Patrick M. Fuller

Sleep state switching.

We take for granted the ability to fall asleep or to snap out of sleep into wakefulness, but these changes in behavioral state require specific switching mechanisms in the brain that allow well-defined state transitions. In this review, we examine the basic circuitry underlying the regulation of sleep and wakefulness and discuss a theoretical framework wherein the interactions between reciprocal neuronal circuits enable relatively rapid and complete state transitions. We also review how homeostatic, circadian, and allostatic drives help regulate sleep state switching and discuss how breakdown of the switching mechanism may contribute to sleep disorders such as narcolepsy.

Neurobiology of the sleep-wake cycle: sleep architecture, circadian regulation, and regulatory feedback

This mini-review article presents the remarkable progress that has been made in the past decade in our understanding of the neural circuitry underlying the regulation of sleep-wake states and circadian control of behaviors. Following a brief introduction to sleep architecture and physiology, the authors describe the neural circuitry and neurotransmitters that regulate sleep and cortical arousal (i.e., wakefulness). They next examine how sleep and wakefulness are regulated by mutual inhibition between sleep-and arousal-promoting circuitry and how this interaction functions analogously to an electronic “flip-flop” switch that ensures behavioral state stability. The authors then discuss the role of circadian and homeostatic processes in the consolidation of sleep, including the physiologic basis of homeostatic sleep drive (i.e., wake-dependent increase in sleep propensity) and the role of the SCN in the circadian regulation of sleep-wake cycles. Finally, they describe the hypothalamic circuitry for the integration of photic and nonphotic environmental time cues and how this integration allows organisms to sculpt patterns of rest-activity and sleep-wake cycles that are optimally adaptive.

Reassessment of the structural basis of the ascending arousal system

The “ascending reticular activating system” theory proposed that neurons in the upper brainstem reticular formation projected to forebrain targets that promoted wakefulness. More recent formulations have emphasized that most neurons at the pontomesencephalic junction that participate in these pathways are actually in monoaminergic and cholinergic cell groups. However, cell‐specific lesions of these cell groups have never been able to reproduce the deep coma seen after acute paramedian midbrain lesions that transect ascending axons at the caudal midbrain level. To determine whether the cortical afferents from the thalamus or the basal forebrain were more important in maintaining arousal, we first placed large cell‐body‐specific lesions in these targets. Surprisingly, extensive thalamic lesions had little effect on electroencephalographic (EEG) or behavioral measures of wakefulness or on c‐Fos expression by cortical neurons during wakefulness. In contrast, animals with large basal forebrain lesions were behaviorally unresponsive and had a monotonous sub‐1‐Hz EEG, and little cortical c‐Fos expression during continuous gentle handling. We then retrogradely labeled inputs to the basal forebrain from the upper brainstem, and found a substantial input from glutamatergic neurons in the parabrachial nucleus and adjacent precoeruleus area. Cell‐specific lesions of the parabrachial‐precoeruleus complex produced behavioral unresponsiveness, a monotonous sub‐1‐Hz cortical EEG, and loss of cortical c‐Fos expression during gentle handling. These experiments indicate that in rats the reticulo‐thalamo‐cortical pathway may play a very limited role in behavioral or electrocortical arousal, whereas the projection from the parabrachial nucleus and precoeruleus region, relayed by the basal forebrain to the cerebral cortex, may be critical for this process. J. Comp. Neurol. 519:933–956, 2011.

Differential Rescue of Light- and Food-Entrainable Circadian Rhythms

When food is plentiful, circadian rhythms of animals are powerfully entrained by the light-dark cycle. However, if animals have access to food only during their normal sleep cycle, they will shift most of their circadian rhythms to match the food availability. We studied the basis for entrainment of circadian rhythms by food and light in mice with targeted disruption of the clock gene Bmal1, which lack circadian rhythmicity. Injection of a viral vector containing the Bmal1 gene into the suprachiasmatic nuclei of the hypothalamus restored light-entrainable, but not food-entrainable, circadian rhythms. In contrast, restoration of the Bmal1 gene only in the dorsomedial hypothalamic nucleus restored the ability of animals to entrain to food but not to light. These results demonstrate that the dorsomedial hypothalamus contains a Bmal1-based oscillator that can drive food entrainment of circadian rhythms.

