NMDA Receptors in the Lateral Preoptic Hypothalamus Are Essential for Sustaining NREM and REM Sleep.

The lateral preoptic (LPO) hypothalamus is a center for NREM and REM sleep induction and NREM sleep homeostasis. Although LPO is needed for NREM sleep, we found that calcium signals were, surprisingly, highest in REM sleep. Furthermore, and equally surprising, NMDA receptors in LPO were the main drivers of excitation. Deleting the NMDA receptor GluN1 subunit from LPO abolished calcium signals in all cells and produced insomnia. Mice of both sexes had highly fragmented NREM sleep-wake patterns and could not generate conventionally classified REM sleep. The sleep phenotype produced by deleting NMDA receptors depended on where in the hypothalamus the receptors were deleted. Deleting receptors from the anterior hypothalamic area (AHA) did not influence sleep-wake states. The sleep fragmentation originated from NMDA receptors on GABA neurons in LPO. Sleep fragmentation could be transiently overcome with sleeping medication (zolpidem) or sedatives (dexmedetomidine; Dex). By contrast, fragmentation persisted under high sleep pressure produced by sleep deprivation (SD), mice had a high propensity to sleep but woke up. By analyzing changes in δ power, sleep homeostasis (also referred to as "sleep drive") remained intact after NMDA receptor ablation. We suggest NMDA glutamate receptor activation stabilizes firing of sleep-on neurons and that mechanisms of sleep maintenance differ from that of the sleep drive itself.SIGNIFICANCE STATEMENT Insomnia is a common affliction. Most insomniacs feel that they do not get enough sleep, but in fact, often have good amounts of sleep. Their sleep, however, is fragmented, and sufferers wake up feeling unrefreshed. It is unknown how sleep is maintained once initiated. We find that in mice, NMDA-type glutamate receptors in the hypothalamus are the main drivers of excitation and are required for a range of sleep properties: they are, in fact, needed for both sustained NREM sleep periods, and REM sleep generation. When NMDA receptors are selectively reduced from inhibitory preoptic (PO) neurons, mice have normal total amounts of sleep but high sleep-wake fragmentation, providing a model for studying intractable insomnia.

Dysfunction of ventral tegmental area GABA neurons causes mania-like behavior.

The ventral tegmental area (VTA), an important source of dopamine, regulates goal- and reward-directed and social behaviors, wakefulness, and sleep. Hyperactivation of dopamine neurons generates behavioral pathologies. But any roles of non-dopamine VTA neurons in psychiatric illness have been little explored. Lesioning or chemogenetically inhibiting VTA GABAergic (VTAVgat) neurons generated persistent wakefulness with mania-like qualities: locomotor activity was increased; sensitivity to D-amphetamine was heightened; immobility times decreased on the tail suspension and forced swim tests; and sucrose preference increased. Furthermore, after sleep deprivation, mice with lesioned VTAVgat neurons did not catch up on lost sleep, even though they were starting from a sleep-deprived baseline, suggesting that sleep homeostasis was bypassed. The mania-like behaviors, including the sleep loss, were reversed by valproate, and re-emerged when treatment was stopped. Lithium salts and lamotrigine, however, had no effect. Low doses of diazepam partially reduced the hyperlocomotion and fully recovered the immobility time during tail suspension. The mania like-behaviors mostly depended on dopamine, because giving D1/D2/D3 receptor antagonists reduced these behaviors, but also partially on VTAVgat projections to the lateral hypothalamus (LH). Optically or chemogenetically inhibiting VTAVgat terminals in the LH elevated locomotion and decreased immobility time during the tail suspension and forced swimming tests. VTAVgat neurons help set an animal's (and perhaps human's) mental and physical activity levels. Inputs inhibiting VTAVgat neurons intensify wakefulness (increased activity, enhanced alertness and motivation), qualities useful for acute survival. In the extreme, however, decreased or failed inhibition from VTAVgat neurons produces mania-like qualities (hyperactivity, hedonia, decreased sleep).

