Chiara Cirelli - US grants

DESCRIPTION (provided by applicant): Sleep is present in all species where it has been studied, but its functions remain unknown. A sufficient amount of sleep constitutes a fundamental biological need. For example, curtailing the amount of sleep in normal sleepers affects performance, vigilance, memory and health. Like all complex behaviors, sleep is both environmentally modulated and genetically determined. However, the responsible genes have not been discovered. To identify them, we have initiated a genetic screening for short sleepers in the fruit fly Drosophila melanogaster. Mutagenesis screening in Drosophila has helped unraveling cellular mechanisms that are highly conserved across species, e.g. those controlling development, aging, stress memory, and circadian rhythms. Over the past few years, our laboratory and others have shown that fly sleep shares many key features with mammalian sleep. As in mammals, sleep in Drosophila is characterized by increased arousal thresholds and by changes in brain electrical activity. Fly sleep is regulated independent of the circadian clock, is modulated by stimulants and hypnotics, and is affected by age. Also, fly sleep is associated with changes in brain gene expression similar to those observed in mammals. Over the past 3 years, we have screened approximately 8000 mutant lines, most of which carry single-gene mutations. We found that the amount and regulation of sleep are highly conserved: almost all flies sleep between 400 and 800 min/24 hours and show increased sleep duration and continuity after sleep deprivation. We have also identified several short sleeper lines, three of which are particularly interesting. Despite the reduced amount of sleep (<230 min/day), these lines show normal day-time performance and vigilance. When sleep deprived, they recover most of the sleep lost, suggesting that it is biologically important. The short sleep mutation is due to the genomic insertion of a P element whose mobilization reverts them to normal sleep, suggesting a single gene effect. We propose to characterize these three lines genetically, molecularly, and behaviorally. We will manipulate the expression of the genes responsible for the short sleep phenotype, investigate the molecular pathways controlled by these genes, and characterize their impact on performance, memory, circadian rhythms and life span. This research will help to identify the molecular mechanisms regulating the need for sleep and provide novel clues to its functions.

behavioral /social science research tag

DESCRIPTION (provided by applicant): During development, brain circuits undergo extensive remodeling, involving both synaptogenesis and pruning. Adolescence, in particular, is thought to be a sensitive period for synaptic pruning in cortical circuits involved in cognitive functions and emotional regulation. Adolescence is also a sensitive period for the pathophysiology of many psychiatric disorders, presumably due to this extensive synaptic remodeling. Thus, it would be useful to be able to track changes in the number and efficacy of synapses longitudinally and non-invasively during adolescence. New evidence suggests that changes in sleep slow wave activity (SWA), which can be assessed longitudinally and non-invasively using electroencephalography (EEG), may parallel neurodevelopmental changes in cortical synaptic density. To develop SWA as a potential marker of synaptic function during developmental sensitive periods requires an animal model in which: i) anatomical, molecular, and physiological changes in cortical synapses can be evaluated directly;ii) a point- to-point, intra-subject correlation can be established between sleep SWA and direct measures of synaptic number/molecular composition/efficacy. In Aim 1 of this project, we will pursue these goals by performing both chronic EEG recordings and repeated in vivo imaging with two-photon microscopy in transgenic mice that express yellow fluorescent protein in cortical neurons. Moreover, we will measure molecular and electrophysiological markers of synaptic strength in these mice throughout development. In addition to monitoring synaptic remodeling in vivo, it is important to begin investigating which factors can influence it during the sensitive period of adolescence. Since major changes in synaptogenesis/pruning during development are correlated with major changes in sleep/wake patterns, it has been hypothesized that changes in behavioral state may not only reflect, but also affect synaptic remodeling. Consistent with this notion, new evidence in animals and humans shows that, in the adult brain, waking is associated with a net increase in synaptic strength, and sleep with a net decrease, and that SWA reflects molecular and physiological changes in synaptic function brought about by wake and sleep. Aim 2 of this proposal will test the hypothesis that sleep/wake behavior affect synaptic structure/function also during development. Specifically, we will determine whether sleep and waking differentially affect synaptogenesis and synaptic pruning, consistent with their effects on synaptic strength in adults. If successful, Aim 1 will lead the foundation for EEG monitoring of synaptic efficacy during neurodevelopment in human subjects at risk or patient populations as an essential aid for both diagnosis and therapy. Aim 2 will open the way to preventive/therapeutic approaches for influencing synaptogenesis/pruning by stabilizing/adjusting sleep/wake patterns in children. PUBLIC HEALTH RELEVANCE: Adolescence is a sensitive period during which the brain undergoes massive elimination or "pruning" of synapses - the connections among neurons in the brain. Adolescence is also a sensitive period for the onset and progression of many psychiatric disorders. This project seeks to establish whether the slow waves that can be recorded with the electroencephalogram during sleep in children and adolescents can be used as a sensitive, well-tolerated indicator of the number and strength of synapses. Moreover, this project will establish whether sleep can affect synaptic pruning during development. If so, changes in sleep and waking routines could help both the prevention and the therapy of mental illness.

