The early formation of sensorimotor circuits is essential for survival. While the development and function of exteroceptive circuits and their associated motor pathways are well characterized, far less is known about the circuits that convey viscerosensory inputs to the brain and transmit visceromotor commands from the central nervous system to internal organs. Technical limitations, such as the development of viscerosensory and visceromotor circuits and the invasiveness of procedures required to access them, have hindered studies of their functional development in mammals. Using larval zebrafish-which are genetically accessible and optically transparent-we tracked, , how cardiosensory and cardiomotor neural circuits assemble and begin to function. We uncovered a staged program. First, a minimal efferent circuit suffices for heart-rate control: direct brain-to-heart vagal motor innervation is required, intracardiac neurons are not, and heart rate is governed exclusively by the motor vagus nerve. Within the hindbrain, we functionally localize a vagal premotor population that drives this early efferent control. Second, sympathetic innervation arrives and enhances the dynamics and amplitude of cardiac responses, as neurons in the most anterior sympathetic ganglia acquire the ability to drive cardiac acceleration. These neurons exhibit proportional, integral, and derivative-like relationships to heart rate, consistent with controller motifs that shape gain and dynamics. Third, vagal sensory neurons innervate the heart. Distinct subsets increase activity when heart rate falls or rises, and across spontaneous fluctuations, responses to aversive stimuli, and optogenetically evoked cardiac perturbations, their dynamics are captured by a single canonical temporal kernel with neuron-specific phase offsets, supporting a population code for heart rate. This temporally segregated maturation isolates three experimentally tractable regimes-unidirectional brain-to-heart communication, dual efferent control, and closed-loop control after sensory feedback engages-providing a framework for mechanistic dissection of organism-wide heart-brain circuits.

Coherent control of purposive actions emerges from the coordination of multiple brain circuits during learning. Dissociable brain circuits and cell-types are thought to preferentially participate in distinct learning mechanisms. For example, the activity of midbrain dopamine (mDA) neurons is proposed to primarily, or even exclusively, reflect reward prediction error signals in well-trained animals. To study the specific contribution of individual circuits requires observing changes before tight functional coordination is achieved. However, little is known about the detailed timing of the emergence of reward-related representations in dopaminergic neurons. Here we recorded activity of identified dopaminergic neurons as naive mice learned a novel stimulus-reward association. We found that at early stages of learning mDA neuron activity reflected both external (sensory) and internal (action initiation) causes of reward expectation. The increasingly precise correlation of action initiation with sensory stimuli rather than an evaluation of outcomes governed mDA neuron activity. Thus, our data demonstrate that mDA neuron activity early in learning does not reflect errors, but is more akin to a Hebbian learning signal - providing new insight into a critical computation in a highly conserved, essential learning circuit.

Animal development is a complex and dynamic process orchestrated by exquisitely timed cell lineage commitment, divisions, migration, and morphological changes at the single-cell level. In the past decade, extensive genetic, stem cell, and genomic studies provided crucial insights into molecular underpinnings and the functional importance of genetic pathways governing various cellular differentiation processes. However, it is still largely unknown how the precise coordination of these pathways is achieved at the whole-organism level and how the highly regulated spatiotemporal choreography of development is established in turn. Here, we discuss the latest technological advances in imaging and single-cell genomics that hold great promise for advancing our understanding of this intricate process. We propose an integrated approach that combines such methods to quantitatively decipher in vivo cellular dynamic behaviors and their underlying molecular mechanisms at the systems level with single-cell, single-molecule resolution.

Emotional processes are central to behavior, yet their deeply subjective nature has been a challenge for neuroscientific study as well as for psychiatric diagnosis. Here we explore the relationships between subjective feelings and their underlying brain circuits from a computational perspective. We apply recent insights from systems neuroscience-approaching subjective behavior as the result of mental computations instantiated in the brain-to the study of emotions. We develop the hypothesis that emotions are the product of neural computations whose motor role is to reallocate bodily resources mostly gated by smooth muscles. This "emotor" control system is analagous to the more familiar motor control computations that coordinate skeletal muscle movements. To illustrate this framework, we review recent research on "confidence." Although familiar as a feeling, confidence is also an objective statistical quantity: an estimate of the probability that a hypothesis is correct. This model-based approach helped reveal the neural basis of decision confidence in mammals and provides a bridge to the subjective feeling of confidence in humans. These results have important implications for psychiatry, since disorders of confidence computations appear to contribute to a number of psychopathologies. More broadly, this computational approach to emotions resonates with the emerging view that psychiatric nosology may be best parameterized in terms of disorders of the cognitive computations underlying complex behavior.

