A. Top: Immunostained sections of the cerebral cortex of mice sacrificed after 6-h of sleep deprivation (SD; left) or after 6-h of SD followed by 2.5-h recovery sleep (RS) period (right). Black=Fos-immunoreactive nuclei; pink=nNOS-immunoreactive cell bodies. Note that, although there are many Fos⁺ nuclei in the awake animal, none of them are located within the nNOS neurons whereas, in the animal that had a chance to sleep, a high proportion of the nNOS neurons contain Fos⁺ nuclei. Bottom: The proportion of Fos⁺/nNOS cells was significantly greater in the RS group (“asleep”) than in the SD group (“awake”) in every cortical region and in the posterior portion of the caudate-putamen. Data are mean ± s.e.m; *P<0.05 compared to corresponding SD group. Abbreviations: Cg Cx, Cingulate Cortex; M Cx, Motor Cortex; S Cx, Somatosensory Cortex; I Cx, Insular Cortex; Pir Cx, Piriform Cortex; Ent Cx, Entorhinal Cortex; TeA Cx, Temporal Association Cortex; V Cx, Visual Cortex; CPu, a, Caudate Putamen (+1.0 mm from bregma); CPu, p, Caudate Putamen (−1.5 mm from bregma). B and C. The proportion of Fos⁺/nNOS-immunoreactive cells (B) in the mouse cortex varied across the 24-h period in relation to prior sleep history, and was related to NREM delta energy (C), which is an index of homeostatic sleep drive. ZT= Zeitgeber Time, the time since light onset. Shaded area indicates the 12-h dark period. D. Using the data from B and C, the percentage of Fos⁺/nNOS cells in the mouse cortex was highly correlated with NREM delta energy across the 24-h period. Reproduced, with permission, from Ref. [42].

A. “Sleep-active” cortical neurons are nNOS cells that express nNOS, NPY and SST, and are likely to be of the Type I class. During wakefulness (illustrated by low amplitude EEG and high amplitude EMG recordings on the right), these cells are inhibited by afferent input from “wake-active” regions such as the histaminergic (HA) tuberomammillary nuclei, noradrenergic (NA) locus coeruleus, serotonergic (5HT) raphé nuclei, and/or basal forebrain cholinergic (ACh) neurons. These inhibitory inputs result in low activity of nNOS neurons during wakefulness. B. During slow wave sleep (SWS; illustrated by high amplitude EEG and low amplitude EMG relative to panel A), inhibitory input from wake-active regions is reduced (indicated by the thinner lines relative to A) and this disinhibition, along with putative excitatory input from sleep promoting substances such as adenosine, cytokines, and the neuropeptides cortistatin and growth hormone-releasing hormone (GHRH), results in activation of nNOS cells. In this model, “+” and “−“ refer to functional activation or inhibition of nNOS cells in vivo rather than direct excitation or inhibition at a cellular level. Functional activation (increased Fos expression) or inhibition could occur either directly or indirectly through disinhibition or disfacilitation, respectively.

A. Upper: Hypnogram illustrating the temporal progression through various stages of sleep across a 7 h night in a young adult. Abbreviations: W, wakefulness; 1–4, NREM Sleep Stages 1–4; R, REM Sleep. Lower: Slow Wave Activity (SWA; mean power density in the 0.75- to 4.0-Hz band of the EEG) determined in 1-min epochs. Note that the amplitude of the recurring peaks of SWA decline across the night. Adapted, with permission, from Ref. [100]. B. The "two-process model" of homeostatic sleep regulation proposed by Borbely [1] in which the homeostatic sleep-related "Process S" is proposed to interact with input from the circadian system ("Process C") to gate the occurrence of sleep and wakefulness. Process S is proposed to be a biochemical process(es) which begins to build up at the onset of wakefulness; SWA during NREM sleep (as illustrated in the lower panel of Figure 1A) is thought to be an index of the declining phase of Process S. C. Schematic illustrating the occurrence of SWA across three nights in basal conditions (top panel) and in response to a night of sleep deprivation (middle panel). Note that the amplitude of the SWA is greater on the recovery night after sleep deprivation than on the baseline night, indicating a homeostatic response. In the bottom panel, a curve has been fitted to the declining phase of Process S on Night 1, the rising phase of Process S on (Day and) Night 2 (during which sleep deprivation occurs), and the declining phase again on Night 3. The dynamics of the sleep/wake-dependent changes in SWA have been quantified with the use of computer simulations and SWA can now be predicted in great detail in several species

Triple staining for (A) nNOS, (B) NPY and (C) Fos immunoreactivity in mouse cortex during recovery sleep reveals that only neurons that express both nNOS and NPY contain Fos (arrows), whereas neurons that express only NPY (triangles) do not contain Fos (adapted, with permission, from Ref. [42]). D. Infrared videomicroscopy image of a cortical interneuron filled intracellularly with a marker (biocytin) in (E) showing a right horizontal dendritic arborization coursing toward the responsive blood vessel (blue gray color). F. Stimulation of a single nNOS interneuron induces reversible dilatation in vitro. Images of a blood vessel before (left) and after (right) onset of evoked firing (start time 0 sec, duration 30 sec); pial surface is upward. Vessel diameter is shown as a yellow line in both panels. G. Time course of the vasodilation illustrated in G, which was reversible (adapted, with permission, from Ref. [58]).