DESCRIPTION (provided by applicant): Synaptic plasticity is a fundamental feature of the nervous system that underlies neural development, adaptation and learning. There is growing evidence that deficits in the mechanisms of synaptic plasticity are involved in the pathophysiology of many psychiatric disorders, from schizophrenia to mood disorders. For this reason, NIMH has established as one of his strategic research priorities the study of brain plasticity at the cellular, synaptic, circuit, and behavioral level, with the final goal of determining the neurobiological bases of these processes. This proposal will study humans and three animal models (flies, mice, rats) to test the novel and provocative idea that synaptic plasticity is adaptive up to a point, but beyond that point, or in vulnerable individuals, it can become maladaptive. The cost of synaptic plasticity is not often considered but may be crucial in the pathophysiology of psychiatric disorders, and will be assessed at the ultrastructural, cellular, circuit, and behavioral level. Our previous NIMH-funded work has established that the overall result of wake plasticity is a net increase in synaptic strength, which is renormalized by sleep. But what happens when plasticity is excessive, for instance because it is extended beyond the physiological range without intervening sleep? Based on preliminary results obtained in both animals and humans, we hypothesize that extended plasticity can lead to negative consequences on neuronal activity (OFF periods, performance deficits) and on cellular function/integrity (cellular stress, ultrastructural abnormalities). Aim 1 will use rats to test whether plasticity-dependent synaptic overload leads to the occurrence of neuronal OFF periods, local EEG slowing during wake, and performance impairment. It will also establish to what extent these effects are a region- specific consequence of plasticity, rather than a general effect of prolonged wake. Aim 2 will use high density (hd) EEG in humans to ask whether the local increase in EEG theta waves, which occurs during wake as a result of extended plasticity in specific brain circuits, leads to local performance deficits, locally increased sleep need, and o sleep-dependent restoration of function. Aim 3 will use flies and mice to test whether extending plasticity by prolonging wakefulness leads to cellular stress and subcellular damage, and whether doing so chronically under sleep restriction conditions leads to lasting cellular damage and cognitive deficits. Plasticity plays a central role in the life of every organism, but its coston neural structure and function may be substantial especially at vulnerable developmental times, such as adolescence, or in vulnerable populations, such as psychiatric patients. Demonstrating the cost of plasticity at the cellular and systems level will have clear practical implications forthe prevention and treatment of mental disorders.

