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Summary All-optical physiology (AOP) manipulates and reports neuronal activities with light, allowing for interrogation of neuronal functional connections with high spatiotemporal resolution. However, contemporary high-speed AOP platforms are limited to single-depth or discrete multi-plane recordings that are not suitable for studying functional connections among densely packed small neurons, such as neurons in Drosophila brains. Here, we constructed a 3D AOP platform by incorporating single-photon point stimulation and two-photon high-speed volumetric recordings with a tunable acoustic gradient-index (TAG) lens. We demonstrated the platform effectiveness by studying the anterior visual pathway (AVP) of Drosophila. We achieved functional observation of spatiotemporal coding and the strengths of calcium-sensitive connections between anterior optic tubercle (AOTU) sub-compartments and >70 tightly assembled 2-μm bulb (BU) microglomeruli in 3D coordinates with a single trial. Our work aids the establishment of in vivo 3D functional connectomes in neuron-dense brain areas., Graphical Abstract, Highlights • All-optical volumetric physiology = precise stimulation + fast volumetric recording • Precise single-photon point stimulation among genetically defined neurons • 3D two-photon imaging by an acoustic gradient-index lens for dense neural structures • Observation of 3D functional connectivity in Drosophila anterior visual pathway, Neuroscience; Techniques in Neuroscience; Biological Sciences Research Methodologies; Biological Sciences Tools

Summary Precise optogenetic control, ideally down to single cells in dense cell populations, is essential in understanding the heterogeneity of cell networks. Devices with such capability, if built in a chip scale, will advance optogenetic studies at cellular levels in a variety of experimental settings. Here we demonstrate optogenetic control of intracellular Ca2+ dynamics at the single cell level using a 16-μm pitched micro-light emitting diode (LED) array that features high brightness, small spot size, fast response, and low voltage operation. Individual LED pixels are able to reliably trigger intracellular Ca2+ transients, confirmed by fluorescence microscopy and control experiments and cross-checked by two genetically coded Ca2+ indicators. Importantly, our array can optogenetically address individual cells that are sub-10 μm apart in densely packed cell populations. These results suggest the possible use of the micro-LED array toward a lab-on-a-chip for single-cell optogenetics, which may allow for pharmaceutical screening and fundamental studies on a variety of cell networks., Graphical Abstract, Highlights • Precise optogenetic control of Ca2+ signaling down to the single cell level • Bright, localized optogenetic stimulus with a high-density micro-LED array • Advancing micro-LED arrays toward a lab-on-a-chip for single-cell optogenetics, Techniques in Genetics; Cellular Neuroscience; Techniques in Neuroscience; Bioelectronics; Electronic Materials

Summary Since the generation of cell-type specific knockout models, the importance of inter-cellular communication between neural, vascular, and microglial cells during neural development has been increasingly appreciated. However, the extent of communication between these major cell populations remains to be systematically mapped. Here, we describe EMBRACE (embryonic brain cell extraction using FACS), a method to simultaneously isolate neural, mural, endothelial, and microglial cells to more than 94% purity in ∼4 h. Utilizing EMBRACE we isolate, transcriptionally analyze, and build a cell-cell communication map of the developing mouse brain. We identify 1,710 unique ligand-receptor interactions between neural, endothelial, mural, and microglial cells in silico and experimentally confirm the APOE-LDLR, APOE-LRP1, VTN-KDR, and LAMA4-ITGB1 interactions in the E14.5 brain. We provide our data via the searchable “Brain interactome explorer”, available at https://mpi-ie.shinyapps.io/braininteractomeexplorer/. Together, this study provides a comprehensive map that reveals the richness of communication within the developing brain., Graphical Abstract, Highlights • Isolation of embryonic neural, mural, endothelial, and microglial cells to >94% purity • Transcriptome analyses of neural, vascular, and microglial cells from E14.5 brain • Generation of inter-cellular communication network with 1,710 unique interactions • Established “Brain interactome explorer,” a searchable cell communication database, Neuroscience; Developmental Neuroscience; Systems Neuroscience; Techniques in Neuroscience