Basal forebrain contributes to default mode network regulation.

The default mode network (DMN) is a collection of cortical brain regions that is active during states of rest or quiet wakefulness in humans and other mammalian species. A pertinent characteristic of the DMN is a suppression of local field potential gamma activity during cognitive task performance as well as during engagement with external sensory stimuli. Conversely, gamma activity is elevated in the DMN during rest. Here, we document that the rat basal forebrain (BF) exhibits the same pattern of responses, namely pronounced gamma oscillations during quiet wakefulness in the home cage and suppression of this activity during active exploration of an unfamiliar environment. We show that gamma oscillations are localized to the BF and that gamma-band activity in the BF has a directional influence on a hub of the rat DMN, the anterior cingulate cortex, during DMN-dominated brain states. The BF is well known as an ascending, activating, neuromodulatory system involved in wake-sleep regulation, memory formation, and regulation of sensory information processing. Our findings suggest a hitherto undocumented role of the BF as a subcortical node of the DMN, which we speculate may be important for switching between internally and externally directed brain states. We discuss potential BF projection circuits that could underlie its role in DMN regulation and highlight that certain BF nuclei may provide potential target regions for up- or down-regulation of DMN activity that might prove useful for treatment of DMN dysfunction in conditions such as epilepsy or major depressive disorder.

A Neural Code That Is Isometric to Vocal Output and Correlates with Its Sensory Consequences.

What cortical inputs are provided to motor control areas while they drive complex learned behaviors? We study this question in the nucleus interface of the nidopallium (NIf), which is required for normal birdsong production and provides the main source of auditory input to HVC, the driver of adult song. In juvenile and adult zebra finches, we find that spikes in NIf projection neurons precede vocalizations by several tens of milliseconds and are insensitive to distortions of auditory feedback. We identify a local isometry between NIf output and vocalizations: quasi-identical notes produced in different syllables are preceded by highly similar NIf spike patterns. NIf multiunit firing during song precedes responses in auditory cortical neurons by about 50 ms, revealing delayed congruence between NIf spiking and a neural representation of auditory feedback. Our findings suggest that NIf codes for imminent acoustic events within vocal performance.

Is xenon a suitable euthanasia agent for mice?

OBJECTIVE: To compare behavioural and electrophysiological variables of mice undergoing gas euthanasia with either xenon (Xe) or carbon dioxide (CO2). STUDY DESIGN: Single animals chronically instrumented for electroencephalography (EEG) recording were randomized to undergo euthanasia with either CO2 or Xe (n = 6 animals per group). ANIMALS: Twelve adult (>6 weeks old) male C57Bl6/n mice. METHODS: Mice were surgically instrumented with EEG and electromyogram electrodes. Following a 7-day recovery period, animals were placed individually in a sealed chamber and a 5-minute baseline recorded in 21% O2. Gas [100% Xe (n = 6) or 100% CO2 (n = 6)] was then added to the chamber at 30% chamber volume minute-1 (2.8 L minute-1) until cessation of breathing. EEG, behaviour (jumping and freezing) and locomotion speed were recorded throughout. RESULTS: Mice undergoing single gas euthanasia with Xe did not show jumping or freezing behaviours and had reduced locomotion speed compared to baseline, in contrast to CO2, which resulted in increases in these variables. EEG recordings revealed sedative effects from Xe but heightened arousal from CO2. CONCLUSIONS: Our data suggest that Xe may be less aversive than CO2 when using a 30% chamber volume minute-1 fill rate and could improve the welfare of mice undergoing gas euthanasia.

Individual differences in heart rate reveal a broad range of autonomic phenotypes in a free-living seabird population.

