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Vagus nerve stimulation (VNS) is a neuromodulation therapy for a broad and expanding set of neurologic conditions. However, the mechanism through which VNS influences central nervous system circuitry is not well described, limiting therapeutic optimization. VNS leads to widespread brain activation, but the effects on behavior are remarkably specific, indicating plasticity unique to behaviorally engaged neural circuits. To understand how VNS can lead to specific circuit modulation, we leveraged genetic tools including optogenetics and in vivo calcium imaging in mice learning a skilled reach task. We find that VNS enhances skilled motor learning in healthy animals via a cholinergic reinforcement mechanism, producing a rapid consolidation of an expert reach trajectory. In primary motor cortex (M1), VNS drives precise temporal modulation of neurons that respond to behavioral outcome. This suggests that VNS may accelerate motor refinement in M1 via cholinergic signaling, opening new avenues for optimizing VNS to target specific disease-relevant circuitry.

Vagus nerve stimulation (VNS) is a neuromodulation therapy for a broad and rapidly expanding set of neurologic conditions. Classically used to treat epilepsy and depression, VNS has recently received FDA approval for stroke rehabilitation and is under preclinical and clinical investigation for other neurologic indications. Despite benefits across a diverse range of neurological disorders, the mechanism through which VNS influences central nervous system circuitry is not well described, limiting therapeutic optimization. A deeper understanding of the influence of VNS on neural circuits and activity is needed to maximize the use of VNS therapy across a broad range of neurologic conditions.To investigate how VNS can influence the neurons and circuits that underlie behavior, we paired VNS with upper limb movement in mice learning a skilled motor task. We leveraged genetic tools to perform optogenetic circuit dissection, as well as longitudinal in vivo imaging of calcium activity in cortical neurons to understand the effect of VNS on neural function. We found that VNS robustly enhanced motor learning when temporally paired with successful movement outcome, while randomly applied VNS impaired learning. This suggests that temporally-precise VNS may act through augmenting outcome cues, such as reinforcement signals. Within motor cortex, VNS paired with movement outcome selectively modulates the neural population that represents outcome, but not other movement-related neurons, across both acute and behaviorally-relevant timescales. Phasic cholinergic signaling from basal forebrain is required both for VNS-driven improvements in motor learning and the effects on neural activity in M1. These results indicate that VNS enhances motor learning through precisely-timed phasic cholinergic signaling to reinforce outcome, resulting in the recruitment of specific, behaviorally-relevant cortical circuits. A deeper understanding of the mechanisms of VNS on neurons, circuits and behavior provides new opportunities to optimize VNS to treat neurologic conditions.

TheDrosophilaventral nerve cord (VNC), the fly equivalent of the spinal cord, is composed of thousands of neurons that are born from a set of individually identifiable stem cells. The VNC harbors neuronal circuits required for the execution of vital behaviors, such as flying and walking. Taking advantage of the lineage-based functional organization of the VNC and genetic tools we developed, we investigated the molecular and developmental basis of behavior by focusing on lineage-specific functions of the homeodomain transcription factor, Unc-4. We found that Unc-4 functions in lineage 11A to promote cholinergic neurotransmitter identity and suppress the GABA fate. In 7B lineage, Unc-4 promotes proper neuronal projections to the leg neuropil, the hub for leg-related neuronal circuits and a specific flight-related take-off behavior. We also uncovered that Unc-4 acts peripherally to promote the development of proprioceptive sense organs and the abilities of flies to execute specific leg-related behaviors such as walking, climbing, and grooming. Our findings, thus, initiates the work on revealing molecular and developmental events that shape the VNC related behaviors.

Summary: Sparse manipulation of neuron excitability during free behavior is critical for identifying neural substrates of behavior. Genetic tools for precise neuronal manipulation exist in the fruit fly, Drosophila melanogaster, but behavioral tools are still lacking to identify potentially subtle phenotypes only detectible using high-throughput and high spatiotemporal resolution. We developed three assay components that can be used modularly to study natural and optogenetically induced behaviors. FlyGate automatically releases flies one at a time into an assay. FlyDetect tracks flies in real time, is robust to severe occlusions, and can be used to track appendages, such as the head. GlobeDisplay is a spherical projection system covering the fly’s visual receptive field with a single projector. We demonstrate the utility of these components in an integrated system, FlyPEZ, by comprehensively modeling the input-output function for directional looming-evoked escape takeoffs and describing a millisecond-timescale phenotype from genetic silencing of a single visual projection neuron type. : Williamson et al. developed a high-throughput device to automatically isolate individual flies and record their behavior with high-speed video. They demonstrate the system’s utility for studying the Drosophila brain using modern genetic tools by discovering behavioral phenotypes that result from manipulation of small groups of neurons. Keywords: neuroscience, Drosophila, behavior, neural substrates, high throughput, neural circuits, high-speed video, visual stimulation, neural activation, escape, machine vision