Summary Recently, many super-resolution technologies have been demonstrated, significantly affecting biological studies by observation of cellular structures down to nanometer precision. However, current super-resolution techniques mostly rely on wavefront engineering or wide-field imaging of signal blinking or fluctuation, and thus imaging depths are limited due to tissue scattering or aberration. Here we present a technique that is capable of imaging through an intact Drosophila brain with 20-nm lateral resolution at ∼200 μm depth. The spatial resolution is provided by molecular localization of a photoconvertible fluorescent protein Kaede, whose red form is found to exhibit blinking state. The deep-tissue observation is enabled by optical sectioning of spinning disk microscopy, as well as reduced scattering from optical clearing. Together these techniques are readily available for many biologists, providing three-dimensional resolution of densely entangled dendritic fibers in a complete Drosophila brain. The method paves the way toward whole-brain neural network studies and is applicable to other high-resolution bioimaging., Graphical Abstract, Highlights • Spatial resolution of 20 nm at ∼ 200 μm imaging depth (may be extended) • Super-resolution tracking of densely entangled neural fibers in an intact brain • All tools (FP, confocal, clearing, localization) are commercially available, Optical Imaging; Neuroscience; Techniques in Neuroscience

Summary The development of optical methods to activate neurons with single-cell resolution has enabled systematic mapping of inhibitory connections. In contrast, optical mapping of excitatory connections between pyramidal neurons (PCs) has been a major challenge due to their high densities in cortical tissue and their weak and stochastic connectivity. Here we present an optogenetic two-photon mapping method in mouse neocortical slices by activating PCs with the red-shifted opsin C1V1 while recording postsynaptic responses in whole-cell configuration. Comparison of delays from triggered action potentials (APs) with those from synaptic inputs allowed us to predict connected PCs in three dimensions. We confirmed these predictions with paired recordings, and used this method to map strong connections among large populations of layer 2/3 PCs. Our method can be used for fast, systematic mapping of synaptic connectivity and weights., Graphical Abstract, Highlights • Two-photon optogenetic mapping of excitatory connectivity and strength in neocortex • Identification of connected neurons in acute slices through numerical optimization • Synaptic delays align with location of connected presynaptic cell • Confirmation of predicted connections by dual patch-clamp recordings, Optical Imaging; Systems Neuroscience; Techniques in Neuroscience

Summary A core goal in cognitive neuroscience is identifying the physical substrates of the patterns of thought that occupy our daily lives. Contemporary views suggest that the landscape of ongoing experience is heterogeneous and can be influenced by features of both the person and the context. This perspective piece considers recent work that explicitly accounts for both the heterogeneity of the experience and context dependence of patterns of ongoing thought. These studies reveal that systems linked to attention and control are important for organizing experience in response to changing environmental demands. These studies also establish a role of the default mode network beyond task-negative or purely episodic content, for example, implicating it in the level of vivid detail in experience in both task contexts and in spontaneous self-generated experiential states. Together, this work demonstrates that the landscape of ongoing thought is reflected in the activity of multiple neural systems, and it is important to distinguish between processes contributing to how the experience unfolds from those linked to how these experiences are regulated., Cognitive Neuroscience; Psychology; Techniques in Neuroscience

Summary Automated seizure detection in long-term video-EEG recordings is far from being integrated into common clinical practice. Here, we leverage classical and state-of-the-art complexity measures to robustly and automatically detect seizures from scalp recordings. Brain activity is scored through eight features, encompassing traditional time domain and novel measures of recurrence. A binary classification algorithm tailored to treat unbalanced dataset is used to determine whether a time window is ictal or non-ictal from its features. The application of the algorithm on a cohort of ten adult patients with focal refractory epilepsy indicates sensitivity, specificity, and accuracy of 90%, along with a true alarm rate of 95% and less than four false alarms per day. The proposed approach emphasizes ictal patterns against noisy background without the need of data preprocessing. Finally, we benchmark our approach against previous studies on two publicly available datasets, demonstrating the good performance of our algorithm., Graphical Abstract, Highlights • Complexity measures are formulated to enhance classical time-domain statistics of EEG • The detection algorithm does not need ad-hoc data preprocessing to address artifacts • Focal seizures are detected 95% of the time with less than four false alarms per day • The approach offers a visual representation of a seizure as a time-evolving network, Computer Application in Medicine; Computer-Aided Diagnosis Method; Clinical Neuroscience; Techniques in Neuroscience; Algorithms

