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Sleep disruption (SD) promotes stress which may mediate the effect of SD induced by noise on bodyweight gain and food intake. We determined if the change in bodyweight during SD caused by noise was driven by stress (assessed by corticosterone) and whether the effects of noise on SD, stress and bodyweight were specific to the method of SD or a consequence of SD per se. We isolated stress from SD due to noise by exposing rats to noise during the dark

Background/objectives: Inadequate sleep increases obesity and environmental noise contributes to poor sleep. However, women may be more vulnerable to noise and hence more susceptible to sleep disruption-induced weight gain than men. In male rats, exposure to environmental (i.e. ambient) noise disrupts sleep and increases feeding and weight gain. However, the effects of environmental noise on sleep and weight gain in female rats are unknown. Thus, this study was designed to determine whether noise exposure would disturb sleep, increase feeding and weight gain and alter the length of the estrous cycle in female rats. Subjects/methods: Female rats (12-weeks old) were exposed to noise for 17d (8h/d during the light period) to determine the effects of noise on weight gain and food intake. In a separate set of females, estrous cycle phase and length, EEG, EMG, spontaneous physical activity and energy expenditure were recorded continuously for 27d during baseline (control, 9d), noise exposure (8h/d, 9d) and recovery (9d) from sleep disruption. Results: Noise exposure significantly increased weight gain and food intake compared to females that slept undisturbed. Noise also significantly increased wakefulness, reduced sleep and resulted in rebound sleep during the recovery period. Total energy expenditure was significantly lower during both noise exposure and recovery due to lower energy expenditure during spontaneous physical activity and sleep. Notably, noise did not alter the estrous cycle length. Conclusions: As previously observed in male rats, noise exposure disrupted sleep and increased weight gain in females but did not alter the length of the estrous cycle. This is the first demonstration of weight gain in female rats during sleep disruption. We conclude that the sleep disruption caused by exposure to environmental noise is a significant tool for determining how sleep loss contributes to obesity in females.

BACKGROUND: Identifying whether components of total energy expenditure (EE) are affected by orexin receptor (OXR1 and OXR2) stimulation or antagonism with dual orexin receptor antagonists (DORAs) has relevance for obesity treatment. Orexin receptor stimulation reduces weight gain by increasing total EE and EE during spontaneous physical activity (SPA). OBJECTIVE: The purpose of this study was to determine if a DORA (TCS-1102) in the ventrolateral preoptic area (VLPO) reduced orexin-A-induced arousal, SPA, total EE and EE during sleep, rest, wake and SPA and whether the DORA alone reduced total EE and its components. We hypothesized that: (1) a DORA would reduce orexin-A induced increases in arousal, SPA, components of total EE, reductions in sleep and the EE during sleep and (2) the DORA alone would reduce baseline (non-stimulated) SPA and total EE. SUBJECTS/METHODS: Sleep, wakefulness, SPA and EE were determined after microinjection of the DORA (TCS-1102) and orexin-A in the VLPO of male Sprague–Dawley rats with a unilateral cannula targeted towards the VLPO. Individual components of total EE were determined based on time-stamped data. RESULTS: The DORA reduced orexin-A-induced increases in arousal, SPA, total EE and EE during SPA, wake, rest and sleep 1 h post injection (P < 0.05). Orexin-A significantly reduced sleep and significantly increased EE during sleep 1 h post injection (P < 0.05). Furthermore, the DORA alone significantly reduced total EE, EE during sleep (NREM and REM) and resting EE 2 h post injection (P < 0.05). CONCLUSIONS: These data suggest that orexin-A reduces weight gain by stimulating total EE through increases in EE during SPA, rest and sleep. Residual effects of the DORA alone include decreases in total EE and EE during sleep and rest, which may promote weight gain.

Neuropeptide B and W (NPB and NPW) are cognate peptide ligands for NPBWR1 (GPR7), a G protein-coupled receptor. In rodents, they have been implicated in the regulation of energy homeostasis, neuroendocrine/autonomic responses, and social interactions. Although localization of these peptides and their receptors in adult rodent brain has been well documented, their expression in mouse brain during development is unknown. Here we demonstrate the transient expression of NPW mRNA in the dorsomedial hypothalamus (DMH) of postnatal mouse brain and its co-localization with neuropeptide Y (NPY) mRNA. Neurons expressing both NPW and NPY mRNAs begin to emerge in the DMH at about postnatal day 0 (P-0) through P-3. Their expression is highest around P-14, declines after P-21, and by P-28 only a faint expression of NPW and NPY mRNA remains. In P-18 brains, we detected NPW neurons in the region spanning the subincertal nucleus (SubI), the lateral hypothalamic (LH) perifornical (PF) areas, and the DMH, where the highest expression of NPW mRNA was observed. The majority of these postnatal hypothalamic NPW neurons co-express NPY mRNA. A cross of NPW-iCre knock-in mice with a Cre-dependent tdTomato reporter line revealed that more than half of the reporter-positive neurons in the adult DMH, which mature from the transiently NPW-expressing neurons, are sensitive to peripherally administrated leptin. These data suggest that the DMH neurons that transiently co-express NPW and NPY in the peri-weaning period might play a role in regulating energy homeostasis during postnatal development.