Animals in the same population consistently differ in their physiology and behaviour, but the underlying mechanisms remain poorly understood. As the autonomic nervous system regulates wide-ranging physiological functions, many of these phenotypic differences may be generated by autonomic activity. We investigated for the first time in a free-living animal population (the streaked shearwater, Calonectris leucomelas, a long-lived seabird) whether individuals consistently differ in autonomic activity, over time and across contexts. We repeatedly recorded electrocardiograms from individual shearwaters, and from heart rate and heart rate variability quantified sympathetic activity, which drives the 'fight-or-flight' response, and parasympathetic activity, which promotes 'rest-and-digest' processes. We found a broad range of autonomic phenotypes that persisted even across years: heart rate consistently differed among individuals during periods of stress and non-stress and these differences were driven by parasympathetic activity, thus identifying the parasympathetic rest-and-digest system as a central mechanism that can drive broad phenotypic variation in natural animal populations.

The low-down on sleeping down low: pigeons shift to lighter forms of sleep when sleeping near the ground.

Sleep in birds is composed of two distinct sub-states, remarkably similar to mammalian slow-wave sleep (SWS) and rapid eye movement (REM) sleep. However, it is unclear whether all aspects of mammalian sleep are present in birds. We examined whether birds suppress REM sleep in response to changes in sleeping conditions that presumably evoke an increase in perceived predation risk, as observed previously in rodents. Although pigeons sometimes sleep on the ground, they prefer to sleep on elevated perches at night, probably to avoid nocturnal mammalian ground predators. Few studies to date have investigated how roosting sites affect sleep architecture. We compared sleep in captive pigeons on days with and without access to high perches. On the first (baseline) day, low and high perches were available; on the second day, the high perches were removed; and on the third (recovery) day, the high perches were returned. The total time spent sleeping did not vary significantly between conditions; however, the time spent in REM sleep declined on the low-perch night and increased above baseline when the pigeons slept on the high perch during the recovery night. Although the amount of SWS did not vary significantly between conditions, SWS intensity was lower on the low-perch night, particularly early in the night. The similarity of these responses between birds and mammals suggests that REM sleep is influenced by at least some ecological factors in a similar manner in both groups of animals.

Bottom-Up versus Top-Down Induction of Sleep by Zolpidem Acting on Histaminergic and Neocortex Neurons.

Zolpidem, a GABAA receptor-positive modulator, is the gold-standard drug for treating insomnia. Zolpidem prolongs IPSCs to decrease sleep latency and increase sleep time, effects that depend on α2 and/or α3 subunit-containing receptors. Compared with natural NREM sleep, zolpidem also decreases the EEG power, an effect that depends on α1 subunit-containing receptors, and which may make zolpidem-induced sleep less optimal. In this paper, we investigate whether zolpidem needs to potentiate only particular GABAergic pathways to induce sleep without reducing EEG power. Mice with a knock-in F77I mutation in the GABAA receptor γ2 subunit gene are zolpidem-insensitive. Using these mice, GABAA receptors in the frontal motor neocortex and hypothalamic (tuberomammillary nucleus) histaminergic-neurons of γ2I77 mice were made selectively sensitive to zolpidem by genetically swapping the γ2I77 subunits with γ2F77 subunits. When histamine neurons were made selectively zolpidem-sensitive, systemic administration of zolpidem shortened sleep latency and increased sleep time. But in contrast to the effect of zolpidem on wild-type mice, the power in the EEG spectra of NREM sleep was not decreased, suggesting that these EEG power-reducing effects of zolpidem do not depend on reduced histamine release. Selective potentiation of GABAA receptors in the frontal cortex by systemic zolpidem administration also reduced sleep latency, but less so than for histamine neurons. These results could help with the design of new sedatives that induce a more natural sleep. SIGNIFICANCE STATEMENT: Many people who find it hard to get to sleep take sedatives. Zolpidem (Ambien) is the most widely prescribed "sleeping pill." It makes the inhibitory neurotransmitter GABA work better at its receptors throughout the brain. The sleep induced by zolpidem does not resemble natural sleep because it produces a lower power in the brain waves that occur while we are sleeping. We show using mouse genetics that zolpidem only needs to work on specific parts and cell types of the brain, including histamine neurons in the hypothalamus, to induce sleep but without reducing the power of the sleep. This knowledge could help in the design of sleeping pills that induce a more natural sleep.