Summary In the mammalian cerebral neocortex, different regions have different cytoarchitecture, neuronal birthdates, and functions. In most regions, neuronal migratory profiles are speculated similar based on observations using thymidine analogs. Few reports have investigated regional migratory differences from mitosis at the ventricular surface. In this study, we applied FlashTag technology, in which dyes are injected intraventricularly, to describe migratory profiles. We revealed a mediolateral regional difference in the migratory profiles of neurons that is dependent on developmental stage; for example, neurons labeled at embryonic day 12.5–15.5 reached their destination earlier dorsomedially than dorsolaterally, even where there were underlying ventricular surfaces, reflecting sojourning below the subplate. This difference was hardly recapitulated by thymidine analogs, which visualize neurogenic gradients, suggesting a biological significance different from the neurogenic gradient. These observations advance our understanding of cortical development and the power of FlashTag in studying migration and are thus resources for future neurodevelopmental studies., Graphical abstract, Highlights • FlashTag visualized mediolateral regional differences of cortical migratory profiles • Mediolateral differences were observed when neurons were labeled at E12.5–15.5 • Late-born neurons transiently sojourned below the dorsolateral subplate (SP) cells • The difference was unclear in reeler cortex, where SP cells position superficially, Biological Sciences ; Neuroscience ; Developmental Neuroscience ; Neuroanatomy ; Techniques in Neuroscience

Summary Resin embedding is widely used to dissect the fine structure of bio-tissue with electron and optical microscopy. However, it is difficult to embed large-volume tissues with resin. Here, we modified the formula of LR-White resin to prevent the sample cracking during polymerization process and applied this method to the intact brains of mouse, ferret, and macaque. Meanwhile, we increased the fluorescence preservation rate for green fluorescent protein (GFP) from 73 ± 4.0% to 126 ± 3.0% and tdTomato from 60 ± 3.3% to 117 ± 2.8%. Combined with the whole-brain imaging system, we acquired the cytoarchitectonic information and the circuit information such as individual axon and boutons which were labeled with multiple fluorescent proteins. This method shows great potential in the study of continuous fine microstructure information in large-volume tissues from different species, which can facilitate the neuroscience research and help the understanding of the structure-function relationship in complex bio-tissues., Graphical Abstract, Highlights • Modified LR-White resin embedding was proposed to embed large-volume tissues • Retarder α-methyl-styrene was added to prevent cracking during polymerization • Resin formula was modified to preserve multiple fluorescent proteins • Microstructure information was acquired from the brains of different species, Techniques in Neuroscience; Tissue Engineering; Biomedical Materials

Summary Optical clearing has emerged as a powerful tool for volume imaging. Although volume imaging with immunostaining have been successful in many protocols, yet obtaining homogeneously stained thick samples remains challenging. Here, we propose a method for label-free imaging of brain slices by enhancing the regional heterogeneity of the optical properties using the tissue clearing principle. We used FxClear, a method for delipidation of brain tissue, to retain a larger proportion of lipids at the white matter (WM). Furthermore, the embedding media affected the contrasts for the lipid-rich or extracellular matrix-rich areas, depending on their chemical properties. Thus, we tailored clearing conditions for the enhancement of the refractive indices (RIs) differences between gray and WM, or several pathological features. RI differences can be imaged using conventional light microscopy or optical coherence tomography. We propose that our protocol is simple, reliable, and flexible for label-free imaging, easily implementable to routine histology laboratory., Graphical abstract, Highlights • Label-free imaging via enhancing RI differences in optically cleared brain • Recognition of lipid- or ECM-rich regions using tailored protocols • Easy and affordable protocols using conventional optics, Physiology; Techniques in Neuroscience; Biology Experimental Methods

Summary In vivo two-photon deep imaging with a broad field of view has revealed functional connectivity among brain regions. Here, we developed a novel observation method that utilizes a polyethylene-oxide-coated CYTOP (PEO-CYTOP) nanosheet with a thickness of ∼130 nm that exhibited a water retention effect and a hydrophilized adhesive surface. PEO-CYTOP nanosheets firmly adhered to brain surfaces, which suppressed bleeding from superficial veins. By taking advantage of the excellent optical properties of PEO-CYTOP nanosheets, we performed in vivo deep imaging in mouse brains at high resolution. Moreover, PEO-CYTOP nanosheets enabled to prepare large cranial windows, achieving in vivo imaging of neural structure and Ca2+ elevation in a large field of view. Furthermore, the PEO-CYTOP nanosheets functioned as a sealing material, even after the removal of the dura. These results indicate that this method would be suitable for the investigation of neural functions that are composed of interactions among multiple regions., Graphical Abstract, Highlights • PEO-CYTOP nanosheet enables in vivo deep brain imaging in a vast field of view • The 130 nm thickness and the hydrophilized surface realize the strong adhesiveness • Suppressions of bleeding from the surface and inflammation in long-term are achieved • The vast and transparent cranial window with natural curvature of the surface, Optical Imaging; Techniques in Neuroscience; Nanomaterials

