Sleep and wakefulness are two basic behavioural states identifiable in virtually all species of animal. Sleep occurs in two alternating stages: non-rapid eye movement (REM) sleep, and REM sleep. The timing of sleep and waking is controlled by both homeostatic need and the circadian clock. Sleep-wake states are also influenced by circulating steroid hormones, such as estrogen. With the long-term goal to understand the neural mechanisms of behavioural state control, we have been working on the following main projects using rodent models.
1. Basal forebrain mechanisms of wake and sleep states and attention
The basal forebrain (BF) has been implicated in cortical activation, attention, learning and memory. These functions have long been ascribed to its cholinergic neurons, a main cell population in this area. However, the BF contains many non-cholinergic neurons as well. Furthermore, recent evidence suggests that, in addition to the cognitive functions, the BF plays an important role in behavioural arousal and sleep homeostasis. For example, the potential sleep promoting factor adenosine selectively accumulates in the BF during prolonged wakefulness, and inhibits BF neurons to promote transition to sleep. We hypothesize that the BF is a major forebrain site for promoting behavioural arousal and that the promotion of arousal by the BF is executed through its ascending and descending pathways to multiple brain regions involved in cognitive, motivational, emotional, and autonomic functions. These pathways might use different transmitters. We are currently investigating which BF neurons and which pathways are responsible for cortical activation, behavioural arousal, and sleep homeostasis, respectively, by using somnographic EEG-EMG recording, selective neurotoxins, and microinjection of drugs.
Although best known for its cholinergic neurons, the BF also contains GABAergic and glutamatergic neurons that project to the cortex and other targets. C olocalization of multiple neurotransmitters in single neurons is now considered to be a norm than an exception. We have been using immunohistochemistry, tract tracing techniques, and confocal microscopy to provide anatomical support for the possibility that cholinergic BF neurons not only release ACh but also glutamate. Our recent data show that BF cholinergic neurons projecting to the amygdala contain a vesicular glutamatergic marker and may release glutamate. This new evidence calls for a change in the way we understand the transmitter phenotype of BF neurons.
Cortical ACh is implicated in attention, and the main extrinsic source of cortical ACh is cholinergic BF neurons. In collaboration with Dr. Doug Rasmusson, we have been investigating whether ACh release in the cortex is controlled in a region- and modality-specific manner. Our in vivo microdialysis data show that ACh release increases differentially in different cortical regions and also in a modality-specific manner, consistent with the role of ACh in attention.
2. Adenosinergic mechanisms of sleep
Adenosine has been thought to be an endogenous sleep-promoting factor. A common experience supporting this notion is the wake-promoting effect of caffeine, which is mediated by blockade of adenosine receptors in the brain. Sleep deprivation increases adenosine levels selectively in the basal forebrain. What mechanisms underlie this increase? We showed previously that the activation of M3 muscarinic receptors can enhance glutamate-evoked adenosine release in the cortex in vitro. We also showed in unanesthetized rats that glutamate, ACh and noradrenaline, major transmitters implicated in behavioural arousal, can increase cortical adenosine concentrations.
To understand the adenosinergic regulation of behavioural state, we have also been mapping the neurons that are activated by psychostimulant doses of caffeine and a selective adenosine receptor antagonist, using the immediate early gene product c-Fos as a marker for neuronal activation. One cell population that is activated by caffeine is a group of neurons containing orexin, a novel neuropeptide implicated for a role in arousal and in the etiology of the sleep disorder narcolepsy. This activation may be a direct or indirect effect, but it appears to be fairly selective, because a few other, but not all, arousal-promoting cell groups are activated by caffeine. T hese different patterns of neuronal activation across brain regions may reflect the cognitive, somatomotor and autonomic activation associated with the arousal response to caffeine.
3. The role of neuropeptides and glutamate in REM sleep control
Rapid eye movement (REM) sleep is a unique stage of sleep in which the brain is active but the body is asleep. REM sleep is thought to be induced by release of ACh in the pontine reticular formation. However, recent evidence suggests that neuropeptides might also play an important role. We investigated the possibility that substance P is co-released with ACh by studying the anatomical colocalization of the peptide with mesopontine cholinergic neurons, and by testing the electrophysiological effect of substance P on reticular neurons. Our data suggest that substance P and ACh might be co-released and co-activate reticular neurons to initiate and maintain REM sleep. We are also investigating the role of glutamate in the pontine reticular formation in inducing and maintaining REM sleep.
4. Circadian control of sleep and wakefulness.
Daily rhythms characterize virtually all aspects of life on Earth, and many of these rhythms are governed by an internal biological clock called the circadian clock. In mammals, the circadian clock is housed in the suprachiasmatic nucleus (SCN) of the hypothalamus. While the presence of a ~24-hour rhythm is a main feature of sleep, the SCN does not directly control sleep or wakefulness but, rather, these states are controlled by a system that is located outside of the SCN and, in fact, is distributed widely throughout the brain. In exploring the linkage between the circadian clock and the sleep/wake-regulatory system, in collaboration with Dr. Ben Rusak we identified and characterized the projections from the SCN to neurons in the ventrolateral preoptic nucleus, a main sleep-promoting nucleus, using electrophysiological recordings from brain slices. These results suggested that the circadian clock housed in the SCN can influence the activity of the preoptic neurons directly to regulate preoptic mechanisms of sleep initiation and maintenance.