Neuropeptide B (NPB) and neuropeptide W (NPW) are endogenous neuropeptide ligands for the G protein-coupled receptors NPBWR1 and NPBWR2. Here we report that the majority of NPW neurons in the mesolimbic region possess tyrosine hydroxylase immunoreactivity, indicating that a small subset of dopaminergic neurons coexpress NPW. These NPW-containing neurons densely and exclusively innervate two limbic system nuclei in adult mouse brain: the lateral bed nucleus of the stria terminalis and the lateral part of the central amygdala nucleus (CeAL). In the CeAL of wild-type mice, restraint stress resulted in an inhibition of cellular activity, but this stress-induced inhibition was attenuated in the CeAL neurons of NPW(-/-) mice. Moreover, the response of NPW(-/-) mice to either formalin-induced pain stimuli or a live rat (i.e., a potential predator) was abnormal only when they were placed in a novel environment: The mice failed to show the normal species-specific self-protective and aversive reactions. In contrast, the behavior of NPW(-/-) mice in a habituated environment was indistinguishable from that of wild-type mice. These results indicate that the NPW/NPBWR1 system could play a critical role in the gating of stressful stimuli during exposure to novel environments.

In humans, narcolepsy is a debilitating and important sleep disorder that was first described in 1877 by Westphal, a German physician, based on his observation of a patient with sudden sleep attacks associated with symptoms of motor incapacity and aphasia. He considered it a form of epilepsy. Three years later, the French neurologist Gelineau (Fig. 24.1) described the disorder for the first time as a distinct clinical entity rather than a symptom of another condition, and proposed the term narcolepsy, meaning “sleep seizure”. In the 1880 paper, Gelineau described a patient who, from the age of 36, began to fall asleep suddenly during the day and had sudden falls or “astasia” (cataplexy), which occurred whenever he became emotional.

Adult neurogenesis persists throughout life in restricted brain regions in mammals and is affected by various physiological and pathological conditions. The tumor suppressor genePtenis involved in adult neurogenesis and is mutated in a subset of autism patients with macrocephaly; however, the link between the role of PTEN in adult neurogenesis and the etiology of autism has not been studied before. Moreover, the role of hippocampus, one of the brain regions where adult neurogenesis occurs, in development of autism is not clear. Here, we show that ablatingPtenin adult neural stem cells in the subgranular zone of hippocampal dentate gyrus results in higher proliferation rate and accelerated differentiation of the stem/progenitor cells, leading to depletion of the neural stem cell pool and increased differentiation toward the astrocytic lineage at later stages.Pten-deleted stem/progenitor cells develop into hypertrophied neurons with abnormal polarity. Additionally,Ptenmutant mice have macrocephaly and exhibit impairment in social interactions and seizure activity. Our data reveal a novel function for PTEN in adult hippocampal neurogenesis and indicate a role in the pathogenesis of abnormal social behaviors.

Summary In the decade since the discovery that pathology of the orexin/hypocretin system is causative for the sleep disorder narcolepsy, considerable progress has been made in understanding the functional role of the neuropeptide. Two, apparently separate functions of orexin have emerged as a consensus from studies to date. The first is the effect on vigilance state boundaries, as exemplified by narcolepsy. Thus the absence of orexin severely limits the ability to maintain prolonged periods of wakefulness or sleep and also allows the unregulated appearance of cataplexy as sudden muscle weakness during wakefulness. The second function is that orexin acts as a signaling molecule in transferring information about physiological disequilibrium to the central nervous system. Orexin activates the central arousal and motor systems during such disequilibrium and so may facilitate the necessary response and adaptation to restore equilibrium. A feasible relationship between these two functions is therefore that the maintenance of prolonged and active wakefulness is an integral part of this adaptive process. Furthermore, the limit placed on the onset of sleep by orexin suggests that these adaptive processes then continue during sleep to become integrated into the development of a coping strategy for the longer term.

金沢大学医薬保健研究域医学系, Orexin-A and orexin-B are hypothalamic neuropeptides that play critical roles in the maintenance of wakefulness. Intracerebroventricular (ICV) administration of orexin-A has been shown to promote wakefulness and suppress both rapid eye movement (REM) sleep and non-REM (NREM) sleep through the orexin receptor-1 (OX1R) and orexin receptor-2 (OX2R). Here, we elucidated the differential roles of orexin receptors in the regulation of sleep and wakefulness by comparing the effects of ICV orexin-A administration in wild-type, OX1R, and OX2R mice. The effects of orexin-A on wakefulness and NREM sleep were significantly attenuated in both knock-out mice as compared with wild-type mice, with substantially larger attenuation in OX2R mice than in OX1R mice. These results suggest that although the OX2R-mediated pathway has a pivotal role in the promotion of wakefulness, OX1R also plays additional roles in promoting arousal. In contrast, suppression of REM sleep by orexin-A administration was slightly and similarly attenuated in both OX1R and OX2R mice, suggesting a comparable contribution of the two receptors to REM sleep suppression. Histological studies demonstrated differential distributions of each receptor subtype in distinct neuronal populations with specific neurotransmitter identities in brainstem cholinergic/monoaminergic neurons. In the laterodorsal tegmental and pedunculopontine tegmental nuclei especially, cholinergic neurons exclusively expressed OX1R mRNA, but OX2R mRNA was expressed mainly in GABAergic putative interneurons. Thus, each orexin receptor subtype plays differential roles in gating NREM and REM sleep through distinct neuronal pathways. © 2011 by the authors.

