Sepsis is a serious medical condition in which immune dysfunction plays a key role. Previous treatments focused on chemotherapy to control immune function; however, a recognized effective compound or treatment has yet to be developed. Recent advances indicate that a neuromodulation approach with nerve stimulation allows developing a therapeutic strategy to control inflammation and improve organ functions in sepsis. As a quick, non-invasive technique of peripheral nerve stimulation, acupuncture has emerged as a promising therapy to provide significant advantages for immunomodulation in acute inflammation. Acupuncture obtains its regulatory effect by activating the somatic-autonomic-immune reflexes, including the somatic-sympathetic-splenic reflex, the somatic-sympathetic-adrenal reflex, the somatic-vagal-splenic reflex and the somatic-vagal-adrenal reflex, which produces a systemic effect. The peripheral nerve stimulation also induces local reflexes such as the somatic-sympathetic-lung-reflex, which then produces local effects. These mechanisms offer scientific guidance to design acupuncture protocols for immunomodulation and inflammation control, leading to an evidence-based comprehensive therapy recommendation.

Modern morphological and structural studies are coming to a new level by incorporating the latest methods of three-dimensional electron microscopy (3D-EM). One of the key problems for the wide usage of these methods is posed by difficulties with sample preparation, since the methods work poorly with heterogeneous (consisting of tissues different in structure and in chemical composition) samples and require expensive equipment and usually much time. We have developed a simple protocol allows preparing heterogeneous biological samples suitable for 3D-EM in a laboratory that has a standard supply of equipment and reagents for electron microscopy. This protocol, combined with focused ion-beam scanning electron microscopy, makes it possible to study 3D ultrastructure of complex biological samples, e.g., whole insect heads, over their entire volume at the cellular and subcellular levels. The protocol provides new opportunities for many areas of study, including connectomics.

Laboratory behavioural tasks are an essential research tool. As questions asked of behaviour and brain activity become more sophisticated, the ability to specify and run richly structured tasks becomes more important. An increasing focus on reproducibility also necessitates accurate communication of task logic to other researchers. To these ends we developed pyControl, a system of open source hardware and software for controlling behavioural experiments comprising; a simple yet flexible Python-based syntax for specifying tasks as extended state machines, hardware modules for building behavioural setups, and a graphical user interface designed for efficiently running high throughput experiments on many setups in parallel, all with extensive online documentation. These tools make it quicker, easier and cheaper to implement rich behavioural tasks at scale. As important, pyControl facilitates communication and reproducibility of behavioural experiments through a highly readable task definition syntax and self-documenting features.

Genetically encoded Ca indicators (GECIs) enable long-term monitoring of cellular and subcellular dynamics of this second messenger in response to environmental and developmental cues without relying on exogenous dyes. Continued development and optimization in GECIs, combined with advances in gene manipulation, offer new opportunities for investigating the mechanism of Ca signaling in fungi, ranging from documenting Ca signatures under diverse conditions and genetic backgrounds to evaluating how changes in Ca signature impact calcium-binding proteins and subsequent cellular changes. Here, we attempted to express multi-color (green, yellow, blue, cyan, and red) circularly permuted fluorescent protein (FP)-based Ca indicators driven by multiple fungal promoters in Fusarium oxysporum, F. graminearum, and Neurospora crassa. Several variants were successfully expressed, with GCaMP5G driven by the Magnaporthe oryzae ribosomal protein 27 (P) and F. verticillioides elongation factor-1α (P) gene promoters being optimal for F. graminearum and F. oxysporum, respectively. Transformants expressing GCaMP5G were compared with those expressing YC3.60, a ratiometric Cameleon Ca indicator. Wild-type and three Ca signaling mutants of F. graminearum expressing GCaMP5G exhibited improved signal-to-noise and increased temporal and spatial resolution and are also more amenable to studies involving multiple FPs compared to strains expressing YC3.60.

Recording of the electrical activity from one of the smallest cells of a mammalian organism- a sperm cell- has been a challenging task for electrophysiologists for many decades. The method known as "spermatozoan patch clamp" was introduced in 2006. It has enabled the direct recording of ion channel activity in whole-cell and cell-attached configurations and has been instrumental in describing sperm cell physiology and the molecular identity of various calcium, potassium, sodium, chloride, and proton ion channels. However, recording from single spermatozoa requires advanced skills and training in electrophysiology. This detailed protocol summarizes the step-by-step procedure and highlights several 'tricks-of-the-trade' in order to make it available to anyone who wishes to explore the fascinating physiology of the sperm cell. Specifically, the protocol describes recording from human and murine sperm cells but can be adapted to essentially any mammalian sperm cell of any species. The protocol covers important details of the application of this technique, such as isolation of sperm cells, selection of reagents and equipment, immobilization of the highly motile cells, formation of the tight (Gigaohm) seal between a recording electrode and the plasma membrane of the sperm cells, transition into the whole-spermatozoan mode (also known as break-in), and exemplary recordings of the sperm cell calcium ion channel, CatSper, from six mammalian species. The advantages and limitations of the sperm patch clamp method, as well as the most critical steps, are discussed.