Reconstruction of vocal interactions in a group of small songbirds.

The main obstacle for investigating vocal interactions in vertebrates is the difficulty of discriminating individual vocalizations of rapidly moving, sometimes simultaneously vocalizing individuals. We developed a method of recording and analyzing individual vocalizations in free-ranging animals using ultraminiature back-attached sound and acceleration recorders. Our method allows the separation of zebra finch vocalizations irrespective of background noise and the number of vocalizing animals nearby.

Sleep-Related Electrophysiology and Behavior of Tinamous (Eudromia elegans): Tinamous Do Not Sleep Like Ostriches.

The functions of slow wave sleep (SWS) and rapid eye movement (REM) sleep, distinct sleep substates present in both mammals and birds, remain unresolved. One approach to gaining insight into their function is to trace the evolution of these states through examining sleep in as many taxonomic groups as possible. The mammalian and avian clades are each composed of two extant groups, i.e., the monotremes (echidna and platypus) and therian (marsupial and eutherian [or placental]) mammals, and Palaeognaths (cassowaries, emus, kiwi, ostriches, rheas, and tinamous) and Neognaths (all other birds) among birds. Previous electrophysiological studies of monotremes and ostriches have identified a unique "mixed" sleep state combining features of SWS and REM sleep unlike the well-delineated sleep states observed in all therian mammals and Neognath birds. In the platypus this state is characterized by periods of REM sleep-related myoclonic twitching, relaxed skeletal musculature, and rapid eye movements, occurring in conjunction with SWS-related slow waves in the forebrain electroencephalogram (EEG). A similar mixed state was also observed in ostriches; although in addition to occurring during periods with EEG slow waves, reduced muscle tone and rapid eye movements also occurred in conjunction with EEG activation, a pattern typical of REM sleep in Neognath birds. Collectively, these studies suggested that REM sleep occurring exclusively as an integrated state with forebrain activation might have evolved independently in the therian and Neognath lineages. To test this hypothesis, we examined sleep in the elegant crested tinamou (Eudromia elegans), a small Palaeognath bird that more closely resembles Neognath birds in size and their ability to fly. A 24-h period was scored for sleep state based on electrophysiology and behavior. Unlike ostriches, but like all of the Neognath birds examined, all indicators of REM sleep usually occurred in conjunction with forebrain activation in tinamous. The absence of a mixed REM sleep state in tinamous calls into question the idea that this state is primitive among Palaeognath birds and therefore birds in general.

Heart rate variability reveals that a decrease in parasympathetic ('rest-and-digest') activity dominates autonomic stress responses in a free-living seabird.

The autonomic stress response, often referred to as the 'fight-or-flight' response, is a highly conserved physiological reaction to stress in vertebrates that occurs via a decrease in parasympathetic (PNS) activity, which promotes self-maintenance 'rest and digest' processes, and an increase in sympathetic (SNS) activity, which prepares an animal for danger ('fight-or-flight'). Though the PNS and SNS both innervate most organs, they often control different tissues and functions within those organs (though the pacemaker of the heart is controlled by both). Moreover the PNS and SNS are regulated independently. Yet until now, most studies of autonomic stress responses in non-model species focused only on the SNS response. We used external electrocardiogram loggers to measure heart rate and heart rate variability indexes that reflect PNS and SNS activity in a seabird, the Streaked Shearwater (Calonectris leucomelas), during the stress of handling, and during recovery in the nest burrow or during restraint in a cloth bag. We show for the first time in a free-living animal that the autonomic stress response is mediated primarily by a rapid decrease in PNS activity: handling stress induced a large and long-lasting depression of PNS 'rest-and-digest' activity that required two hours to recover. We also found evidence for a substantially smaller and shorter-lasting SNS 'fight-or-flight' response. Confinement in a cloth bag was less stressful for birds than handling, but more stressful than recovering in nest burrows. We show that quantifying autonomic activity from heart rate variability is effective for non-invasively studying stress physiology in free-living animals.

