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Living organisms synchronize their biological activities with the earth’s rotation through the circadian clock, a molecular mechanism that regulates biology and behavior daily. This synchronization factually maximizes positive activities (e.g., social interactions, feeding) during safe periods, and minimizes exposure to dangers (e.g., predation, darkness) typically at night. Beyond basic circadian regulation, some behaviors like sleep have an additional layer of homeostatic control, ensuring those essential activities are fulfilled. While sleep is predominantly governed by the circadian clock, a secondary homeostatic regulator, though not well-understood, ensures adherence to necessary sleep amounts and hints at a fundamental biological function of sleep beyond simple energy conservation and safety. Here we explore sleep regulation across seven Drosophila species with diverse ecological niches, revealing that while circadian-driven sleep aspects are consistent, homeostatic regulation varies significantly. The findings suggest that in Drosophilids, sleep evolved primarily for circadian purposes. The more complex, homeostatically regulated functions of sleep appear to have evolved independently in a species-specific manner, and are not universally conserved. This laboratory model may reproduce and recapitulate primordial sleep evolution. We all feel tired without sleep, but we still don’t know why. Is tiredness a crucial, evolutionarily conserved feature of sleep? Joyce et al. show that some species have circadian but not homeostatic regulation of rest and suggest this is the ancestral drive of sleep evolution.

Here, we present the use of ethoscopes, which are machines for high-throughput analysis of behavior in Drosophila and other animals. Ethoscopes provide a software and hardware solution that is reproducible and easily scalable. They perform, in real-time, tracking and profiling of behavior by using a supervised machine learning algorithm, are able to deliver behaviorally triggered stimuli to flies in a feedback-loop mode, and are highly customizable and open source. Ethoscopes can be built easily by using 3D printing technology and rely on Raspberry Pi microcomputers and Arduino boards to provide affordable and flexible hardware. All software and construction specifications are available at http://lab.gilest.ro/ethoscope.

In all animals, sleep pressure is under continuous tight regulation. It is universally accepted that this regulation arises from a two-process model, integrating both a circadian and a homeostatic controller. Here we explore the role of environmental social signals as a third, parallel controller of sleep homeostasis and sleep pressure. We show that, in Drosophila melanogaster males, sleep pressure after sleep deprivation can be counteracted by raising their sexual arousal, either by engaging the flies with prolonged courtship activity or merely by exposing them to female pheromones. Humans spend one-third of their lifetime sleeping, but why we (and other animals) need to sleep remains an unresolved mystery of biology. Our desire to sleep changes depending on how much sleep we’ve already had. If we’ve had a long nap during the day, we may find it harder to fall asleep at night; conversely, if we stay up all night partying, we’ll have a difficult time staying alert the next day. This change in the pressure to sleep is known as “sleep homeostasis”. Can sleep homeostasis be suppressed? We know that some migratory birds are able to resist sleep while flying over the ocean. In addition, males of an Arctic bird species forgo sleep for courtship during the three-week window every year when females of its species are fertile. These examples suggest that some behavioral or environmental factors may influence sleep homeostasis. Beckwith et al. now show that sexual arousal can disrupt sleep homeostasis in fruit flies. In “blind date” experiments, young male fruit flies were kept in a small tube with female fruit flies, prompting a 24-hour period of courtship and mating. The males went without sleep during that period, and they did not make up for the lost sleep afterward. In other experiments, male fruit flies were kept awake by a robot that disturbed them every time they tried to sleep. After such treatment, the flies normally attempted to nap. But if the sleep-deprived flies were exposed to a chemical emitted by female flies that increased their sexual arousal, they no longer needed to sleep. Overall, the results presented by Beckwith et al. show that sleep is a biological drive that can be overcome under certain conditions. This will be important for sleep researchers to remember, because it means that it’s possible to affect sleep regulation (perhaps by making the animal stressed or aroused) without activating the brain circuits directly involved in regulating sleep.

