**2.3 Mechanisms of sleep regulation**

158 Neuroimaging – Cognitive and Clinical Neuroscience

important for the memory consolidation and reconsolidation during SWS (Diekelmann &

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Fig. 1. Electrophysiological signatures of distinct sleep-wake stages: EEG recorded from Fz,

Unlike non-REM sleep electrophysiology, REM sleep EEG signatures (Fig. 1) include lowvoltage desynchronized wake-like EEG activity comprising theta and fast (beta and gamma) rhythms accompanied by a swift occurrence of rapid eye movements (REM) upon lack of muscle tone (Aserinsky & Kleitman, 1953; Cantero et al., 2003; Clemens et al., 2009). Hippocampal theta rhythm is a prominent REM sleep EEG signature in rodents (Gottesmann, 1999; Kirov & Moyanova, 2002) and felines (Hobson & Pace Schott, 2002), while in human hippocampus and neocortex it is less coherent (Cantero et al., 2003). Further, REM sleep is hallmarked by ponto-geniculo-occipital (PGO) waves. PGO waves are driven by intense bursts of synchronized activity that propagate from the pontine brainstem mainly to the lateral geniculate nucleus and visual cortex (Callaway et al., 1987; Hobson & Pace-Schott, 2002; Pace-Schott & Hobson, 2002). They occur in temporal association with

Cz and Pz, vertical (v) and horizontal (h) eye movements, and electromyogram (m).

Stage 1 of non-REM Sleep Stage 2 of non-REM Sleep

Slow Wave Sleep REM Sleep

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Active Wake Quiet Wake

Born, 2010).

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100 µV 100 µV 100 µV 100 µV 100 µV 100 µV The regulation of sleep is active in its own rights, and is closely related to sleep's physiology and functions (Hobson, 2005; Pace-Schott & Hobson, 2002). The respective neurobiological mechanisms are represented by complex reciprocal interactions between different neuronal populations and their chemical modulators and transmitters in distinct functional states across sleep-wake cycle leading to distinct functional states (Gottesmann, 1999; Hobson et al., 1975; Hobson & Peace-Schott, 2002; Peace-Schott & Hobson, 2002). Two major brain regions are mostly considered in sleep regulation, especially when functions of sleep are concerned (Pace-Schott & Hobson, 2002). The first engages neuronal populations located in the diencephalon, in particular, the hypothalamus, mostly involved in the circadian regulation of sleep. The second brain region engages brainstem or meso-pontine and basal forebrain nuclei spread in the reticular ascending system (RAS) and projecting noradrenaline (NA), serotonin (5-Hydroxytryptamine, 5-HT) and acetylcholine (Ach) neuromodulatory signals to upper brain structures including the basal ganglia and amygdala, thalamus, hippocampus, and cortex. These mechanisms are essential for the ultradian alternating expression of non-REM sleep-REM sleep periods (Gottesmann, 1999; Hobson et al., 1975; Pace-Schott & Hobson, 2002). Briefly, during wake, brainstem/mesopontine NA, 5-HT, Ach, and hypothalamic histamine (HIS) neurons projecting to upper brain structures and cortex, are all active, thus sustaining functional brain states optimal to the environmental requirements (Gottesmann, 1999; Hobson & Pace-Schott, 2002; Pace-Schott & Hobson, 2002). As sleep deepens from stages 1 and 2 to SWS, all these neuromodulators progressively decrease their activities, with their lowest levels observed during SWS. This leads to strongly diminished or lacking RAS neuromodulation of upper brain structures and cortex, which in turn, is responsible for the appearance of non-REM sleep EEG signals represented by sleep spindles, K-complexes and SWA, all originating from thalamo-cortical and cortico-cortical interplay (McCormick & Bal, 1997; Pace-Schott & Hobson, 2002; Steriade & Timofeev, 2003). In REM sleep, all NA, 5-HT and HIS neurons cease their firing. In contrast, Ach excessive over-activity emerges projecting to the cortex and all sub-cortical structures, which produces the electrophysiological signatures of REM sleep (Gottesmann, 1999; Pace-Schott & Hobson, 2002).

#### **2.4 Neuroimaging of sleep and wake**

Several neuroimaging studies using either fMRI or positron-emission tomography (PET) have investigated the pattern of brain activation across wake, non-REM and REM sleep. These studies have demonstrated that anterior cingulate cortex, right and left amygdaloid complexes, pons, parahippocampal cortex, and extrastriate visual cortex are more active during REM sleep compared with wake and non-REM sleep, whereas the activation of other brain areas including right and left dorsolateral prefrontal cortices, right and left parietal cortices and precuneus, posterior cingulate cortex, and primary visual cortex, is suppressed in REM sleep compared with wake. All these brain regions have been shown to be the most

The Memory, Cognitive and Psychological Functions of Sleep:

dependent memory consolidation.

McCormick, 2010; Reis et al., 2008).