Wearable devices that record brain signals may present privacy concerns for consumers. Industry leaders discussed four such concerns with us that pertain to data collection and management, user autonomy and information transparency, exceptionalism of brain data, and regulatory systems., Graphical Abstract, Techniques in Neuroscience; Biotechnology; Bioethics; Bioengineering; Biodevices

Summary Fibronectin intrabodies generated with mRNA display (FingRs) are a recently developed tool for labeling excitatory or inhibitory synapses, with the benefit of not altering endogenous synaptic protein expression levels or synaptic transmission. Here, we generated a viral vector FingR toolbox that allows for multi-color, neuron-type-specific labeling of excitatory or inhibitory synapses in multiple brain regions. We screened various fluorophores, FingR fusion configurations, and transcriptional control regulations in adeno-associated virus (AAV) and retrovirus vector designs. We report the development of a red FingR variant and demonstrated dual labeling of excitatory and inhibitory synapses in the same cells. Furthermore, we developed cre-inducible FingR AAV variants and demonstrated their utility, finding that the density of inhibitory synapses in aspiny striatal cholinergic interneurons remained unchanged in response to dopamine depletion. Finally, we generated FingR retroviral vectors, which enabled us to track the development of excitatory and inhibitory synapses in hippocampal adult-born granule cells., Graphical Abstract, Highlights • AAV FingR vectors globally label excitatory or inhibitory synapses in neurons • AAV FingRs dually label excitatory and inhibitory synapses in the same neuron • Cre-inducible FingR AAV vectors label synapses in defined neuron populations • Retroviral FingR vectors allow tracking of synaptic development in newborn neurons, Biological Sciences; Neuroscience; Techniques in Neuroscience

Summary Elucidating fine architectures and functions of cellular and synaptic connections requires development of new flexible methods. Here, we created a concept called the “battle of transgenes,” based on which we generated strategies using genetically engineered battles of multiple recombinases. The strategies enabled split-tunable allocation of multiple transgenes. We demonstrated the versatility of these strategies and technologies in inducing strong and multi-sparse allocations of multiple transgenes. Furthermore, the combination of our transgenic strategy and expansion microscopy enabled three-dimensional high-resolution imaging of whole synaptic structures in the hippocampus with simultaneous visualizations of endogenous synaptic proteins. These strategies and technologies based on the battle of genes may accelerate the analysis of whole synaptic and cellular connections in diverse life science fields., Graphical Abstract, Highlights • Generation of BATTLE-recombinase systems for allocation of multiple transgenes • Split-tunable allocation in BATTLE-1 and multi-sparse allocation in BATTLE-2 • Clear and strong labeling of dendrites and axons using BATTLE-2 • 3D high-resolution imaging of whole synapses in hippocampus in BATTLE-1EX, Molecular Biology; Molecular Neuroscience; Neuroanatomy; Techniques in Neuroscience

Summary Implanted electrodes provide one of the most important neurotechniques for fundamental and translational neurosciences by permitting time-resolved electrical detection of individual neurons in vivo. However, conventional rigid electrodes typically cannot provide stable, long-lasting recordings. Numerous interwoven biotic and abiotic factors at the tissue-electrode interface lead to short- and long-term instability of the recording performance. Making neural electrodes flexible provides a promising approach to mitigate these challenges on the implants and at the tissue-electrode interface. Here we review the recent progress of ultraflexible neural electrodes and discuss the engineering principles, the material properties, and the implantation strategies to achieve stable tissue-electrode interface and reliable unit recordings in living brains., Graphical Abstract, Techniques in Neuroscience; Bioelectronics; Bioelectrical Engingeering