PTEN (phosphatase and tensin homolog deleted on chromosome ten) is a lipid phosphatase that counteracts the function of phosphatidylinositol-3 kinase (PI3K). Loss of function of PTEN results in constitutive activation of AKT and downstream effectors and correlates with many human cancers, as well as various brain disorders, including macrocephaly, seizures, Lhermitte–Duclos disease, and autism. We previously generated a conditionalPtenknock-out mouse line withPtenloss in limited postmitotic neurons in the cortex and hippocampus.Pten-null neurons developed neuronal hypertrophy and loss of neuronal polarity. The mutant mice exhibited macrocephaly and behavioral abnormalities reminiscent of certain features of human autism. Here, we report that rapamycin, a specific inhibitor of mammalian target of rapamycin complex 1 (mTORC1), can prevent and reverse neuronal hypertrophy, resulting in the amelioration of a subset of PTEN-associated abnormal behaviors, providing evidence that the mTORC1 pathway downstream of PTEN is critical for this complex phenotype.

The conceptual understanding of hippocampal function has been challenged recently by the finding that new granule cells are born throughout life in the mammalian dentate gyrus (DG). The number of newborn neurons is dynamically regulated by a variety of factors. Kainic acid-induced seizures, a rodent model of human temporal lobe epilepsy, strongly induce the proliferation of DG neurogenic progenitor cells and are also associated with long-term cognitive impairment. We show here that the antiepileptic drug valproic acid (VPA) potently blocked seizure-induced neurogenesis, an effect that appeared to be mainly mediated by inhibiting histone deacetylases (HDAC) and normalizing HDAC-dependent gene expression within the epileptic dentate area. Strikingly, the inhibition of aberrant neurogenesis protected the animals from seizure-induced cognitive impairment in a hippocampus-dependent learning task. We propose that seizure-generated granule cells have the potential to interfere with hippocampal function and contribute to cognitive impairment caused by epileptic activity within the hippocampal circuitry. Furthermore, our data indicate that the effectiveness of VPA as an antiepileptic drug may be partially explained by the HDAC-dependent inhibition of aberrant neurogenesis induced by seizure activity within the adult hippocampus.

Study objectives The ventrolateral preoptic area (VLPO) and the orexin/hypocretin neuronal system are key regulators of sleep onset, transitions between vigilance states, and energy homeostasis. Reciprocal projections exist between the VLPO and orexin/hypocretin neurons. Although the importance of the VLPO to sleep regulation is clear, it is unknown whether VLPO neurons are involved in energy balance. The purpose of these studies was to determine if the VLPO is a site of action for orexin-A, and which orexin receptor subtype(s) would mediate these effects of orexin-A. We hypothesized that orexin-A in the VLPO modulates behaviors (sleep and wakefulness, feeding, spontaneous physical activity [SPA]) to increase energy expenditure. Design and measurements Sleep, wakefulness, SPA, feeding, and energy expenditure were determined after orexin-A microinjection in the VLPO of male Sprague-Dawley rats with unilateral cannulae targeting the VLPO. We also tested whether pretreatment with a dual orexin receptor antagonist (DORA, TCS-1102) or an OX2R antagonist (JNJ-10397049) blocked the effects of orexin-A on the sleep/wake cycle or SPA, respectively. Results Orexin-A injected into the VLPO significantly increased wakefulness, SPA, and energy expenditure (SPA-induced and total) and reduced NREM sleep and REM sleep with no effect on food intake. Pretreatment with DORA blocked the increase in wakefulness and the reduction in NREM sleep elicited by orexin-A, and the OX2R antagonist reduced SPA stimulated by orexin-A. Conclusions These data show the ventrolateral preoptic area is a site of action for orexin-A, which may promote negative energy balance by modulating sleep/wakefulness and stimulating spontaneous physical activity and energy expenditure.

Neuropeptide B (NPB) and neuropeptide W (NPW) have been recently identified as ligands for the G protein-coupled receptor (GPR) 7 and GPR8. The precise in vivo role of this neuropeptide-receptor pathway has not been fully demonstrated. In this paper, we report that NPB-deficient mice manifest a mild adult-onset obesity, similar to that reported in GPR7-null mice. NPB-deficient mice also exhibit hyperalgesia in response to inflammatory pain. Hyperalgesia was not observed in response to chemical pain, thermal pain, or electrical stimulation. NPB-deficient mice demonstrated intact behavioral responses to pain, and learning from the negative reinforcement of electrical stimulation was unaltered. Baseline anxiety was also unchanged as measured in both the elevated plus maze and time spent immobile in a novel environment. These data support the idea that NPB is a factor in the modulation of responses to inflammatory pain and body weight homeostasis.

Narcolepsy-cataplexy, a neurological disorder associated with the absence of hypothalamic orexin (hypocretin) neuropeptides, consists of two underlying problems: inability to maintain wakefulness and intrusion of rapid eye movement (REM) sleep into wakefulness. Here we document, using behavioral, electrophysiological, and pharmacological criteria, two distinct classes of behavioral arrests exhibited by mice deficient in orexin-mediated signaling. Both OX2R(-/-) and orexin(-/-) mice are similarly affected with behaviorally abnormal attacks of non-REM sleep ("sleep attacks") and show similar degrees of disrupted wakefulness. In contrast, OX2R(-/-) mice are only mildly affected with cataplexy-like attacks of REM sleep, whereas orexin(-/-) mice are severely affected. Absence of OX2Rs eliminates orexin-evoked excitation of histaminergic neurons in the hypothalamus, which gate non-REM sleep onset. While normal regulation of wake/non-REM sleep transitions depends critically upon OX2R activation, the profound dysregulation of REM sleep control unique to the narcolepsy-cataplexy syndrome emerges from loss of signaling through both OX2R-dependent and OX2R-independent pathways.

