtained after microinjection of appropriate agents in dorsal pontine structures of rats. 4. Our findings indicated that a waking state possibly associated with episodes of SWS, occurred at FD2 and FD14, i.e. at launch and after exposure of the animal to microgravity. It appeared also that at the reentry (R + 1) rather than at launch (FD2), an increase in Fos and FRA expression affected the noradrenergic LC neurons, as well as several related structures. These findings probably resulted from the acceleration stress, or immobilization stress as shown by the appearance of a starle reaction (or arrest reaction) which occurred after landing. This condition of stress was followed after landing by an increase in Fos and FRA expression which affected ventromedial medullary reticular structures, whose descending projections are involved in the suppression of postural activity during PS. Moreover, their ascending projections were likely to increase the FRA expression in the neocortex as well as in several regions of the limbic system, such as the dentate gyrus and the hippocampus, which lead to EEG desynchronization and the theta activity during PS. FRA expression affected also at the reentry pontine and diencephalic structures, such as the lateral parabrachial nucleus and the central nucleus of the amygdala, which are known to contribute to the occurrence of pontine waves and the related bursts of REM. 5. Observations made on the various components of the vestibular complex indicated that no Fos and FRA expression occurred in the LVN at the four different mission time points. However, an increase in Fos and FRA expression occurred particularly in the medial (MVN) and spinal vestibular nuclei (SpVN) at FD2 and at R + 1, i.e. 1 day after launch and 12-24 hours after landing, respectively. The pattern of FRA expression observed in the VN during the space flight was generally similar to that of Fos, except at the reentry, when FRA positive cells were observed throughout the whole SpVN, but not the MVN, which showed only a few labeled cells in its rostral part. In contrast to this finding, a prominent Fos expression was found not only in the SpVN, but also throughout the entire MVN. In this case the Fos labeling affected not only the caudal but also the rostral part of this structure, including the dorsal (MVePc) rather than the ventral aspect (MVeMc). Grounded on their different time of persistence, both Fos and FRA expression which occurred in the SpVe could be attributed to the increase in gravity force experienced during take-off and landing, while the Fos pattern which affected particularly the MVN soon after the reentry could additionally be attributed to the rebound episode of PS following the forced period of waking which occurred after landing and after the prolonged (12 days) exposure to microgravity. 6. The results of the present experiments provide the first molecular evidence that pontine activity sources producing rhythmic discharges of vestibulo-ocular neurons during REM sleep may substitute for labyrinthine signals after prolonged (12 days) exposure to microgravity, thus contributing to activity-related plastic changes in the VN leading to readaptation of the vestibular system to 1 G.     

Differential effects of water and saline intake on water deprivation-induced c-Fos staining in the rat

We studied c-Fos staining in adult male rats after 48 h of water deprivation and after 46 h of water deprivation with 2 h of access to water or physiological saline. Controls were allowed ad libitum access to water and physiological saline. For immunocytochemistry, anesthetized rats were perfused with a commercially available antibody for c-Fos. Dehydration significantly increased plasma vasopressin (AVP), osmolality, plasma renin activity (PRA), hematocrit, and sodium concentration and decreased urinary volume. Fos staining was significantly increased in the median preoptic nucleus, organum vasculosum of the lamina terminalis, supraoptic nucleus (SON), and magnocellular and parvocellular paraventricular nucleus (PVN), as well as the area postrema, nucleus of the solitary tract (NTS), and rostral ventrolateral medulla (RVL). Rehydration with water significantly decreased AVP levels and Fos staining in the SON, PVN, and RVL and significantly increased Fos expression in the perinuclear zone of the SON, NTS, and parabrachial nucleus. Rehydration with water was associated with decreased urinary sodium concentration and hypotonicity, and hematocrit and PRA were comparable to levels seen after dehydration. After rehydration with saline, plasma osmolality, hematocrit, and PRA were not different from control, but plasma AVP and urinary sodium concentration were increased. In the SON, Fos staining was significantly increased, with a great percentage of the Fos cells also stained for oxytocin compared with water deprivation. Changes in Fos staining were also observed in the NTS, RVL, parabrachial nucleus, and PVN. Rehydration with water or saline produces differential effects on plasma AVP, Fos staining, and sodium concentration.     