NaChBac, the first bacterial voltage-gated Na+ (Nav) channel to be characterized, has been the prokaryotic prototype for studying the structure–function relationship of Nav channels. Discovered nearly two decades ago, the structure of NaChBac has not been determined. Here we present the single particle electron cryomicroscopy (cryo-EM) analysis of NaChBac in both detergent micelles and nanodiscs. Under both conditions, the conformation of NaChBac is nearly identical to that of the potentially inactivated NavAb. Determining the structure of NaChBac in nanodiscs enabled us to examine gating modifier toxins (GMTs) of Nav channels in lipid bilayers. To study GMTs in mammalian Nav channels, we generated a chimera in which the extracellular fragment of the S3 and S4 segments in the second voltage-sensing domain from Nav1.7 replaced the corresponding sequence in NaChBac. Cryo-EM structures of the nanodisc-embedded chimera alone and in complex with HuwenToxin IV (HWTX-IV) were determined to 3.5 and 3.2 Å resolutions, respectively. Compared to the structure of HWTX-IV–bound human Nav1.7, which was obtained at an overall resolution of 3.2 Å, the local resolution of the toxin has been improved from ∼6 to ∼4 Å. This resolution enabled visualization of toxin docking. NaChBac can thus serve as a convenient surrogate for structural studies of the interactions between GMTs and Nav channels in a membrane environment.

BACKGROUND: Epigenetic mechanisms play fundamental roles in brain function and behavior and stressors such as social isolation can alter animal behavior via epigenetic mechanisms. However, due to cellular heterogeneity, identifying cell-type-specific epigenetic changes in the brain is challenging. Here, we report the first use of a modified isolation of nuclei tagged in specific cell type (INTACT) method in behavioral epigenetics of Drosophila melanogaster, a method we call mini-INTACT.

RESULTS: Using ChIP-seq on mini-INTACT purified dopaminergic nuclei, we identified epigenetic signatures in socially isolated and socially enriched Drosophila males. Social experience altered the epigenetic landscape in clusters of genes involved in transcription and neural function. Some of these alterations could be predicted by expression changes of four transcription factors and the prevalence of their binding sites in several clusters. These transcription factors were previously identified as activity-regulated genes, and their knockdown in dopaminergic neurons reduced the effects of social experience on sleep.

CONCLUSIONS: Our work enables the use of Drosophila as a model for cell-type-specific behavioral epigenetics and establishes that social environment shifts the epigenetic landscape in dopaminergic neurons. Four activity-related transcription factors are required in dopaminergic neurons for the effects of social environment on sleep.

Chromophores that absorb in the tissue-penetrant far-red/near-infrared window have long served as photocatalysts to generate singlet oxygen for photodynamic therapy. However, the cytotoxicity and side reactions associated with singlet oxygen sensitization have posed a problem for using long-wavelength photocatalysis to initiate other types of chemical reactions in biological environments. Herein, silicon-Rhodamine compounds (SiRs) are described as photocatalysts for inducing rapid bioorthogonal chemistry using 660 nm light through the oxidation of a dihydrotetrazine to a tetrazine in the presence of cyclooctene dienophiles. SiRs have been commonly used as fluorophores for bioimaging but have not been applied to catalyze chemical reactions. A series of SiR derivatives were evaluated, and the Janelia Fluor-SiR dyes were found to be especially effective in catalyzing photooxidation (typically 3%). A dihydrotetrazine/tetrazine pair is described that displays high stability in both oxidation states. A protein that was site-selectively modified by cyclooctene was quantitatively conjugated upon exposure to 660 nm light and a dihydrotetrazine. By contrast, a previously described methylene blue catalyst was found to rapidly degrade the protein. SiR-red light photocatalysis was used to cross-link hyaluronic acid derivatives functionalized by dihydrotetrazine and cyclooctenes, enabling 3D culture of human prostate cancer cells. Photoinducible hydrogel formation could also be carried out in live mice through subcutaneous injection of a Cy7-labeled hydrogel precursor solution, followed by brief irradiation to produce a stable hydrogel. This cytocompatible method for using red light photocatalysis to activate bioorthogonal chemistry is anticipated to find broad applications where spatiotemporal control is needed in biological environments.