During rapid eye movement (REM) sleep, activity of non-retinal origin is propagated into central visual-system pathways in a manner similar, in pattern and intensity, to central visual-system activity that is exogenously generated in waking. It has been hypothesized that REM sleep, which is more abundantly represented early in life than later, functions to provide adjunct 'afferent' input for shaping synaptic connectivity during brain maturation. Here we present data that support this proposal. Recent studies have described a developmentally regulated form of in vitro long-term potentiation (LTP) in the visual cortex that is experience- and age-dependent. In immature rats, suppression of retinal activation of the visual system by removal of visual experience (dark rearing) extends the age when the developmentally regulated form of LTP can be produced. This study tests whether suppression of REM-state activation of the visual system also lengthens the developmental period in which this specific form of LTP can be elicited. Young rats were deprived of REM sleep by the multiple-small-platforms-over-water method during the typically latest week for induction of such LTP in slices of visual cortex. After this week, we could still induce LTP in slices from nearly all the REM-sleep-deprived rats (8/9) but not from age-matched rats that had not lost REM sleep (0/5). The control rats had been housed on large platforms that allow the animals to obtain REM sleep. Only body weights and the concentration of thyrotrophin-releasing hormone in the hypothalamus distinguished home-caged, normal-sleeping controls from both groups of platform animals. On all measures, stress levels were not dissimilar in the two platforms groups. After 7 days of behavioral suppression of REM sleep in immature rats, and consequent reduction of the intense, extra-retinal activity endogenously generated during this sleep state, we found that the period was extended in which developmentally regulated synaptic plasticity (LTP) could be elicited in slices of visual neocortex. These studies support the role of REM sleep and its associated neuronal activity in brain maturation.

Orexins are bioactive peptides, which have been shown to play a pivotal role in vigilance state transitions: the loss of orexin-producing neurons (orexin neurons) leads to narcolepsy with cataplexy in the human. However, the effect of the need for sleep (i.e., sleep pressure) on orexin neurons remains largely unknown. Here, we found that immunostaining intensities of the α1 subunit of the GABAA receptor and neuroligin 2, which is involved in inhibitory synapse specialization, on orexin neurons of mouse brain were significantly increased by 6-h sleep deprivation. In contrast, we noted that immunostaining intensities of the α2, γ2, and β2/3 subunits of the GABAA receptor and Huntingtin-associated protein 1, which is involved in GABAAR trafficking, were not changed by 6-h sleep deprivation. Using a slice patch recording, orexin neurons demonstrated increased sensitivity to a GABAA receptor agonist together with synaptic plasticity changes after sleep deprivation when compared with an ad lib sleep condition. In summary, the GABAergic input property of orexin neurons responds rapidly to sleep deprivation. This molecular response of orexin neurons may thus play a role in the changes that accompany the need for sleep following prolonged wakefulness, in particular the decreased probability of a transition to wakefulness once recovery sleep has begun.

Pyridostigmine bromide (PB), a reversible inhibitor of acetylcholinesterase (AChE), is used for the treatment of myasthenia gravis. PB has also been provided to military personnel for preexposure protection against potential soman release. The entry of PB into the brain is typically minimal, but recently published data in mice suggest that a brief forced swim stress increases the permeability of the blood-brain barrier to PB. From these results, PB administered under stressful conditions was proposed to induce long-lasting central cholinergic deficits, potentially explaining the neurological and neuropsychological symptoms presented by some Gulf War veterans. In undertaking to replicate these results in the Long-Evans rat, no evidence of a stress-potentiated central effect of PB, administered at doses up 5.0 mg/kg ip, was found. Three stress protocols were used: restraint, forced swim, or a combined restraint/forced swim. Wistar rats were also tested in some of the protocols to ensure that the results were generalizable across rat strains, and plasma corticosterone levels were measured to test the effectiveness of the stressors employed. In contrast to the previously reported findings in the mouse, stress significantly reduced the entry of PB into rat brain, as measured by reduced inhibition of AChE activity: a 12.5% reduction in whole brain AChE activity after treatment with 5.0 mg/kg PB under control conditions declined to 9% after stress exposure. It is apparent, therefore, that the interaction between stress and PB requires further study, and previous data should be reassessed before they are used as a basis for interpreting symptoms presented by veterans.

Neurochemical and functional abnormalities of the striatum have been reported in schizophrenic brains, but the cellular substrates of these changes are not known. We hypothesized that schizophrenia may involve an abnormality in one of the key modulators of striatal output, the cholinergic interneuron. We measured the densities of cholinergic neurons in the striatum in schizophrenic and control brains in a blind analysis, using as a marker of this cell population immunoreactivity for choline acetyltransferase, the synthetic enzyme of acetylcholine. As an independent marker, we used immunoreactivity for calretinin, a protein which is co-localized with choline acetyltransferase in virtually all of the cholinergic interneurons of the striatum. A significant decrease in choline acetyltransferase-positive and calretinin-positive cell densities was found in the schizophrenic cases compared with controls in the striatum as a whole [for the choline acetyltransferase-positive cells: controls: 3.21 +/- 0.48 cells/mm2 (mean +/- S.D.), schizophrenics: 2.43 +/- 0.68 cells(mm2; P < 0.02]. The decrease was patchy in nature and most prominent in the ventral striatum (for the choline acetyltransferase-positive cells: controls: 3.47 +/- 0.59 cells/mm2, schizophrenics: 2.52 +/- 0.64 cells/ mm2; P < 0.005) which included the ventral caudate nucleus and nucleus accumbens region. Three of the schizophrenic cases with the lowest densities of cholinergic neurons had not been treated with neuroleptics for periods from more than a month to more than 20 years. A decrease in the number or function of the cholinergic interneurons of the striatum may disrupt activity in the ventral striatal-pallidal-thalamic-prefrontal cortex pathway and thereby contribute to abnormalities in function of the prefrontal cortex in schizophrenia.