During sleep, most animal species enter a state of reduced consciousness characterized by a marked sensory disconnect. Yet some processing of the external world must remain intact, given that a sleeping animal can be awoken by intense stimuli (for example, a loud noise or a bright light) or by soft but qualitatively salient stimuli (for example, the sound of a baby cooing or hearing one’s own name 1 – 3 ). How does a sleeping brain retain the ability to process the quality of sensory information? Here we present a paradigm to study the functional underpinnings of sensory discrimination during sleep in Drosophila melanogaster . We show that sleeping vinegar flies, like humans, discern the quality of sensory stimuli and are more likely to wake up in response to salient stimuli. We also show that the salience of a stimulus during sleep can be modulated by internal states. We offer a prototypical blueprint detailing a circuit involved in this process and its modulation as evidence that the system can be used to explore the cellular underpinnings of how a sleeping brain experiences the world. The authors develop a paradigm to study sensory discrimination during sleep in Drosophila melanogaster .

We present ethoscopes, machines for high-throughput analysis of behaviour in Drosophila and other animals. Ethoscopes provide a software and hardware solution that is reproducible and easily scalable; they perform, in real-time, tracking and profiling of behaviour using a supervised machine learning algorithm; they can deliver behaviourally-triggered stimuli to flies in a feedback-loop mode; and they are highly customisable and open source. Ethoscopes can be built easily using 3D printing technology and rely on Raspberry Pi microcomputers and Arduino boards to provide affordable and flexible hardware. All software and construction specifications are available at http://lab.gilest.ro/ethoscope .

Most living organisms have evolved to synchronize their biological activities with the earth’s rotation, a daily regulation of biology and behaviour controlled by an evolutionary conserved molecular machinery known as the circadian clock. For most animals, circadian mechanisms are meant to maximize their exposure to positive activities (e.g.: social interactions, mating, feeding – generally during the day) and minimize their exposure to peril (e.g.: predation, weather, darkness – generally during the night1). On top of circadian regulation, some behaviours also feature a second layer of homeostatic control acting as a fail-safe to ensure important activities are not ignored. Sleep is one of these behaviours: largely controlled by the circadian clock for its baseline appearance, it is at the same time modulated by a – poorly understood – homeostatic regulator ensuring animals obey their species-specific amount of daily sleep2. An evolutionary conserved homeostatic control is often considered the main evidence for a core biological function of sleep beyond the trivial one (that is: keeping us out of trouble by limiting our energy expenditure and exposure to danger3,4) and it is hypothesized that sleep evolved around this mysterious basic biological function. Here we characterize sleep regulation in a group of seven species of the Drosophila genus at key evolutionary distances and representing a variety of ecological niche adaptations. We show that the spontaneous circadian-driven aspects of sleep are conserved among all species but the homeostatic regulation, unexpectedly, is not. We uncover differences in the behavioural, cell-biological and neuro-pharmacological aspects of sleep and suggest that, in Drosophilids, sleep primarily evolved to satisfy a circadian role, keeping animals immobile during dangerous hours of the day. The homeostatic functions of sleep evolved independently, in a species-specific fashion, and are not conserved.

Behaviour is the ultimate output of neural circuit computations, and therefore its analysis is a cornerstone of neuroscience research. However, every animal and experimental paradigm requires different illumination conditions to capture and, in some cases, manipulate specific behavioural features. This means that researchers often develop, from scratch, their own solutions and experimental set-ups. Here, we present OptoPi, an open source, affordable (∼ £600), behavioural arena with accompanying multi-animal tracking software. The system features highly customisable and reproducible visible and infrared illumination and allows for temporally precise optogenetic stimulation. OptoPi acquires images using a Raspberry Pi camera, features motorised LED-based illumination, Arduino control, as well as spectrum and irradiance monitoring to fine-tune illumination conditions with real time feedback. Our open-source software (BIO) can be used to simultaneously track multiple animals while accurately keeping individual animal’s identity both in on-line and off-line modes. We demonstrate the functionality of OptoPi by recording and tracking under different illumination conditions the spontaneous behaviour of larval zebrafish as well as adult Drosophila flies and their first instar larvae, an experimental animal that due to its small size and transparency has classically been hard to track. Further, we showcase OptoPi’s optogenetic capabilities through a series of experiments using transgenic Drosophila larvae.