Summary Three-dimensional (3D) in vitro cultures recapitulate key features of the brain including morphology, cell-cell and cell-extracellular matrix interactions, gradients of factors, and mechanical properties. However, there remains a need for experimental and computational tools to investigate network functions in these 3D models. To address this need, we present an experimental system based on 3D scaffold-based cortical neuron cultures in which we expressed the genetically encoded calcium indicator GCaMP6f to record neuronal activity at the millimeter-scale. Functional neural network descriptors were computed with graph-theory-based network analysis methods, showing the formation of functional networks at 3 weeks of culture. Changes to the functional network properties upon perturbations to glutamatergic neurotransmission or GABAergic neurotransmission were quantitatively characterized. The results illustrate the applicability of our 3D experimental system for the study of brain network development, function, and disruption in a biomimetic microenvironment., Graphical Abstract, Highlights • 3D biomimetic in vitro cultures engineered to study functional neural networks • Millimeter-scale network analysis using GCaMP6f expression and widefield microscopy • Connected, unspecialized networks form in the developing 3D neural culture • Network properties change upon pharmacological perturbations of neurotransmission, Optical Imaging; Techniques in Neuroscience; Biomedical Engineering

Advances in microfabrication technologies and biomaterials have enabled a growing class of electronic devices that can stimulate and record bioelectronic signals. Many of these devices have been developed for humans or vertebrate animals, where miniaturization allows for implantation within the body. There are, however, another class of bioelectronic interfaces that exploit microfabrication and nanoelectronics to record signals from tiny, millimeter-sized organisms. In these cases, rather than implanting a device inside an animal, animals themselves are loaded in large numbers into bioelectronic devices for neural circuit and behavioral interrogation. These scalable interfaces provide platforms to develop new therapeutics as well as better understand basic principles of bioelectronic communication, neuroscience, and behavior. Here we review recent progress in these bioelectronic technologies and describe how they can complement on-chip optical, mechanical, and chemical interrogation methods to achieve high-throughput, multimodal studies of millimeter-sized small animals., Graphical Abstract, Bioelectronics; Electronic Materials; Systems Neuroscience; Techniques in Neuroscience

Summary Although a variety of remarkable molecular tools for studying neural circuits have recently been developed, the ability to deploy them in particular neuronal subtypes is limited by the fact that native promoters are almost never specific enough. We recently showed that one can generate transgenic mice with anatomical specificity surpassing that of native promoters by combining enhancers uniquely active in particular brain regions with a heterologous minimal promoter, an approach we call EDGE (Enhancer-Driven Gene Expression). Here we extend this strategy to the generation of viral (rAAV) vectors, showing that some EDGE rAAVs can recapitulate the specificity of the corresponding transgenic lines in wild-type animals, even of another species. This approach thus holds the promise of enabling circuit-specific manipulations in wild-type animals, not only enhancing our understanding of brain function, but perhaps one day even providing novel therapeutic avenues to approach disorders of the brain., Graphical Abstract, Highlights • rAAVs with enhancers unique to a brain region specify cell types of that brain region • This requires viral constructs optimized to express only with enhancers • One rAAV distinguishes distinct subtypes of excitatory neurons in a cortical layer • The same specificity is seen in wild-type animals of at least two species, Molecular Biology; Neuroscience; Techniques in Neuroscience

Summary Recording the electrical activity of multiple neurons simultaneously would greatly facilitate studies on the function of neuronal circuits. The combination of the fast scanning by random-access multiphoton microscopy (RAMP) and the latest two-photon-compatible high-performance fluorescent genetically encoded voltage indicators (GEVIs) has enabled action potential detection in deep layers in in vivo brain. However, neuron connectivity analysis on optically recorded action potentials from multiple neurons in brain tissue has yet to be achieved. With high expression of a two-photon-compatible GEVI, ASAP3, via in utero electroporation and RAMP, we achieved voltage recording of spontaneous activities from multiple neurons in brain slice. We provide evidence for the developmental changes in intralaminar horizontal connections in somatosensory cortex layer 2/3 with a greater sensitivity than calcium imaging. This method thus enables investigation of neuronal network connectivity at the cellular resolution in brain tissue., Graphical Abstract, Highlights • Kilohertz-order two-photon voltage imaging with single-trial acquisition • Readout spontaneous activity from multi-cells or subcellular regions simultaneously • Long-duration voltage imaging insuring functional studies of neuron circuit • Functional connections analysis with greater precision than calcium imaging, Optical Imaging; Developmental Neuroscience; Cellular Neuroscience; Techniques in Neuroscience