Neurons containing the neuropeptide orexin (hypocretin) are located exclusively in the lateral hypothalamus and send axons to numerous regions throughout the central nervous system, including the major nuclei implicated in sleep regulation. Here, we report that, by behavioral and electroencephalographic criteria, orexin knockout mice exhibit a phenotype strikingly similar to human narcolepsy patients, as well as canarc-1 mutant dogs, the only known monogenic model of narcolepsy. Moreover, modafinil, an anti-narcoleptic drug with ill-defined mechanisms of action, activates orexin-containing neurons. We propose that orexin regulates sleep/wakefulness states, and that orexin knockout mice are a model of human narcolepsy, a disorder characterized primarily by rapid eye movement (REM) sleep dysregulation.

This study examined whether damage to dopamine (DA) nerve terminals via inhibition of energy metabolism in the striatum would result in the retrograde loss of cell bodies in the substantia nigra. Infusion of 2 micromol malonate into the left striatum of rats resulted in a 67% loss of striatal DA and a 40% loss of tyrosine hydroxylase (TH)-positive neurons in the substantia nigra. No change in the number of Nissl-positive-TH-negative neurons was observed. These findings demonstrate the retrograde destruction of DA cell bodies in the substantia nigra resulting from energy impairment at their terminal projection site.

Sleep is regulated by homeostatic mechanisms, and the low-frequency power in the electroencephalogram (delta power) during non-rapid eye movement sleep reflects homeostatic sleep need. Additionally, sleep is limited by circadian and environmentally influenced arousal. Little is known, however, about the underlying neural substrates for sleep homeostasis and arousal and about the potential link between them. Here, we subjected C57BL/6 mice to 6 h of sleep deprivation using two different methods: gentle handling and continual cage change. Both groups were deprived of sleep to a similar extent (>99%), and, as expected, the delta power increase during recovery sleep was quantitatively similar in both groups. However, in a multiple sleep latency test, the cage change group showed significantly longer sleep latencies than the gentle handling group, indicating that the cage change group had a higher level of arousal despite the similar sleep loss. To investigate the possible biochemical correlates of these behavioral changes, we screened for arousal-related and sleep need-related phosphoprotein markers from the diencephalon. We found that the abundance of highly phosphorylated forms of dynamin 1, a presynaptic neuronal protein, was associated with sleep latency in the multiple sleep latency test. In contrast, the abundance of highly phosphorylated forms of N -myc downstream regulated gene 2, a glial protein, was increased in parallel with delta power. The changes of these protein species disappeared after 2 h of recovery sleep. These results suggest that homeostatic sleep need and arousal can be dissociated behaviorally and biochemically and that phosphorylated N -myc downstream regulated gene 2 and dynamin 1 may serve as markers of homeostatic sleep need and arousal, respectively.

The calcium-binding protein calbindin-D28k (CB) is located in midbrain dopaminergic (DA) neurons that are less vulnerable to degeneration in Parkinson's disease and in an animal model of the disorder, the MPTP-treated monkey. The present study sought to determine whether CB-containing DA neurons are also less vulnerable to degeneration in the MPTP-treated mouse. Double-labelling immunocytochemical staining and computer imaging techniques were employed to map and quantify the tyrosine hydroxylase-, CB- and CB-containing tyrosine hydroxylase neurons in portions of nucleus A9 and nucleus A10 (ventral tegmental area and central linear nucleus) following MPTP treatment in the C57BL/6 mouse. A cumulative dose of 140 mg/kg MPTP produced a significantly greater loss of DA neurons that lack CB in both nucleus A9 (71 +/- 4%) and the ventral tegmental area (70 +/- 4%), compared to the loss of DA neurons that contain CB (44 +/- 6% and 25 +/- 14%, respectively). In the central linear nucleus there was no loss of CB-containing DA neurons. These data demonstrate that the presence of CB in midbrain DA neurons identifies a population of cells in the mouse that are less vulnerable to MPTP-induced degeneration. The mouse, therefore, can serve as a useful model in which to investigate the putative neuroprotective effects of CB in an animal model of Parkinson's disease.