前列腺素D2与睡眠调节

前列腺素D2（PGD2）是目前已知最有潜力的内源性促睡眠物质之一。鼠脑脊液（CSF）中PGD2浓度与睡眠－觉醒周期一致,呈现节律性改变,并且随睡眠剥夺期间嗜睡倾向增加而增加,催化PGH2转变为PGD2的特生酶有两种：Lipocalin型前列腺素D合成酶（L－PGDS）和脾型PGDS。L－PGDS主要在大脑蛛网膜和脉络丛产生,并分泌入CSF。L－PGDS和PGD2在脑室系统、蛛网膜下腔及细胞外间隙中循环,循环中的PGD2可与前脑头端基底腹内侧面的化学感受器中的前列腺素D2受体（DPR）相互作用,通过活化具有腺苷A2a受体的元以产生促进睡眠的信号。PGD2敏感区内DPR的活化可导致腹侧视前区（VLPO）内神经元的活化,其可通过抑制结节乳头核（TMN）而促进睡眠。相反,PGE2在维持大脑清醒状态下起主要作用。PGD2和PGE2的平衡对正常睡眠－觉醒周期的维持十分关键。     

Microinjection of melanin concentrating hormone into the lateral preoptic area promotes non-REM sleep in the rat

The ventrolateral preoptic area (VLPO) has been recognized as one of the key structures responsible for the generation of non-REM (NREM) sleep. The melanin-concentrating hormone (MCH)-containing neurons, which are located in the lateral hypothalamus and incerto-hypothalamic area, project widely throughout the central nervous system and include projections to the VLPO. The MCH has been associated with the central regulation of feeding and energy homeostasis. In addition, recent findings strongly suggest that the MCHergic system promotes sleep. The aim of the present study was to determine if MCH generates sleep by regulating VLPO neuronal activity. To this purpose, we characterized the effect of unilateral and bilateral microinjections of MCH into the VLPO on sleep and wakefulness in the rat. Unilateral administration of MCH into the VLPO and adjacent dorsal preoptic area did not modify sleep. On the contrary, bilateral microinjections of MCH (100 ng) into these areas significantly increased light sleep (LS, 39.2 ± 4.8 vs. 21.6 ± 2.5 min, P < 0.05) and total NREM sleep (142.4 ± 23.2 vs. 86.5 ± 10.5 min, P < 0.05) compared to control (saline) microinjections. No effect was observed on REM sleep. We conclude that MCH administration into the VLPO and adjacent dorsal lateral preoptic area promotes the generation of NREM sleep.     

Key roles of the histaminergic system in sleep-wake regulation

Histamine plays an important role in mediating wakefulness in mammals. Based on the findings from gene-manipulated mice, we provide several lines of evidence showing the roles of the histaminergic system in the somnogenic effects of prostaglandin (PG) D2 and adenosine, and in the arousal effects of PGE2 and orexin. PGD2 activates DP1 receptors (R) to promote sleep by stimulating them to release adenosine. The released adenosine activates adenosine A2AR and subsequently excites the ventrolateral preoptic area (VLPO), one of the sleep centers in the anterior hypothalamus. VLPO neurons then send inhibitory signals to downregulate the histaminergic tuberomammillary nucleus (TMN), which contributes to arousal. A1R is expressed in histaminergic neurons of the rat TMN. Adenosine in the TMN inhibits the histaminergic system via A1R and promotes non–rapid eye movement sleep. Conversely, both endogenous PGE2 and orexin activate the histaminergic system through EP4R and OX-2R, respectively, to promote wakefulness via histamine H1R. Furthermore, the arousal effect of ciproxifan, H3R antagonist, depends on the activation of histaminergic systems. These findings indicate that VLPO and TMN regulate sleep and wakefulness by means of a “flip-flop” mechanism operating in an anti-coincident manner during sleep–wake state transitions.