Summary Multiple mazes are routinely used to test the performance of animals because each has disadvantages inherent to its shape. However, the maze shape cannot be flexibly and rapidly reproduced in a repeatable and scalable way in a single environment. Here, to overcome the lack of flexibility, scalability, reproducibility, and repeatability, we develop a reconfigurable maze system that consists of interlocking runways and an array of accompanying parts. It allows experimenters to rapidly and flexibly configure a variety of maze structures along the grid pattern in a repeatable and scalable manner. Spatial navigational behavior and hippocampal place coding were not impaired by the interlocking mechanism. As a proof-of-principle demonstration, we demonstrate that the maze morphing induces location remapping of the spatial receptive field. The reconfigurable maze thus provides flexibility, scalability, repeatability, and reproducibility, therefore facilitating consistent investigation into the neuronal substrates for learning and memory and allowing screening for behavioral phenotypes., Graphical Abstract, Highlights • The reconfigurable maze enables flexibly in building a variety of maze paths • The maze ensures reproducibility, repeatability, and scalability • The maze does not impair spatial navigational behavior or neuronal activity • Various tests of learning and memory can be conducted in a single environment, Neuroscience; Behavioral Neuroscience; Techniques in Neuroscience

Summary Long-term potentiation and depression, inferred from analysis on brain slices, are considered the cellular processes underlying learning and memory formation. They have not so far been demonstrated in human stem cell-derived neurons. By expressing channelrhodopsin in hESCs-derived glutamate neurons and co-culturing them with GABA neurons, we found that blue light stimulation increased the frequency of miniature excitatory postsynaptic currents (mEPSCs) and decreased the ratio of paired pulse facilitation (PPF) in non-ChR2-expressing GABA neurons, indicating a facilitating action at the presynaptic terminals. When paired with postsynaptic depolarization, the repetitive stimulation significantly increased the amplitude of light-evoked EPSCs that persisted during the period, indicating long-term potentiation (LTP). In contrast, low-frequency light stimulation induced long-term depression (LTD). These effects were blocked by N-methyl-D-aspartic acid (NMDA) receptor antagonists, suggesting NMDA receptor-mediated synaptic plasticity in human neural networks. Furthermore, induced pluripotent stem cell (iPSC)-derived neurons of patient with Down syndrome showed absence of LTP or LTD. Thus, our platform offers a versatile model for assessing human neural plasticity under physiological and pathological conditions., Graphical Abstract, Highlights • Repetitive stimulation induces LTP in hPSC-derived neural networks • Low-frequency light stimulation induces LTD in hPSCs-derived neural networks • The LTP/LTD in human neural networks are NMDAR dependent • Down syndrome neural networks exhibit defective LTP/LTD, Neuroscience; Cellular Neuroscience; Techniques in Neuroscience

Summary The neuromuscular junction is a synapse critical for muscle strength and coordinated motor function. Unlike CNS injuries, motor neurons mount robust regenerative responses after peripheral nerve injuries. Conversely, motor neurons selectively degenerate in diseases such as amyotrophic lateral sclerosis (ALS). To assess how these insults affect motor neurons in vivo, we performed ribosomal profiling of mouse motor neurons. Motor neuron-specific transcripts were isolated from spinal cords following sciatic nerve crush, a model of acute injury and regeneration, and in the SOD1G93A ALS model. Of the 267 transcripts upregulated after nerve crush, 38% were also upregulated in SOD1G93A motor neurons. However, most upregulated genes in injured and ALS motor neurons were context specific. Some of the most significantly upregulated transcripts in both paradigms were chemokines such as Ccl2 and Ccl7, suggesting an important role for neuroimmune modulation. Collectively these data will aid in defining pro-regenerative and pro-degenerative mechanisms in motor neurons., Graphical abstract, Highlights • A translatomic method evaluated motor neuron gene expression after injury and in ALS • This approach highlighted shared and divergent pathways during injury and degeneration • Motor neurons upregulate chemokines during axon regeneration and ALS degeneration • Regeneration upregulates neuropeptides, whereas ALS triggers developmental pathways, Biological sciences; Neuroscience; Sensory neuroscience; Techniques in neuroscience