The neurotoxin MPTP has been used to create an animal model of Parkinson's disease in the mouse, in part, because it causes a significant loss of dopaminergic neurons in the substantia nigra (nucleus A9). The purpose of the present study was to determine whether MPTP also causes degeneration of midbrain dopaminergic neurons in nuclei A8 and A10 in the mouse, as occurs in humans with Parkinson's disease. Two commonly used strains of mice were used: FVB/N and C57BL/6. MPTP was administered in cumulative doses of 50—300 mg/kg. Seven days later, dopamine concentrations were measured in the striatum using high performance liquid chromatography, and midbrain dopaminergic neurons were identified using an antibody against tyrosine hydroxylase. The cell locations were mapped with a computer imaging system. In the FVB/N strain, there was a dose-dependent decrease in striatal dopamine concentrations. Although the highest dose (300 mg/kg) caused an 86% reduction in striatal dopamine concentrations, there was only a moderate and non-significant loss of midbrain dopaminergic neurons. In the C57BL/6 strain, however, a high dose of MPTP (240 mg/kg) caused a significant reduction in both striatal dopamine concentrations (95%), and midbrain dopaminergic cells; 69% loss of nucleus A8 cells, 75% loss of nucleus A9 cells, and in nucleus A10 subnuclei there was 42% loss of ventral tegmental area cells, 55% loss of interfascicular nucleus cells, and no loss of cells in the central linear nucleus. These data (1) provide further evidence for differential susceptibility to MPTP toxicity among different mouse strains, (2) indicate that a significant depletion of striatal dopamine is not necessarily due to degeneration of midbrain dopaminergic neurons, (3) provide the precise locations of midbrain dopaminergic cells that are vulnerable to MPTP, which will aid future studies that seek to determine the mechanism/s by which MPTP selectively destroys only certain midbrain dopaminergic neurons, and (4) indicate that MPTP produces midbrain dopaminergic neuronal degeneration in the same nuclei in the C57BL16 mouse that degenerate in humans with Parkinson's disease..

The dopaminergic (DA) neurons in the midbrain play a role in cognition, affect and movement. The purpose of the present study was to map and quantify the number of DA neurons in the midbrain, within the nuclei that constitute cell groups A8, A9 and A10, in the mouse. Two strains of mice were used; the C57BL/6 strain was chosen because it is commonly used in neurobiological studies, and the FVB/N strain was chosen because it is used frequently in transgenic studies. DA neurons were identified, in every fifth 20-microns-thick coronal section, using an antibody against tyrosine hydroxylase. Cell locations were entered into a computer imaging system. The FVB/N strain has 42% more midbrain DA neurons than the C57BL/6 strain; on one side of the brain there were 15,135 +/- 356 neurons (mean +/- S.E.M.) in the FVB/N strain, and 10,645 +/- 315 neurons in the C57BL/6 strain. In both strains, approximately 11% of the neurons were located in nucleus A8 (the DA neurons in the retrorubral field), 38% in nucleus A9 (the DA neurons in the substantia nigra pars compacta, pars reticulata, and pars lateralis), and 51% in nucleus A10 (the DA neurons in midline regions such as the ventral tegmental area, central linear nucleus, and interfascicular nucleus). The number of midbrain DA cells, and their distribution within the three nuclear groups, is discussed with respect to findings in other species.

Sleep and wakefulness are regulated primarily by inhibitory interactions between the hypothalamus and brainstem. The expression of the states of rapid eye movement (REM) sleep and non-REM (NREM) sleep also are correlated with the activity of groups of REM-off and REM-on neurons in the dorsal brainstem. However, the contribution of ventral brainstem nuclei to sleep regulation has been little characterized to date. Here we examined sleep and wakefulness in mice deficient in a homeobox transcription factor, Goosecoid-like ( Gscl ), which is one of the genes deleted in DiGeorge syndrome or 22q11 deletion syndrome. The expression of Gscl is restricted to the interpeduncular nucleus (IP) in the ventral region of the midbrain–hindbrain transition. The IP has reciprocal connections with several cell groups implicated in sleep/wakefulness regulation. Although Gscl −/− mice have apparently normal anatomy and connections of the IP, they exhibited a reduced total time spent in REM sleep and fewer REM sleep episodes. In addition, Gscl −/− mice showed reduced theta power during REM sleep and increased arousability during REM sleep. Gscl −/− mice also lacked the expression of DiGeorge syndrome critical region 14 ( Dgcr14 ) in the IP. These results indicate that the absence of Gscl and Dgcr14 in the IP results in altered regulation of REM sleep.

The complexity of the pontine neuronal mechanisms that control and modulate expression of rapid eye movement (REM) sleep has been only recently recognized. In this review of some of these mechanisms, results from studies in the cat and rat are summarized and compared, and the differences between neuronal populations that might be effective, modulatory, or executive for REM sleep control are considered. One of the principal pontine areas that has been found to be important for the induction of REM sleep is located in the nucleus pontis oralis. Another is located in a more dorsocaudal area. The latter, designated the peri-locus coeruleus α in the cat, corresponds most closely with the sublaterodorsal (SLD) nucleus in the rat. In both areas in the cat, cholinergic mechanisms are important for REM sleep induction, whereas in the rat, only the pontis oralis area is cholinergically sensitive. In contrast, the SLD is responsive to glutamatergic processes. The emerging recognition of the importance of GABAergic modulation of these sites in both species is reviewed, and from this work, some preliminary conclusions can be drawn. In the caudal pontis oralis of rat, GABAergic mechanisms can affect the release of acetylcholine, inferring that transmitter release in a terminal region can be decoupled from impulse flow. This is important in light of the concept of changes in the discharge rate of reciprocally interacting populations of neurons being correlated with differences in vigilance state. A second conclusion, based on the work in the SLD, is that network interactions are critical for REM sleep control. The latter can be extended to conclude that activation of neurons that are effective for at least two of the signs of REM sleep may be sufficient to induce the state. Distributed networks would obviate the need for a single locus of executive control for REM sleep. But another possibility derived from the SLD studies is that executive control of REM sleep may actually represent the coordination of effective mechanisms, explaining the difficulty that has been encountered in finding a REM sleep “generator.”