Update from Electroencephalographic and Neuroimaging Studies 161

particular, has been proposed as an essential mechanism underlying the consolidation of hippocampus-dependent memories (Buzsáki, 1989; Marshall & Born, 2007; Mölle et al., 2004; Steriade, 2001). In human SWS, "up" and "down" EEG states of SO are shown be dissimilarly associated with a number of electrophysiological events. Specifically, the "up" state of the SO is marked by increased occurrence of delta slow waves and sleep spindles, whereas during the 'down" state of SO, delta slow wave and sleep spindle activities markedly decrease (Mölle et al., 2002; 2004). Thus, human SO are demonstrated to group both slow waves and spindles. Further, a rapid increase of the underlying neuronal activity (depolarizing state) and a rapid decrease in it (hyperpolarizing state) have recently been shown to characterize human SO "up" and "down" wave forms, respectively (Nir et al., 2011). Finally, human studies have demonstrated time and phase coupling between SO, slow waves, sleep spindles, and hippocampal SWR bursts (Clemens et al., 2007; 2011; Nir et al., 2011). Collectively, these findings strongly indicate that non-REM sleep/SWS SO represent an EEG mechanism involved in plastic changes sub-serving the hippocampus-

Indeed, SO have been shown to be strongly associated with both procedural (Huber et al., 2004) and declarative (Mölle et al., 2004) memory consolidation taking place in human non-REM sleep/SWS. Later, to verify the specific role for SO in hippocampus-dependent memory consolidation, a series of studies, in which brain rhythms have been modulated using trans-cranial direct current stimulation (tDCS), has been conducted in humans. This method is now recognized as a reliable tool for modulating both the internally generated brain rhythms and the activity of underlying neuronal populations, depolarized under anodal tDCS and hyperpolarized under cathodal tDCS, respectively (Fröhlich &

Initially, weak (not perceived by subjects) anodal tDCS oscillating at 0.75 Hz (slow oscillation stimulation, SOS) has been delivered during the transition from stage 2 of non-REM sleep to SWS after declarative and procedural learning before sleep. Compared with a sham condition, stimulation has selectively produced a gain in only declarative memory after sleep. Importantly, it has also produced a substantial increase in SO (~ 0.75 Hz) and frontal slow alpha spindle (8-12 Hz) activity, possibly by entraining these sleep EEG rhythms (Marshall et al., 2006). These findings have provided strong evidence for the role of SO and/or frontal slow spindle activity for the hippocampus-dependent memory consolidation. However, they have not addressed the question of whether SOS itself or whether endogenous SO boosted by the SOS have resulted in improvement of the consolidation of declarative memory found. This question was addressed in two later studies. In these studies, weak anodal tDCS oscillating at frequencies not common for the respective functional brain states was applied. In particular, SOS oscillating at 0.75 Hz was applied during quiet or resting wake retention period after learning declarative and procedural tasks. SOS did not affect either declarative or procedural memory consolidation, nor did it affect working memory and mood at retest. However, in contrast to its EEG effects during non-REM sleep (Marshall et al., 2006), it produced only a local (at the frontal sites of stimulation) increase in EEG power in SO (0.4-1.2 Hz) frequency band, accompanied by a widespread and strong increase in theta (4-8 Hz) EEG power. Further, when delivered in active wake state during encoding of a verbal learning memory task, the 0.75 Hz SOS produced virtually the same EEG effects as during quite wake (local increase in SO and widespread increase in theta power), but it significantly improved encoding of declarative verbal information (Kirov et al., 2009). Recently, tDCS oscillating at 5 Hz (wake and/or REM

suppressed during non-REM sleep (Broun et al., 1997; 1998; Maquet et al., 1996; Miyauchi et al., 2009; Nofzinger et al., 1997). Yet, other brain structures are shown to specifically increase their activation in relation to distinct non-REM sleep stages and their EEG signatures. For example, blood oxygen level-dependent (BOLD) signal from the thalamus is strongest during spindle activity in stage 2 of non-REM sleep (Schabus et al., 2007), whereas during SO in SWS, brain functional activation is strong in the medial temporal cortex, the parahippocampal cortex, and neocortical areas (Dang-Vu et al., 2005; 2008; Maquet et al., 1997).

### **2.5 Mental characteristics of the sleep-wake stages**

Notably, from a cognitive point of view, the mental characteristics of sleep-wake stages well correspond to their brain activation patterns found in neuroimaging studies (Fosse et al., 2001; 2004; Hobson & Pace-Schott, 2002; Hobson et al., 2000; Stickgold et al., 2001). Thus, wake is characterized by strongest and most logic thoughts in the presence of sensory input, executive control and goal-directed behavior. Sleep onset is hallmarked by the so called hypnagogic hallucinations, and as sleep deepens from stage 2 of non-REM sleep to SWS, thinking becomes more and more scares, and almost absent during SWS. Yet, there are logic thoughts mostly associated with previous wake experiences (Fosse et al., 2004; Hobson, 2005; Hobson et al., 2000; Stickgold et al., 2001). During REM sleep, mental activity is likely hallucinatory, and is behaviorally expressed in vivid, bizarre and elusive dreams (Fosse et al., 2004; Hobson et al., 2000; Stickgold et al., 2001). Also, REM sleep mentality is characterized by a most salient emotional tone upon lack of sensory input (Hobson et al., 2000) and executive control, as evidenced by the neuroimaging data (suppression of the dorsolateral prefrontal cortex, e.g., Maquet et al., 1996).