Natural Killer (NK) cells are innate immune cells that secrete lytic granules to directly kill virus-infected or transformed cells across an immune synapse. However, a major gap in understanding this process is in establishing how lytic granules pass through the mesh of cortical actin known to underlie the NK cell membrane. Research has been hampered by the resolution of conventional light microscopy, which is too low to resolve cortical actin during lytic granule secretion. Here we use two high-resolution imaging techniques to probe the synaptic organisation of NK cell receptors and filamentous (F)-actin. A combination of optical tweezers and live cell confocal microscopy reveals that microclusters of NKG2D assemble into a ring-shaped structure at the centre of intercellular synapses, where Vav1 and Grb2 also accumulate. Within this ring-shaped organisation of NK cell proteins, lytic granules accumulate for secretion. Using 3D-structured illumination microscopy (3D-SIM) to gain super-resolution of ~100 nm, cortical actin was detected in a central region of the NK cell synapse irrespective of whether activating or inhibitory signals dominate. Strikingly, the periodicity of the cortical actin mesh increased in specific domains at the synapse when the NK cell was activated. Two-colour super-resolution imaging revealed that lytic granules docked precisely in these domains which were also proximal to where the microtubule-organising centre (MTOC) polarised. Together, these data demonstrate that remodelling of the cortical actin mesh occurs at the central region of the cytolytic NK cell immune synapse. This is likely to occur for other types of cell secretion and also emphasises the importance of emerging super-resolution imaging technology for revealing new biology.

Scientific results are often presented as ‘surprising’ as if that is a good thing. Is it? And if so, why? What is the value of surprise in science? Discussions of surprise in science have been limited, but surprise has been used as a way of defending the epistemic privilege of experiments over simulations. The argument is that while experiments can ‘confound’, simulations can merely surprise (Morgan, 2005). Our aim in this paper is to show that the discussion of surprise can be usefully extended to thought experiments and theoretical derivations. We argue that in focusing on these features of scientific practice, we can see that the surprise-confoundment distinction does not fully capture surprise in science. We set out how thought experiments and theoretical derivations can bring about surprises that can be disruptive in a productive way, and we end by exploring how this links with their future fertility.

Ocean warming threatens the persistence of tropical corals and the biologically diverse ecosystems they sustain. While field-based studies on heat impact have predominantly focused on quantifying coral bleaching, a symptom of thermal stress, less attention has been given to understanding trends in coral mortality, a critical metric for assessing and predicting the long-term effects of rising temperatures. Consequently, the relationships between varying heat exposures and resultant coral cover changes remain poorly quantified. Such trends are challenging to establish in the Tropical and Subtropical Western Atlantic (TWA), because climate change impacts are compounded by local anthropogenic and natural disturbances. Additionally, many coral communities have already been substantially altered, with those remaining dominated by relatively resilient species. This study addresses this issue by quantifying coral cover loss as a function of maximum Degree Heating Weeks (DHW) exposure using observed in-situ coral cover changes across reefs in the TWA. Of the five locations assessed (the Florida Keys, Dry Tortugas, Puerto Rico, the US Virgin Islands, and the East and West Flower Garden Banks), all exhibited significant declines in mean coral cover with increasing DHW exposure. Rates ranged from 0.3% to 2.4% annual loss in relative mean cover per unit DHW, with spatial variability largely reflecting pre-impact system conditions and variations in the life-history traits of geographically distinct coral assemblages (e.g., Puerto Rico vs East and West Flower Garden Banks). Variation in responses to DHW among species was also observed across locations. By establishing site-specific coral loss parameters, this study contributes to our understanding of how future coral cover may evolve under escalating thermal stress in the TWA. It also provides practical guidance for targeted restoration by identifying species that have fared comparatively well across locations, grounding efforts in what is possible under current and future conditions rather than idealised historical baselines.