Cyclin-dependent kinase 5 (Cdk5) is a proline-directed serine/threonine kinase that has been implicated in learning, synaptic plasticity, neurotransmission, and numerous neurological disorders. We previously showed that conditional loss of Cdk5 in adult mice enhanced hippocampal learning and plasticity via modulation of calpain-mediated N-methyl-D-aspartic acid receptor (NMDAR) degradation. In the present study, we characterize the enhanced synaptic plasticity and examine the effects of long-term Cdk5 loss on hippocampal excitability in adult mice. Field excitatory post-synaptic potentials (fEPSPs) from the Schaffer collateral CA1 subregion of the hippocampus (SC/CA1) reveal that loss of Cdk5 altered theta burst topography and enhanced post-tetanic potentiation. Since Cdk5 governs NMDAR NR2B subunit levels, we investigated the effects of long-term Cdk5 knockout on hippocampal neuronal excitability by measuring NMDAR-mediated fEPSP magnitudes and population-spike thresholds. Long-term loss of Cdk5 led to increased Mg(2+)-sensitive potentials and a lower threshold for epileptiform activity and seizures. Biochemical analyses were performed to better understand the role of Cdk5 in seizures. Induced-seizures in wild-type animals led to elevated amounts of p25, the Cdk5-activating cofactor. Long-term, but not acute, loss of Cdk5 led to decreased p25 levels, suggesting that Cdk5/p25 may be activated as a homeostatic mechanism to attenuate epileptiform activity. These findings indicate that Cdk5 regulates synaptic plasticity, controls neuronal and behavioral stimulus-induced excitability and may be a novel pharmacological target for cognitive and anticonvulsant therapies.

Interrupted sleep, fragmented sleep or restricted sleep is a corollary of many psychiatric, neurological and respiratory disorders and also results from disruptive environments such as that of the intensive care unit (ICU). Recent rodent studies have revealed that sleep interruption (SI) can have more significant consequences for cognitive and neurophysiological variables than were expected and may even be equivalent to those of total sleep deprivation. Results from this research are therefore being increasingly recognized for their implications, which may include delayed recovery from critical illness in the ICU. Here we describe in detail a method for interrupting sleep in a murine model, which we had previously adopted to show an increase in mortality after septic insult. Interrupting sleep for 30s every 2 min over 48 h significantly decreased rapid eye movement (REM) and non-rapid eye movement (NREM) sleep. The technique, which is based on using a standard laboratory orbital shaker to oscillate the cage containing the mouse, can easily be adapted to use different parameters for SI. During recovery, mice exhibited a rebound in REM sleep time and an increase in the depth of NREM sleep as measured by delta (1-4 Hz) power in the electroencephalogram. The changes in sleep both during and after SI showed some differences from those previously observed in the rat using the same SI parameters. In conclusion, the mouse may provide a useful alternative model for studying the effects of SI.

Sleep deprivation is a common problem in the intensive care unit. Animal models have demonstrated that sleep deprivation alone is associated with increased mortality. We have previously shown that septic insult with sleep deprivation results in increased mortality in a murine model. The aging process is known to reduce the restorative phases of sleep. The purpose of this study was to evaluate the effect of age on mortality with sleep deprivation during recovery from septic insult.C57BL/6J male mice aged 2 months (young) or 9 months (old) underwent cecal ligation and puncture (CLP). Animals were randomized to receive sleep interruption (SI) for 48 hours or standard recovery (no SI). Sham animals underwent laparotomy and cecal manipulation without puncture. SI was achieved by securing animal housing to an orbital shaker set to repeatedly cycle at 30 rpm over 120 seconds (30 seconds on/90 seconds off). The primary outcome was survival at 5 days post-CLP. Kaplan-Meier survival analysis with log-rank test was used to explore differences in mortality.SI resulted in an increase in time awake for both light and dark cycles (p0.001). Mortality after CLP with SI (n = 30) was 57% and mortality after CLP without SI (controls; n = 33) was 24%. SI was associated with a greater than 3-fold increase in mortality after CLP (RR = 3.29; 95% CI, 1.42-7.63). Young mice (n = 28) had a mortality of 31% with CLP alone increasing to 67% with SI (p = 0.03). Old mice (n = 35) had a mortality of 18% with CLP alone increasing to 50% with SI (p = 0.05). There was no difference in survival between young and old mice undergoing SI (p = 0.49).Sleep deprivation after septic insult increases mortality in both young and old mice. However, sleep deprivation after septic insult does not have a more profound effect on mortality in either age group. These findings suggest that sleep deprivation experienced in the intensive care unit setting during recovery from critical illness may increase mortality. This effect appears independent of increased age. Further studies evaluating extremes of age are warranted.

Two of the primary symptoms of narcolepsy, cataplexy and sleep attacks, have been amenable to study in canine and rodent models of narcolepsy. The canine model has proved to be an important research tool in the pathophysiology since the disorder was first identified in this species in 1973. Neurotransmitter differences in the narcoleptic canines, combined with evaluation of compounds that can both decrease and increase the incidence of cataplectic episodes, have provided insight into possible brain changes associated with the disorder, and suggested new treatment regimes. Since canine narcolepsy was found to be transmissible in at least two breeds, this model also provided a feasible tool for a genetic study of the disorder. A decade-long positional cloning effort eventually resulted in the identification of the gene that was mutated in dogs: a receptor for the orexin (also called hypocretin) neuropeptides. Coincidentally, but independently, a mouse line was created with the orexin gene deleted. In a remarkable concordance of results, the discovery of both the canine gene and the narcoleptic phenotype of the orexin knockout mouse occurred within weeks, if not days, of each other. These studies resulted in an unequivocal linkage between the pathophysiology in the orexin system and the narcolepsy symptoms. Studies on these models are continued to understand how orexin might normally prevent the dysregulated transitions between different vigilance states, which appear to be at the core of the narcoleptic symptomatology. The etiology of the disorder, and the nature and mechanism of cataplexy remain important issues that are amenable to research with the animal models.