Summary Cerebral tracts connect separated regions within a brain and serve as fundamental structures that support integrative brain functions. However, understanding the mechanisms of cerebral tract development, macro-circuit formation, and related disorders has been hampered by the lack of an in vitro model. Here, we developed a human stem cell-derived model of cerebral tracts, which is composed of two spheroids of cortical neurons and a robust fascicle of axons linking these spheroids reciprocally. In a microdevice, two spheroids of cerebral neurons extended axons into a microchannel between the spheroids and spontaneously formed an axon fascicle, mimicking a cerebral tract. We found that the formation of axon fascicle was significantly promoted when two spheroids extended axons toward each other compared with axons extended from only one spheroid. The two spheroids were able to communicate electrically through the axon fascicle. This model tissue could facilitate studies of cerebral tract development and diseases., Graphical Abstract, Highlights • A cerebral tract model was generated from iPS cell-derived cortical spheroids • Two spheroids were spontaneously connected with an axon fascicle in a microdevice • An axon fascicle electrically connected two cortical spheroids • Knockdown of L1CAM disrupted axon fascicle formation in the model tissue, Neuroscience; Techniques in Neuroscience; Biodevices

Summary Theoretically, by controlling neural membrane potential (Vm) in vivo, motion, sensation, and behavior can be controlled. Until now, there was no available technique that can increase or decrease ion concentration in vivo in real time to change neural membrane potential. We introduce a method that we coin electro-ionic modulation (EIM), wherein ionic concentration around a nerve can be controlled in real time and in vivo. We used an interface to regulate the Ca2+ ion concentration around the sciatic nerve of a frog and thus achieved stimulation and blocking with higher resolution and lower current compared with electrical stimulation. As EIM achieves higher controllability of Vm, it has potential to replace conventional methods used for the treatment of neurological disorders and may bring a new perspective to neuromodulation techniques., Graphical Abstract, Highlights • EIM regulates extracellular ion concentration in vivo in real time • EIM stimulates or blocks the nerve via Ca2+ ion depletion or enhancement • EIM achieves selective stimulation or blocking of large or small axons • EIM is the most superior neuromodulation method for real-life applications, Bioelectrical Engineering; Bioengineering; Biological Sciences; Techniques in Neuroscience

Summary Traumatic brain injury (TBI) involves complex secondary injury processes following the primary injury. The secondary injury is often associated with rapid metabolic shifts and impaired brain function immediately after the initial tissue damage. Magnetic resonance spectroscopic imaging (MRSI) coupled with hyperpolarization of 13C-labeled substrates provides a unique opportunity to map the metabolic changes in the brain after traumatic injury in real-time without invasive procedures. In this report, we investigated two patients with acute mild TBI (Glasgow coma scale 15) but no anatomical brain injury or hemorrhage. Patients were imaged with hyperpolarized [1-13C]pyruvate MRSI 1 or 6 days after head trauma. Both patients showed significantly reduced bicarbonate (HCO3–) production, and one showed hyperintense lactate production at the injured sites. This study reports the feasibility of imaging altered metabolism using hyperpolarized pyruvate in patients with TBI, demonstrating the translatability and sensitivity of the technology to cerebral metabolic changes after mild TBI.

Summary Epileptic seizures constitute a common neurological disease primarily diagnosed by characteristic rhythms or waves in the local field potentials (LFPs) of cerebral cortices or electroencephalograms. With a basolateral amygdala (BLA) kindling model, we found that the dominant frequency of BLA oscillations is in the delta range (1–5 Hz) in both normal and seizure conditions. Multi-unit discharges are increased with higher seizure staging but remain phase-locked to the delta waves in LFPs. Also, the change in synchrony precedes and outlasts the changes in discharging units as well as behavioral seizures. One short train of stimuli readily drives the pyramidal-inhibitory neuronal networks in BLA slices into prolonged reverberating activities, where the burst and interburst intervals may concurrently set a “natural wavelength” for delta frequencies. Seizures thus could be viewed as erroneous temporospatial continuums to normal oscillations in a system with a built-in synchronizing and resonating nature for information relay., Graphical Abstract, Highlights • Delta oscillations constitute the natural frequencies in basolateral amygdala • Neuronal burst and interburst intervals set the wavelength for delta oscillations • Delta oscillations may be readily transmitted distantly and translated behaviorally • Synchrony precedes and outlasts increase in discharges and seizures in ictogenesis, Behavioral Neuroscience; Systems Neuroscience; Techniques in Neuroscience