Visual projection neurons (VPNs) provide an anatomical connection between early visual processing and higher brain regions. Here we characterize lobula columnar (LC) cells, a class of Drosophila VPNs that project to distinct central brain structures called optic glomeruli. We anatomically describe 22 different LC types and show that, for several types, optogenetic activation in freely moving flies evokes specific behaviors. The activation phenotypes of two LC types closely resemble natural avoidance behaviors triggered by a visual loom. In vivo two-photon calcium imaging reveals that these LC types respond to looming stimuli, while another type does not, but instead responds to the motion of a small object. Activation of LC neurons on only one side of the brain can result in attractive or aversive turning behaviors depending on the cell type. Our results indicate that LC neurons convey information on the presence and location of visual features relevant for specific behaviors. DOI: http://dx.doi.org/10.7554/eLife.21022.001, eLife digest Many animals rely heavily on what they can see to interact with the world around them. But how does the brain use such visual information to guide behavior? Light-sensitive neurons in the eye cannot distinguish between the visual signals associated with, say, an approaching predator or a source of food. Yet the brain can make this distinction. Networks of neurons in the brain perform computations to extract information from a visual scene that indicates the need for a particular behavior, such as an escape response. These networks are found in regions of the brain that communicate closely with the eyes. Cells known as visual projection neurons then relay the output of these networks to more central parts of the brain. By studying visual projection neurons, it is possible to work out what the eye tells the brain, and how the brain uses this information to control behavior. The fruit fly Drosophila is a suitable model organism in which to study these phenomena. This insect shows a range of behavioral responses to visual stimuli, and can be studied using sophisticated genetic tools. Wu, Nern et al. set out to explore how a group of visual projection neurons known as lobula columnar cells help fruit flies respond appropriately to visual stimuli. Experiments revealed that individual subtypes of lobula columnar cells convey information about the presence and general location of specific visual features. Wu, Nern et al. identified a number of lobular columnar subtypes involved in triggering escape responses to specific stimuli – such as walking backwards or taking off in flight – as well as others that can trigger the flies to approach a target. A next step is to map the circuits of neurons that act upstream and downstream of lobula columnar cells. This can help to reveal how these neurons detect specific visual features and how the fly then chooses and executes an appropriate behavior in response. Such studies in flies can provide insights into general principles of how brains use sensory information to guide behavior. DOI: http://dx.doi.org/10.7554/eLife.21022.002

Summary Animals rely on dedicated sensory circuits to extract and encode environmental features. How individual neurons integrate and translate these features into behavioral responses remains a major question. Here, we identify a visual projection neuron type that conveys predator approach information to the Drosophila giant fiber (GF) escape circuit. Genetic removal of this input during looming stimuli reveals that it encodes angular expansion velocity, whereas other input cell type(s) encode angular size. Motor program selection and timing emerge from linear integration of these two features within the GF. Linear integration improves size detection invariance over prior models and appropriately biases motor selection to rapid, GF-mediated escapes during fast looms. Our findings suggest feature integration, and motor control may occur as simultaneous operations within the same neuron and establish the Drosophila escape circuit as a model system in which these computations may be further dissected at the circuit level. Video Abstract

Interindividual differences in neuronal wiring may contribute to behavioral individuality and affect susceptibility to neurological disorders. To investigate the causes and potential consequences of wiring variation in Drosophila melanogaster, we focused on a hemilineage of ventral nerve cord interneurons that exhibits morphological variability. We find that late-born subclasses of the 12A hemilineage are highly sensitive to genetic and environmental variation. Neurons in the second thoracic segment are particularly variable with regard to two developmental decisions, whereas its segmental homologs are more robust. This variability “hotspot” depends on Ultrabithorax expression in the 12A neurons, indicating variability is cell-intrinsic and under genetic control. 12A development is more variable and sensitive to temperature in long-established laboratory strains than in strains recently derived from the wild. Strains with a high frequency of one of the 12A variants also showed a high frequency of animals with delayed spontaneous flight initiation, whereas other wing-related behaviors did not show such a correlation and were thus not overtly affected by 12A variation. These results show that neurodevelopmental robustness is variable and under genetic control in Drosophila and suggest that the fly may serve as a model for identifying conserved gene pathways that stabilize wiring in stressful developmental environments. Moreover, some neuronal lineages are variation hotspots and thus may be more amenable to evolutionary change.