There is considerable potential for X-ray free electron lasers (XFELs) to enable determination of macromolecular crystal structures that are difficult to solve using current synchrotron sources. Prior XFEL studies often involved the collection of thousands to millions of diffraction images, in part due to limitations of data processing methods. We implemented a data processing system based on classical post-refinement techniques, adapted to specific properties of XFEL diffraction data. When applied to XFEL data from three different proteins collected using various sample delivery systems and XFEL beam parameters, our method improved the quality of the diffraction data as well as the resulting refined atomic models and electron density maps. Moreover, the number of observations for a reflection necessary to assemble an accurate data set could be reduced to a few observations. These developments will help expand the applicability of XFEL crystallography to challenging biological systems, including cases where sample is limited.

The role of cerebellum in controlling eye movements is well established, but its contribution to more complex forms of visual behavior has remained elusive. To study cerebellar activity during visual attention we recorded extracellular activity of dentate nucleus (DN) neurons in two non-human primates (NHPs). NHPs were trained to read the direction indicated by a peripheral visual stimulus while maintaining fixation at the center, and report the direction of the cue by performing a saccadic eye movement into the same direction following a delay. We found that single-unit DN neurons modulated spiking activity over the entire time course of the task, and that their activity often bridged temporally separated intra-trial events, yet in a heterogeneous manner. To better understand the heterogeneous relationship between task structure, behavioral performance, and neural dynamics, we constructed a behavioral, an encoding, and a decoding model. Both NHPs showed different behavioral strategies, which influenced the performance. Activity of the DN neurons reflected the unique strategies, with the direction of the visual stimulus frequently being encoded long before an upcoming saccade. Moreover, the latency of the ramping activity of DN neurons following presentation of the visual stimulus was shorter in the better performing NHP. Labeling with the retrograde tracer Cholera Toxin B in the recording location in the DN indicated that these neurons predominantly receive inputs from Purkinje cells in the D1 and D2 zones of the lateral cerebellum as well as neurons of the principal olive and medial pons, all regions known to connect with neurons in the prefrontal cortex contributing to planning of saccades. Together, our results highlight that DN neurons can dynamically modulate their activity during a visual attention task, comprising not only sensorimotor but also cognitive attentional components.

The role of cerebellum in controlling eye movements is well established, but its contribution to more complex forms of visual behavior has remained elusive. To study cerebellar activity during visual attention we recorded extracellular activity of dentate nucleus (DN) neurons in two non-human primates (NHPs). NHPs were trained to read the direction indicated by a peripheral visual stimulus while maintaining fixation at the center, and report the direction of the cue by performing a saccadic eye movement into the same direction following a delay. We found that single unit DN neurons modulated spiking activity over the entire time-course of the task, and that their activity often bridged temporally separated intra-trial events, yet in a heterogeneous manner. To better understand the heterogeneous relationship between task structure, behavioral performance and neural dynamics, we constructed a behavioral, an encoding and a decoding model. Both NHPs showed different behavioral strategies, which influenced the performance. Activity of the DN neurons reflected the unique strategies, with the direction of the visual stimulus frequently being encoded long before an upcoming saccade. Retrograde labeling of the recording location indicated that these neurons receive predominantly inputs from Purkinje cells in the lateral cerebellum as well as neurons of the principal olive and medial pons, all regions known to connect with neurons in the prefrontal cortex contributing to planning of saccades. Together, our results highlight that DN neurons can dynamically modulate their activity during a visual attention task, comprising not only sensorimotor but also cognitive attentional components.