GABAergic RIP-Cre Neurons in the Arcuate Nucleus Selectively Regulate Energy Expenditure

Neural regulation of energy expenditure is incompletely understood. By genetically disrupting GABAergic transmission in a cell-specific fashion, and by combining this with selective pharmacogenetic activation and optogenetic mapping techniques, we have uncovered an arcuate-based circuit that selectively drives energy expenditure. Specifically, mice lacking synaptic GABA release from RIP-Cre neurons have reduced energy expenditure, become obese and are extremely sensitive to high-fat diet-induced obesity, the latter due to defective diet-induced thermogenesis. Leptins ability to stimulate thermogenesis, but not to reduce feeding, is markedly attenuated. Acute, selective activation of arcuate GABAergic RIP-Cre neurons, which monosynaptically innervate PVH neurons projecting to the NTS, rapidly stimulates brown fat and increases energy expenditure but does not affect feeding. Importantly, this response is dependent upon GABA release from RIP-Cre neurons. Thus, GABAergic RIP-Cre neurons in the arcuate selectively drive energy expenditure, contribute to leptins stimulatory effect on thermogenesis, and protect against diet-induced obesity.

The pontine REM switch : past and present

Rapid eye movement (REM) sleep is a behavioural state characterized by activation of the cortical and hippocampal EEG, rapid eye movements and muscle atonia. For the past 30 years, the most widely accepted neural circuitry model for the regulation of REM sleep has emphasized reciprocal inhibitory interactions between pontine brainstem monoaminergic and cholinergic neurons. In general support of the reciprocal interaction model, neuropharmacological studies have shown that cholinergic agonists promote REM sleep and muscarinic antagonists and monoamines inhibit REM sleep. It has nevertheless proven difficult to reconcile both the theoretical framework of this model and the pharmacological data with the fact that selective lesions of either cholinergic or monoaminergic (noradrenergic, serotoninergic or dopaminergic) nuclei in the brainstem have relatively limited effects on REM sleep. Recent work by our laboratory has revealed the presence of non‐cholinergic and non‐monoaminergic mutually inhibitory REM‐off and REM‐on areas in the mesopontine tegmentum that may form the neuroanatomical basis of the switching circuitry for REM sleep. These findings posit a REM switching circuitry model that is analogous to an electronic ‘flip‐flop’ switch. In this flip‐flop switch arrangement, GABAergic REM‐on neurons (located in the sublaterodorsal tegmental nucleus (SLD)) inhibit GABAergic REM‐off neurons (located in the ventrolateral periaqueductal grey matter (vlPAG) and lateral pontine tegmentum (LPT)) and vice versa. In the REM‐on area are two populations of glutamatergic neurons, the first of which projects to the basal forebrain and regulates EEG components of REM sleep and the second of which projects to the ventromedial medulla and spinal cord and regulates atonia during REM sleep. Our findings demonstrating independent pathways mediating atonia and the EEG components of REM provide a basis for their occasional dissociation in pathological states, e.g. REM sleep behaviour disorder.

The GABAergic parafacial zone is a medullary slow wave sleep–promoting center

Work in animals and humans has suggested the existence of a slow wave sleep (SWS)-promoting/electroencephalogram (EEG)-synchronizing center in the mammalian lower brainstem. Although sleep-active GABAergic neurons in the medullary parafacial zone (PZ) are needed for normal SWS, it remains unclear whether these neurons can initiate and maintain SWS or EEG slow-wave activity (SWA) in behaving mice. We used genetically targeted activation and optogenetically based mapping to examine the downstream circuitry engaged by SWS-promoting PZ neurons, and we found that this circuit uniquely and potently initiated SWS and EEG SWA, regardless of the time of day. PZ neurons monosynaptically innervated and released synaptic GABA onto parabrachial neurons, which in turn projected to and released synaptic glutamate onto cortically projecting neurons of the magnocellular basal forebrain; thus, there is a circuit substrate through which GABAergic PZ neurons can potently trigger SWS and modulate the cortical EEG.

Medullary circuitry regulating rapid eye movement sleep and motor atonia.