Altered activity in the central medial thalamus precedes changes in the neocortex during transitions into both sleep and propofol anesthesia.

How general anesthetics cause loss of consciousness is unknown. Some evidence points toward effects on the neocortex causing "top-down" inhibition, whereas other findings suggest that these drugs act via subcortical mechanisms, possibly selectively stimulating networks promoting natural sleep. To determine whether some neuronal circuits are affected before others, we used Morlet wavelet analysis to obtain high temporal resolution in the time-varying power spectra of local field potentials recorded simultaneously in discrete brain regions at natural sleep onset and during anesthetic-induced loss of righting reflex in rats. Although we observed changes in the local field potentials that were anesthetic-specific, there were some common changes in high-frequency (20-40 Hz) oscillations (reductions in frequency and increases in power) that could be detected at, or before, sleep onset and anesthetic-induced loss of righting reflex. For propofol and natural sleep, these changes occur first in the thalamus before changes could be detected in the neocortex. With dexmedetomidine, the changes occurred simultaneously in the thalamus and neocortex. In addition, the phase relationships between the low-frequency (1-4 Hz) oscillations in thalamic nuclei and neocortical areas are essentially the same for natural sleep and following dexmedetomidine administration, but a sudden change in phase, attributable to an effect in the central medial thalamus, occurs at the point of dexmedetomidine loss of righting reflex. Our data are consistent with the central medial thalamus acting as a key hub through which general anesthesia and natural sleep are initiated.

Staying awake--a genetic region that hinders α2 adrenergic receptor agonist-induced sleep.

How external stimuli prevent the onset of sleep has been little studied. This is usually considered to be a non-specific type of phenomenon. However, the hypnotic drug dexmedetomidine, an agonist at α2 adrenergic receptors, has unusual properties that make it useful for investigating this question. Dexmedetomidine is considered to produce an 'arousable' sleep-like state, so that patients or animals given dexmedetomidine become alert following modest stimulation. We hypothesized that it might be more difficult to make mice unconscious with dexmedetomidine if there was a sufficient external stimulus. Employing a motorized rotating cylinder, which provided a continuous and controlled arousal stimulus, we quantitatively measured the ability of such a stimulus to prevent dexmedetomidine loss of righting reflex in two inbred strains of mice (C57BL/6 and 129X1). We found that whereas the C57BL/6 strain required a strong stimulus to prevent dexmedetomidine-induced hypnosis, the 129X1 strain stayed awake even with minimal stimuli. Remarkably, this could be calibrated as a simple threshold trait, i.e. a binary 'yes-no' response, which after crossing the two mouse strains behaved as a dominant-like trait. We carried out a genome-wide linkage analysis on the F2 progeny to determine if the ability of a stimulus to prevent dexmedetomidine hypnosis could be mapped to one or more chromosomal regions. We identified a locus on chromosome 4 with an associated Logarithm of Odds score exceeding the pre-established threshold level. These results show that complex traits, such as the ability of a stimulus to reverse drug-induced hypnosis, may have precise genetic determinants.

Neurophysiological and behavioral synchronization in group-living and sleeping mice.

Social interactions profoundly influence animal development, physiology, and behavior. Yet, how sleep-a central behavioral and neurophysiological process-is modulated by social interactions is poorly understood. Here, we characterized sleep behavior and neurophysiology in freely moving and co-living mice under different social conditions. We utilized wireless neurophysiological devices to simultaneously record multiple individuals within a group for 24 h, alongside video acquisition. We first demonstrated that mice seek physical contact before sleep initiation and sleep while in close proximity to each other (hereafter, "huddling"). To determine whether huddling during sleep is a motivated behavior, we devised a novel behavioral apparatus allowing mice to choose whether to sleep in close proximity to a conspecific or in solitude, under different environmental conditions. We also applied a deep-learning-based approach to classify huddling behavior. We demonstrate that mice are willing to forgo their preferred sleep location, even under thermoneutral conditions, to gain access to social contact during sleep. This strongly suggests that the motivation for prolonged physical contact-which we term somatolonging-drives huddling behavior. We then characterized sleep architecture under different social conditions and uncovered a social-dependent modulation of sleep. We also revealed coordination in multiple neurophysiological features among co-sleeping individuals, including in the timing of falling asleep and waking up and non-rapid eye movement sleep (NREMS) intensity. Notably, the timing of rapid eye movement sleep (REMS) was synchronized among co-sleeping male siblings but not co-sleeping female or unfamiliar mice. Our findings provide novel insights into the motivation for physical contact and the extent of social-dependent plasticity in sleep.