Despite this interesting evidence, the SCN is not known to project directly to any other structures with primary roles in sleep-wake regulation. This led us to hypothesize that much of the circadian signal from the SCN to the sleep-wake system is mediated through indirect pathways. Using dual tract-tracing techniques in rat, we have shown that several preoptic and hypothalamic nuclei are well positioned to serve as intermediary nuclei to convey the circadian signal from the SCN to sleep- and wake-regulatory cell groups in the forebrain and brainstem. These indirect neuronal pathways might play a major role in circadian control of sleep-wake cycles. Building up on these data, we are currently investigating the distribution of arousal/sleep-related afferents to specific compartments of the SCN.
5. Cellular and behavioural impact of sleep deprivation, and its modulation by female sex steroids
Sleep disorders are more prevalent in women than in men, and specific associations have been proposed between sleep disturbances and women’s changing endocrine status across the menstrual cycle, in pregnancy and during menopause. Despite its potential significance, the mechanism underlying the proposed interactions between hormonal status and sleep loss in females has remained largely unexplored. In collaboration with Drs. Ben Rusak and Mike Wilkinson, w e have studied patterns of neuronal activation resulting from acute (3 or 6 h) sleep deprivation starting at different times of day in male rats. We demonstrated that sleep deprivation early in the day (normal rest phase) increases the expression of immediate-early genes (IEGs) such as c-Fos in several brain regions, including the preoptic area and amygdala. To investigate the interaction of hormonal status with sleep loss, we have been studying sleep deprivation-induced IEG expression after sleep deprivation in hormonally manipulated female rats and intact male rats. We are also using polygraphic-video recordings to characterize the architecture of their recovery sleep and EEG power spectra following sleep deprivation.
Although sleep loss appears to affect virtually all aspects of life, its effects on cognitive function are particularly problematic. Indeed, normal sleep has been proposed to play a critical role in the formation and storage of new memories and skills. We have been using one relatively well characterized model of neuronal plasticity, i.e., cell proliferation and survival in the dentate gyrus of the hippocampus, to examine the cellular effects of sleep loss that may contribute to the negative impact of sleep deprivation on cognitive functions. E strogen has been suggested to influence learning and memory, and there is evidence that estrogen enhances cell proliferation in the hippocampus . We have therefore been manipulating estrogen levels in female rats and assessing how estrogen alters the effects of sleep loss on this measure of hippocampal neuronal plasticity.
Research Support
Canadian Institutes of Health Research
Natural Sciences and Engineering Council of Canada
Representative publications
Fournier, G.N., Materi, L.M., Semba, K. and Rasmusson, D.D. (2004) Cortical acetylcholine release and EEG activation evoked by ionotropic glutamate receptor agonists in the rat basal forebrain. Neuroscience, 123:785-792.
Fournier, G.N., Semba, K. and Rasmusson, D.D. (2004) Modality- and region-specific acetylcholine release in the rat cortex. Neuroscience, 126:257-262.
Semba, K. (2004) Phylogenetic and ontogenetic aspects of the basal forebrain cholinergic neurons and their innervation of the cerebral cortex. Progress in Brain Research, Volume 145, Elsevier, Amsterdam, pp. 3-43.
Deurveilher, S. and Semba, K. (2005) Indirect projections from the suprachiasmatic nucleus to major arousal-promoting cell groups in rat: Implications for the circadian control of behavioural state. Neuroscience, 130:165-183.
Deurveilher, S., Lo, H., Murphy, J.A., Burns, J. and Semba, K. (2006) Differential c-Fos immunoreactivity in arousal-promoting cell groups following systemic administration of caffeine in rats. J. Comp. Neurol., 498, 667-689.
Nickerson Poulin, A., Guerci, A., El Mestikawy, S. and Semba, K. (2006) Vesicular glutamate transporter 3 immunoreactivity is present in cholinergic basal forebrain neurons projecting to the basolateral amygdala in rat. J. Comp. Neurol., 498: 690-711.
Deurveilher, S. and Semba, K. (2006) Mapping sleep-wake control with the transcription factor c-Fos. R. Pinaud and L. Tremere (eds.) Immediate Early Genes in Sensory Processing, Cognitive Performance and Neurological Disorders. Springer, New York, pp. 113-136.
Rasmusson, D.D., Smith, S.A. and Semba, K. (2007) Inactivation of prefrontal cortex abolishes cortical acetylcholine release evoked by sensory or sensory pathway stimulation in the rat. Neuroscience, 149: 232-241
Kaur, S., Junek, A., Black, M.A., and Semba, K. (2008) Effects of ibotenate and 192IgG-saporin lesions of the nucleus basalis magnocellularis/substantia innominata on spontaneous sleep and wake states and on recovery sleep after sleep deprivation in rats. J. Neurosci., 28:491-504.
Lindley, J., Deurveilher, S., Rusak, B., and Semba, K. (2008) Transforming growth factor- and glial fibrillary acidic protein in the hamster circadian system: daily profile and cellular localization. Brain Research 1197(1): 94-105.
Deurveilher, S. and Semba, K. (2008) Reciprocal connections between the suprachiasmatic nucleus and the midbrain raphe nuclei: A putative role in the circadian control of behavioral states. S.R. Pandi, J.M. Monti, B.L. Jacobs, D.J. Nutt (Eds): Serotonin and sleep: Molecular, Functional and Clinical Aspects. Birkhäuser, Basel, pp. 103-131.
Members of the Lab
Samuel Deurveilher, Research Associate
Frederick Li, Graduate Student (Ph.D.)
Michelle Black, Graduate Student (M.Sc.)
Nabilah Chowdhury (Summer Student)
Ahmed Rostom (Summer Student)
Ilia Pavlovski (Summer Student)
Joan Burns, Technician
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HALIFAX,
NOVA SCOTIA | CANADA 