Considerable data support a role for glycinergic ventromedial medulla neurons in the mediation of the postsynaptic inhibition of spinal motoneurons necessary for the motor atonia of rapid-eye movement (REM) sleep in cats. These data are, however, difficult to reconcile with the fact that large lesions of the rostral ventral medulla do not result in loss of REM atonia in rats. In the present study, we sought to clarify which medullary networks in rodents are responsible for REM motor atonia by retrogradely tracing inputs to the spinal ventral horn from the medulla, ablating these medullary sources to determine their effects on REM atonia and using transgenic mice to identify the neurotransmitter(s) involved. Our results reveal a restricted region within the ventromedial medulla, termed here the “supraolivary medulla” (SOM), which contains glutamatergic neurons that project to the spinal ventral horn. Cell-body specific lesions of the SOM resulted in an intermittent loss of muscle atonia, taking the form of exaggerated phasic muscle twitches, during REM sleep. A concomitant reduction in REM sleep time was observed in the SOM-lesioned animals. We next used mice with lox-P modified alleles of either the glutamate or GABA/glycine vesicular transporters to selectively eliminate glutamate or GABA/glycine neurotransmission from SOM neurons. Loss of SOM glutamate release, but not SOM GABA/glycine release, resulted in exaggerated muscle twitches during REM sleep that were similar to those observed after SOM lesions in rats. These findings, together, demonstrate that SOM glutamatergic neurons comprise key elements of the medullary circuitry mediating REM atonia.

Basal ganglia control of sleep–wake behavior and cortical activation

The basal ganglia (BG) are involved in numerous neurobiological processes that operate on the basis of wakefulness, including motor function, learning, emotion and addictive behaviors. We hypothesized that the BG might play an important role in the regulation of wakefulness. To test this prediction, we made cell body‐specific lesions in the striatum and globus pallidus (GP) using ibotenic acid. We found that rats with striatal (caudoputamen) lesions exhibited a 14.95% reduction in wakefulness and robust fragmentation of sleep–wake behavior, i.e. an increased number of state transitions and loss of ultra‐long wake bouts (> 120 min). These lesions also resulted in a reduction in the diurnal variation of sleep–wakefulness. On the other hand, lesions of the accumbens core resulted in a 26.72% increase in wakefulness and a reduction in non‐rapid eye movement (NREM) sleep bout duration. In addition, rats with accumbens core lesions exhibited excessive digging and scratching. GP lesions also produced a robust increase in wakefulness (45.52%), and frequent sleep–wake transitions and a concomitant decrease in NREM sleep bout duration. Lesions of the subthalamic nucleus or the substantia nigra reticular nucleus produced only minor changes in the amount of sleep–wakefulness and did not alter sleep architecture. Finally, power spectral analysis revealed that lesions of the striatum, accumbens and GP slowed down the cortical electroencephalogram. Collectively, our results suggest that the BG, via a cortico‐striato‐pallidal loop, are important neural circuitry regulating sleep–wake behaviors and cortical activation.

Brainstem and Spinal Cord Circuitry Regulating REM Sleep and Muscle Atonia

Background Previous work has suggested, but not demonstrated directly, a critical role for both glutamatergic and GABAergic neurons of the pontine tegmentum in the regulation of rapid eye movement (REM) sleep. Methodology/Principal Findings To determine the in vivo roles of these fast-acting neurotransmitters in putative REM pontine circuits, we injected an adeno-associated viral vector expressing Cre recombinase (AAV-Cre) into mice harboring lox-P modified alleles of either the vesicular glutamate transporter 2 (VGLUT2) or vesicular GABA-glycine transporter (VGAT) genes. Our results show that glutamatergic neurons of the sublaterodorsal nucleus (SLD) and glycinergic/GABAergic interneurons of the spinal ventral horn contribute to REM atonia, whereas a separate population of glutamatergic neurons in the caudal laterodorsal tegmental nucleus (cLDT) and SLD are important for REM sleep generation. Our results further suggest that presynaptic GABA release in the cLDT-SLD, ventrolateral periaqueductal gray matter (vlPAG) and lateral pontine tegmentum (LPT) are not critically involved in REM sleep control. Conclusions/Significance These findings reveal the critical and divergent in vivo role of pontine glutamate and spinal cord GABA/glycine in the regulation of REM sleep and atonia and suggest a possible etiological basis for REM sleep behavior disorder (RBD).