Summary Cardiac stimulation via sympathetic neurons can potentially trigger arrhythmias. We present approaches to study neuron-cardiomyocyte interactions involving optogenetic selective probing and all-optical electrophysiology to measure activity in an automated fashion. Here we demonstrate the utility of optical interrogation of sympathetic neurons and their effects on macroscopic cardiomyocyte network dynamics to address research targets such as the effects of adrenergic stimulation via the release of neurotransmitters, the effect of neuronal numbers on cardiac behavior, and the applicability of optogenetics in mechanistic in vitro studies. As arrhythmias are emergent behaviors that involve the coordinated activity of millions of cells, we image at macroscopic scales to capture complex dynamics. We show that neurons can both decrease and increase wave stability and re-entrant activity in culture depending on their induced activity—a finding that may help us understand the often conflicting results seen in experimental and clinical studies., Graphical Abstract, Highlights • A methodology to study neuron-cardiac interactions at multicellular/tissue level • Cardiac sympathetic stellate neurons can alter cardiac wave stability • Optogenetic control of sympathetic neurons offers temporal and spatial control • Increasing neuron density in co-cultures affects cardiac firing frequency, Optical Imaging; Neuroscience; Techniques in Neuroscience

Summary Spatial representations of the sensory world are important for brain function. Timing is an essential component of sensory information. Many brain circuits transform the temporal sequence of input activity into spatial maps; however, the mechanisms underlying this transformation are unclear. Different N-methyl-D-aspartate receptor (NMDAR) response magnitudes result in synaptic potentiation or depression. We asked whether NMDAR response magnitude also affects the transformation of temporal information into directional spatial maps. We quantified retinotectal axon branch dynamics in Xenopus optic tectum in response to temporal sequences of visual stimulation. Reducing NMDAR responses by 50% inverts the spatial distribution of branch dynamics along the rostrocaudal axis in response to temporal patterns of input, suggesting that the magnitude of NMDAR signaling encodes the temporal sequence of inputs and translates the temporal code into a directional spatial map using structural plasticity-based branch dynamics. We discuss how this NMDAR-dependent decoding mechanism retrieves spatial information from sequential afferent activity., Graphical Abstract, Highlights • NMDAR response magnitude encodes the temporal sequence of inputs • NMDAR mechanism decodes spatial information from sequential input activity • NMDAR attenuation inverts the temporal to spatial transformation • NMDAR activity alters the spatial distribution of dynamic and stable branches, Developmental Neuroscience; Sensory Neuroscience ; Techniques in Neuroscience

Summary Central to our subjective lives is the experience of different emotions. Recent behavioral work mapping emotional responses to 2,185 videos found that people experience upward of 27 distinct emotions occupying a high-dimensional space, and that emotion categories, more so than affective dimensions (e.g., valence), organize self-reports of subjective experience. Here, we sought to identify the neural substrates of this high-dimensional space of emotional experience using fMRI responses to all 2,185 videos. Our analyses demonstrated that (1) dozens of video-evoked emotions were accurately predicted from fMRI patterns in multiple brain regions with different regional configurations for individual emotions; (2) emotion categories better predicted cortical and subcortical responses than affective dimensions, outperforming visual and semantic covariates in transmodal regions; and (3) emotion-related fMRI responses had a cluster-like organization efficiently characterized by distinct categories. These results support an emerging theory of the high-dimensional emotion space, illuminating its neural foundations distributed across transmodal regions., Graphical Abstract, Highlights • Dozens of video-evoked emotions were predicted from fMRI patterns in multiple regions • Categories better predicted cortical and subcortical responses than dimensions • Emotion-related responses had a cluster-like organization characterized by categories • Neural representation of emotion is high-dimensional, categorical, and distributed, Neuroscience; Cognitive Neuroscience; Techniques in Neuroscience

Summary: Emerging 3D correlative light and electron microscopy approaches enable studying neuronal structure-function relations at unprecedented depth and precision. However, established protocols for the correlation of light and electron micrographs rely on the introduction of artificial fiducial markers, such as polymer beads or near-infrared brandings, which might obscure or even damage the structure under investigation. Here, we report a general applicable “flat embedding” preparation, enabling high-precision overlay of light and scanning electron micrographs, using exclusively endogenous landmarks in the brain: blood vessels, nuclei, and myelinated axons. Furthermore, we demonstrate feasibility of the workflow by combining in vivo 2-photon microscopy and focused ion beam scanning electron microscopy to dissect the role of astrocytic coverage in the persistence of dendritic spines. : Neuroscience; Techniques in Neuroscience; Biological Sciences Research Methodologies; Biological Sciences Tools Subject Areas: Neuroscience, Techniques in Neuroscience, Biological Sciences Research Methodologies, Biological Sciences Tools