Melanin-concentrating hormone (MCH) is a hypothalamic neuropeptide that has been implicated in energy homeostasis. Pharmacological studies with MCH and its receptor antagonists have suggested additional behavioral roles for the neuropeptide in the control of mood and vigilance states. These suggestions have been supported by a report of modified sleep in the MCH-1 receptor knockout mouse. Here we found that MCH knockout (MCH(-)(/)(-)) mice slept less during both the light and dark phases under baseline conditions. In response to fasting, MCH(-)(/)(-) mice exhibited marked hyperactivity, accelerated weight loss and an exaggerated decrease in rapid eye movement (REM) sleep. Following a 6-h period of sleep deprivation, however, the sleep rebound in MCH(-)(/)(-) mice was normal. Thus MCH(-)(/)(-) mice adapt poorly to fasting, and their loss of bodyweight under this condition is associated with behavioral hyperactivity and abnormal expression of REM sleep. These results support a role for MCH in vigilance state regulation in response to changes in energy homeostasis and may relate to a recent report of initial clinical trials with a novel MCH-1 receptor antagonist. When combined with caloric restriction, the treatment of healthy, obese subjects with this compound resulted in some subjects experiencing vivid dreams and sleep disturbances.

Orexin (also called hypocretin) is a neurotransmitter that influences central arousal and vigilance state, especially by coupling the level of arousal to energy balance. Localized uniquely to neurons with cell bodies in the lateral hypothalamus, orexin has widespread projections throughout the neuraxis, including the raphe nuclei. Dense orexinergic innervation has been noted in the dorsal raphe, where direct axodendritic contacts between orexinergic fibers and serotoninergic processes have also been described. Orexin depolarizes dorsal raphe serotoninergic cells, an effect that is consistent with activation of the canonical form of the transient receptor potential channel. Thus, the effect of orexin will be integrated with other neurotransmitters that also depolarize serotoninergic cells through this channel, including noradrenaline and histamine. However, in addition to the direct action, orexin at higher concentrations also depolarizes GABAergic interneurons and reduces glutamatergic release at local terminals through a retrograde endocannabinoid signal. The functional consequences of these interactions between orexin and serotonin remain to be elucidated, although they are consistent with synaptic modification. Hence orexin may modulate the serotoninergic influence in the forebrain as part of an adaptive response to homeostatic disequilibrium.

Pharmacological approaches to our understanding of sleep have been at the forefront of sleep research for many years. Traditional techniques have included the use of pharmacological agonists and antagonists, as well as transmitter-specific lesions. These have been enhanced by the introduction of molecular genetics and the use of transgenes and targeted gene deletion. Neurochemistry of Sleep and Wakefulness is an exceptional, single source of information on the role of the major mammalian neurotransmitter systems involved in the regulation of sleep and waking. With contributions from internationally recognized experts, this book clearly describes how researchers have made use of the myriad techniques in their armamentarium to characterize the role of a given neurotransmitter in the regulation of sleep and waking. Suitable for experimental and clinical pharmacologists, the book will have wider appeal to sleep researchers, psychiatrists and any professional interested in the interdisciplinary areas of neurobiology and pharmacology.

Conditional deletion of Pten (phosphatase and tensin homolog on chromosome ten) in differentiated cortical and hippocampal neurons in the mouse results in seizures, macrocephaly, social interaction deficits and anxiety, reminiscent of human autism spectrum disorder. Here we extended our previous examination of these mice using electroencephalogram/electromyogram (EEG/EMG) monitoring and found age-related increases in spontaneous seizures, which were correlated with cellular dispersion in the hippocampal dentate gyrus. Increased spontaneous locomotor activity in the open field on the first and the second day of a 3-day continuous study suggested heightened anxiety in Pten mutant mice. In contrast, the mutants exhibited decreased wheel running activity, which may reflect reduced adaptability to a novel environment. Synchronization to the light-dark cycle was normal, but for up to 28 days under constant darkness, the Pten mutants maintained a significantly lengthened and remarkably constant free-running period of almost exactly 24 h. This result implies the involvement of Pten in the maintenance of circadian rhythms, which we interpret as being due to an effect on the phosphatidylinositol 3-kinase (PI3K) signaling cascade.

An exciting recent development in sleep research was the discovery of the importance for vigilance state control and narcolepsy-cataplexy of neurons containing orexin (also known as hypocretin) neuropeptides. Narcolepsycataplexy is a chronic, debilitating sleep disorder that is characterized by excessive daytime sleepiness, manifested as attacks of daytime somnolence at inappropriate times. 1–3 Narcoleptics also show symptoms that are considered indicative of abnormal REM (Rapid Eye Movement) sleep expression. These latter symptoms include cataplexy, hypnogogic hallucinations, sleep-onset REM periods, and sleep paralysis. In contrast to daytime sleepiness, the nighttime sleep of narcoleptics is fragmented and of poor quality, typically demonstrating lengthy periods of wakefulness after sleep onset