Liquid-liquid phase separation (LLPS) has emerged in recent years as an important physicochemical process for organizing diverse processes within cells via the formation of membraneless organelles termed biomolecular condensates. Emerging evidence now suggests that the formation and regulation of biomolecular condensates are also intricately linked to cancer formation and progression. We review the most recent literature linking the existence and/or dissolution of biomolecular condensates to different hallmarks of cancer formation and progression. We then discuss the opportunities that this condensate perspective provides for cancer research and the development of novel therapeutic approaches, including the perturbation of condensates by small-molecule inhibitors.

Many animals use coordinated limb movements to interact with and navigate through the environment. To investigate circuit mechanisms underlying locomotor behavior, we used serial-section electron microscopy (EM) to map synaptic connectivity within a neuronal network that controls limb movements. We present a synapse-resolution EM dataset containing the ventral nerve cord (VNC) of an adult female Drosophila melanogaster. To generate this dataset, we developed GridTape, a technology that combines automated serial-section collection with automated high-throughput transmission EM. Using this dataset, we reconstructed 507 motor neurons, including all those that control the legs and wings. We show that a specific class of leg sensory neurons directly synapse onto the largest-caliber motor neuron axons on both sides of the body, representing a unique feedback pathway for fast limb control. We provide open access to the dataset and reconstructions registered to a standard atlas to permit matching of cells between EM and light microscopy data. We also provide GridTape instrumentation designs and software to make large-scale EM data acquisition more accessible and affordable to the scientific community.

Anisotropic environments can drastically alter the spectroscopy and photochemistry of molecules, leading to complex structure-function relationships. We examined this using fluorescent proteins as easy-to-modify model systems. Starting from a single scaffold, we have developed a range of 27 photochromic fluorescent proteins that cover a broad range of spectroscopic properties, including the determination of 43 crystal structures. Correlation and principal component analysis confirmed the complex relationship between structure and spectroscopy, but also allowed us to identify consistent trends and to relate these to the spatial organization. We find that changes in spectroscopic properties can come about through multiple underlying mechanisms, of which polarity, hydrogen bonding and presence of water molecules are key modulators. We anticipate that our findings and rich structure/spectroscopy dataset can open opportunities for the development and evaluation of new and existing protein engineering methods.

Changes in walking speed are characterized by changes in both the animal's gait and the mechanics of its interaction with the ground. Here we study these changes in walking . We measured the fly's center of mass (CoM) movement with high spatial resolution and the position of its footprints. Flies predominantly employ a modified tripod gait that only changes marginally with speed. The mechanics of a tripod gait can be approximated with a simple model - angular and radial spring-loaded inverted pendulum (ARSLIP) - which is characterized by two springs of an effective leg that become stiffer as the speed increases. Surprisingly, the change in the stiffness of the spring is mediated by the change in tripod shape rather than a change in stiffness of the individual leg. The effect of tripod shape on mechanics can also explain the large variation in kinematics among insects, and ARSLIP can model these variations.

Animals evolved in complex environments, producing a wide range of behaviors, including navigation, foraging, prey capture, and conspecific interactions, which vary over timescales ranging from milliseconds to days. Historically, these behaviors have been the focus of study for ecology and ethology, while systems neuroscience has largely focused on short timescale behaviors that can be repeated thousands of times and occur in highly artificial environments. Thanks to recent advances in machine learning, miniaturization, and computation, it is newly possible to study freely moving animals in more natural conditions while applying systems techniques: performing temporally specific perturbations, modeling behavioral strategies, and recording from large numbers of neurons while animals are freely moving. The authors of this review are a group of scientists with deep appreciation for the common aims of systems neuroscience, ecology, and ethology. We believe it is an extremely exciting time to be a neuroscientist, as we have an opportunity to grow as a field, to embrace interdisciplinary, open, collaborative research to provide new insights and allow researchers to link knowledge across disciplines, species, and scales. Here we discuss the origins of ethology, ecology, and systems neuroscience in the context of our own work and highlight how combining approaches across these fields has provided fresh insights into our research. We hope this review facilitates some of these interactions and alliances and helps us all do even better science, together.