ABSTRACT Sleep is thought to be essential to restore brain functions, and converging evidence suggests that a key function may be to rebalance cellular changes triggered by plasticity during wake. This evidence is consistent with the hypothesis that sleep and wake may occur, be regulated, and perform their functions at the level of individual neurons. Recently, using multi-array recordings in freely moving rats, we obtained direct evidence that sleep can occur locally within a group of cortical neurons, while the rest of the brain remains awake, and that such local sleep increases with the duration of wake. If so, many questions arise: does local sleep also occur in species very different from mammals, such as flies? Are the mechanisms underlying the occurrence of local sleep similar to those that are known to regulate sleep need, specifically intense neural plasticity leading to tiredness, which requires sleep to enforce synaptic renormalization? Or does local sleep reflect temporary neuronal fatigue due to intense neural activity and short-lasting depletion of energy or calcium stores, and thus not qualify properly as sleep? Finally, it is unknown whether there are cellular/ultrastructural signatures that sleep has occurred and presumably performed its functions. In this proposal, we will test whether local sleep exists in flies by using in vivo calcium imaging to monitor simultaneously the activity of dozens of neurons in specific neuronal circuits. We will then test whether local sleep, like sleep proper, is regulated by intense synaptic plasticity (tiredness) or instead by mere activity (fatigue). To do so we will first establish if local sleep increases during extended wake in a complex enriched environment (fly mall), expected to lead to tiredness in the mushroom bodies, relative to extended wake in an impoverished environment (single tube). Next, we will induce local dTrpA1-mediated activation during extended wake in single tubes, as well as during sleep. These 2 conditions of intense activity with little plasticity are expected to lead to fatigue, and their effects on local sleep will again be compared with those of extended wake in single tubes. To further decouple the effects of plasticity from those of activity we will repeat the experiments in learning mutants, in which exposure to the fly mall should not lead to plasticity/tiredness. Finally, we will use SBF-SEM to test whether there are ultrastructural signatures that can distinguish neurons of flies that have been awake from those of flies that slept, and compare the effects of wake with plasticity leading to tiredness with those of wake associated with dTrpA1-mediated intense firing leading to fatigue. Altogether, these studies in flies will complement those in mice in Project II, which use similar or the very same methods. Together, they will establish if sleep and wake are regulated homeostatically at the single neuron level, and if they leave ultrastructural signatures that reflect their consequences and functions for individual cells.

SUMMARY The SBF-SEM Analysis Core will be directed by Dr. Chiara Cirelli and will provide the hardware, software, and technical personnel resources to assist Projects I (flies) and II (mice) in the analysis of ultrastructural (SBF- SEM) data.

Summary The 2018 meeting will be the third GRC on ?Sleep Regulation and Function?, the second Sleep GRC to include a GRS, and the first conference after we were promoted from a GRC-sponsored meeting to a continuing GRC. As in 2014 and 2016, the 2018 GRC/GRS will be held at the GRC site in Galveston TX. There are many large meetings for sleep professionals, but sleep GRC/GRS clearly meet a widely-felt need: they are the only existing small-format conference focused broadly on the latest basic sleep research, with the specific goal of fostering forward-thinking and innovative approach to sleep science. Both previous sleep GRCs scored in the highest-performing group among all GRCs held in their respective year, with >85% of respondents ranking the meetings as ?excellent? in all categories. Participation was outstanding (199 applications accepted for 2014, 233 for 2016), with 22% (2014) and 25% (2016) of attendees coming from Europe and Asia. Sleep GRCs contributed significantly to grow the next generation of sleep researchers: trainees were 30% of all attendees in 2014 and 40% in 2016. Sleep GRCs also strongly promoted diversity: women represented 44% (2014) and 49% (2016) of all attendees. We believe this success will continue in 2018. The Chair (Dr. Chiara Cirelli, U Wisconsin ? Madison), Vice-Chair (Dr. Paul Shaw, Washington U - St. Louis), and the Steering Committee of 12 eminent sleep researchers planned an outstanding program that spans the breath of the field and reaches out to outside experts, including the Keynote Speaker, Dr. Cristina Alberini (NYU) and Drs Matthew Colonnese (George Washington U) and Heiko Luhmann (U Mainz Germany). None of them has studied sleep per se, but their work on memory consolidation and on the maturation of brain activity soon after birth, when sleep is the predominant behavior, is highly relevant for many of us. The other 7 sessions will discuss the evolving roles of catecholamines in sleep/wake regulation, the cellular and systems? costs of wake for brain and body, the new paradoxes of REM sleep, the global and local determinants of sleep need, the consequences of enhancing/disrupting specific sleep rhythms, the metabolic and vascular effects of sleep disruption. These topics were specifically chosen because thought-provoking and, in line with the subtitle for the 2018 meeting, they are expected to trigger an intense debate on the ?How and Why? of sleep, with the goal of leading to new insights. In addition to including a GRS, the 2018 GRC program will strongly promote the training of young sleep researchers by including 8 early career scientists as GRC speakers or discussion leaders, and 7 trainees (graduate students and post-docs) as GRC speakers. Both Chair and Keynote speaker are women, and so are 38% of the speakers and 78% of the Discussion leaders (~33% for both in 2016). Sleep research is thriving, unraveling the specific mechanisms by which sleep benefits brain and body, from learning and memory to metabolic and cardiovascular function. We expect this meeting to be a crucial milestone that helps the field to move forward in an open-minded and innovative manner.