Sleep and Thermoregulation in Birds: Cold Exposure Reduces Brain Temperature but Has Little Influence on Sleep Time and Sleep Architecture in Jackdaws (Coloeus monedula).

Birds have an electrophysiological sleep state that resembles mammalian rapid-eye-movement (REM) sleep. However, whether its regulation and function are similar is unclear. In the current experiment, we studied REM sleep regulation in jackdaws (Coloeus monedula) by exposing the birds to low ambient temperature, a procedure that selectively suppresses REM sleep in mammals. Eight jackdaws were equipped with electrodes to record brain activity and neck muscle activity and a thermistor to record cortical brain temperature. Recordings covered a three-day period starting with a 24 h baseline day at an ambient temperature of 21 °C, followed by a 12 h cold night at 4 °C, after which the ambient temperature was restored to 21 °C for the remaining recovery period. Cold exposure at night caused a significant drop in brain temperature of 1.4 °C compared to the baseline night. However, throughout the cold night, jackdaws expressed NREM sleep and REM sleep levels that were not significantly different from the baseline. Also, EEG spectral power during NREM sleep was unaffected by cold exposure. Thus, while cold exposure had a clear effect on brain temperature in jackdaws, it did not have the same REM sleep suppressing effect reported for mammals. These findings suggest that the REM-sleep-like state in birds, unlike REM sleep in mammals, is protected against the influence of low temperature.

Semelparous marsupials reduce sleep for sex.

Sleep is a prominent, seemingly universal animal behavior. Although sleep maintains optimal waking performance, the biological drive to sleep may be incompatible with the life history of some species. In a multi-year study on semelparous marsupials in Australia, we provide the first direct evidence of ecological sleep restriction in a terrestrial mammal. Dusky (Antechinus swainsonii) and agile (A. agilis) antechinus have an unusual reproductive strategy characterized by the synchronous death of all males at the end of their only breeding season. Using accelerometry, electrophysiology, and metabolomics, we show that males, but not females, increase their activity during the breeding season by reducing sleep. In a trade-off between the neurophysiological requirements for sleep and evolutionary necessity for reproduction, strong sexual selection might drive males to sacrifice sleep to increase access to fertile females and ultimately maximize their fitness.

A prefrontal-thalamic circuit encodes social information for social recognition.

Social recognition encompasses encoding social information and distinguishing unfamiliar from familiar individuals to form social relationships. Although the medial prefrontal cortex (mPFC) is known to play a role in social behavior, how identity information is processed and by which route it is communicated in the brain remains unclear. Here we report that a ventral midline thalamic area, nucleus reuniens (Re) that has reciprocal connections with the mPFC, is critical for social recognition in male mice. In vivo single-unit recordings and decoding analysis reveal that neural populations in both mPFC and Re represent different social stimuli, however, mPFC coding capacity is stronger. We demonstrate that chemogenetic inhibitions of Re impair the mPFC-Re neural synchronization and the mPFC social coding. Projection pathway-specific inhibitions by optogenetics reveal that the reciprocal connectivity between the mPFC and the Re is necessary for social recognition. These results reveal an mPFC-thalamic circuit for social information processing.

A REM-active basal ganglia circuit that regulates anxiety.

Rapid eye movement (REM) sleep has been hypothesized to promote emotional resilience, but any neuronal circuits mediating this have not been identified. We find that in mice, somatostatin (Som) neurons in the entopeduncular nucleus (EPSom)/internal globus pallidus are predominantly active during REM sleep. This unique REM activity is both necessary and sufficient for maintaining normal REM sleep. Inhibiting or exciting EPSom neurons reduced or increased REM sleep duration, respectively. Activation of the sole downstream target of EPSom neurons, Vglut2 cells in the lateral habenula (LHb), increased sleep via the ventral tegmental area (VTA). A simple chemogenetic scheme to periodically inhibit the LHb over 4 days selectively removed a significant amount of cumulative REM sleep. Chronic, but not acute, REM reduction correlated with mice becoming anxious and more sensitive to aversive stimuli. Therefore, we suggest that cumulative REM sleep, in part generated by the EP → LHb → VTA circuit identified here, could contribute to stabilizing reactions to habitual aversive stimuli.

Months-long tracking of neuronal ensembles spanning multiple brain areas with Ultra-Flexible Tentacle Electrodes.

We introduce Ultra-Flexible Tentacle Electrodes (UFTEs), packing many independent fibers with the smallest possible footprint without limitation in recording depth using a combination of mechanical and chemical tethering for insertion. We demonstrate a scheme to implant UFTEs simultaneously into many brain areas at arbitrary locations without angle-of-insertion limitations, and a 512-channel wireless logger. Immunostaining reveals no detectable chronic tissue damage even after several months. Mean spike signal-to-noise ratios are 1.5-3x compared to the state-of-the-art, while the highest signal-to-noise ratios reach 89, and average cortical unit yields are ~1.75/channel. UFTEs can track the same neurons across sessions for at least 10 months (longest duration tested). We tracked inter- and intra-areal neuronal ensembles (neurons repeatedly co-activated within 25 ms) simultaneously from hippocampus, retrosplenial cortex, and medial prefrontal cortex in freely moving rodents. Average ensemble lifetimes were shorter than the durations over which we can track individual neurons. We identify two distinct classes of ensembles. Those tuned to sharp-wave ripples display the shortest lifetimes, and the ensemble members are mostly hippocampal. Yet, inter-areal ensembles with members from both hippocampus and cortex have weak tuning to sharp wave ripples, and some have unusual months-long lifetimes. Such inter-areal ensembles occasionally remain inactive for weeks before re-emerging.

GABAergic inhibition of histaminergic neurons regulates active waking but not the sleep-wake switch or propofol-induced loss of consciousness.

The activity of histaminergic neurons in the tuberomammillary nucleus (TMN) of the hypothalamus correlates with an animal's behavioral state and maintains arousal. We examined how GABAergic inputs onto histaminergic neurons regulate this behavior. A prominent hypothesis, the "flip-flop" model, predicts that increased and sustained GABAergic drive onto these cells promotes sleep. Similarly, because of the histaminergic neurons' key hub-like place in the arousal circuitry, it has also been suggested that anesthetics such as propofol induce loss of consciousness by acting primarily at histaminergic neurons. We tested both these hypotheses in mice by genetically removing ionotropic GABA(A) or metabotropic GABA(B) receptors from histidine decarboxylase-expressing neurons. At the cellular level, histaminergic neurons deficient in synaptic GABA(A) receptors were significantly more excitable and were insensitive to the anesthetic propofol. At the behavioral level, EEG profiles were recorded in nontethered mice over 24 h. Surprisingly, GABAergic transmission onto histaminergic neurons had no effect in regulating the natural sleep-wake cycle and, in the case of GABA(A) receptors, for propofol-induced loss of righting reflex. The latter finding makes it unlikely that the histaminergic TMN has a central role in anesthesia. GABA(B) receptors on histaminergic neurons were dispensable for all behaviors examined. Synaptic inhibition of histaminergic cells by GABA(A) receptors, however, was essential for habituation to a novel environment.