The Walking Control of a Crab Biorobot in Amphibious Environment.

This article describes development of a crab biorobot that is capable of traversing diverse environments including both land and water. We have transformed a living rainbow crab into a walking biorobot by attaching wireless controller. An anatomical and physiological investigation revealed the rainbow crabs have sensory system on the carapace. Based on this finding, we implanted electrodes into the carapace. The walking direction of the robot is controlled through electrical stimulation provided by the controller. Depending on this site, the crab biorobot is induced to walk forward, leftward, and rightward in both terrestrial and underwater conditions. There is no significant difference in the mean walking direction between the two conditions. Smooth transition from land to water of the crab biorobot further demonstrates the adaptability to amphibious environment. This biorobot is compact, measuring 5 cm in carapace and weighing 50 g including the wireless controller. The crab biorobot in this scale has a potential for application narrow and unstructured in waterfront environments.

A Navigation Algorithm to Enable Sustainable Control of Insect-Computer Hybrid Robot with Stimulus Signal Regulator and Habituation-Breaking Function.

The insect-computer hybrid soft robots are receiving increasing attention due to their excellent motor capabilities, small size, and low power consumption. However, the effective control of insects is limited to minutes since the response from insects is reduced as the number of stimulus signal increase. This phenomenon is known as habituation, which causes the loss of control of robots and hinders their application in practical tasks such as search and rescue missions that require several hours. It has been shown that constantly switching the pattern of stimulus signals can slow down the onset of habituation. Moreover, when habituation occurs, applying a different stimulus signal can break the habituation. Based on this, we have designed a navigation algorithm that can extend the control time of insects to several hours. The algorithm is composed of a stimulation decision-making core responsible for deciding on the type of stimulus signal (left, right, acceleration), a stimulation parameters adjustment (SPA) core responsible for adjusting the stimulus signal voltage constantly to delay the occurrence of habituation, and a reactivation function (RF), as a different stimulus signal from the normal stimulus signal, is used to break insects' habituation to the normal stimulus signal. Experiments have shown that our SPA regulator and RF can significantly extend the control time of insects. Navigation experiments demonstrating effective control of the insects for up to 3 h verified the effectiveness of the navigation algorithm, which strikes a balance between control accuracy and control time.

Toll-like receptor 9-positive plasmacytoid dendritic cells promote Th17 immune responses in oral lichen planus stimulated by epithelium-derived cathepsin K.

Oral lichen planus (OLP) is a chronic inflammatory disease associated with T cell infiltration. The crosstalk between oral epithelium and mucosal T cells was considered to be crucial in the pathogenesis of OLP. Here, we selectively extracted the normal epithelium (NE) and lesional epithelium (LE) of buccal mucosa specimens from three patients with OLP by laser capture microdissection due to identify the pathogenic factors. Cathepsin K (CTSK) was identified as one of common upregulated genes in the LE by DNA microarray. Immunohistochemically, CTSK was distinctly detected in and around the LE, while it was rarely seen in the NE. Recent studies showed that CTSK enhanced Toll-like receptor 9 (TLR9) signaling in antigen-presenting cells, leading to Th17 cell differentiation. TLR9 expression mainly co-localized with CD123+ plasmacytoid dendritic cells (pDCs). The number of RORγt-positive cells correlated with that of CTSK-positive cells in OLP tissues. CD123+ pDCs induced the production of Th17-related cytokines (IL-6, IL-23, and TGF-ß) upon stimulation with TLR9 agonist CpG DNA. Moreover, single cell RNA-sequencing analysis revealed that TLR9-positive pDCs enhanced in genes associated with Th17 cell differentiation in comparison with TLR9-negative pDCs. CTSK could induce Th17-related production of CD123+ pDCs via TLR9 signaling to promote the pathogenesis of OLP.

Performance-Dependent Consolidation of Learned Vocal Changes in Adult Songbirds.

Motor skills learned through practice are consolidated at later time, which can include nighttime, but the time course of motor memory consolidation and its underlying mechanisms remain poorly understood. We investigated neural substrates underlying motor memory consolidation of learned changes in birdsong, a tractable model system for studying neural basis of motor skill learning. Previous studies in male zebra finches and Bengalese finches have demonstrated that adaptive changes in adult song structure learned through a reinforcement paradigm are initially driven by a cortical-basal ganglia circuit, and subsequently consolidated into downstream cortical motor circuitry. However, the time course of the consolidation process, including whether it occurs offline during nighttime or online during daytime, remains unclear and even controversial. Here, we provide in both species experimental evidence of virtually no consolidation of learned vocal changes during nighttime. We demonstrate instead that the consolidation occurs during daytime and the amount of consolidation is strongly correlated with the amount of learning, suggesting online, performance-dependent mechanisms of consolidation of learned vocal changes. Moreover, by using computer simulations based on our experimental results, we demonstrate that such online, performance-dependent consolidation can account for the contradicting conclusions concerning the time course of consolidation process reached by previous studies. These results thus reconcile a controversy in the study of vocal motor consolidation in songbirds, and illustrate the neural substrates through which newly learned motor skills initially implemented by cortical-basal ganglia circuits become encoded in the cortical motor circuitry.SIGNIFICANCE STATEMENT Motor skills learned through repetitive practice become stable and are consolidated into cortical motor circuits. We investigate neural substrates of this "motor memory consolidation" in adult songbirds, which produce songs that are complex motor skills learned and maintained through repetitive vocal practice. We demonstrate that learned changes in song acoustic structure are consolidated into the cortical motor circuits predominantly during daytime, but not during nighttime, depending on ongoing song performance. These consolidation mechanisms reconcile seemingly contradicting results of previous studies regarding the time course of vocal learning consolidation, and provide fundamental insights into the process through which learned performance of complex motor skills is consolidated and encoded in in motor circuits.

CD163+ M2 Macrophages Promote Fibrosis in IgG4-Related Disease Via Toll-like Receptor 7/Interleukin-1 Receptor-Associated Kinase 4/NF-κB Signaling.

OBJECTIVE: IgG4-related disease (IgG4-RD) is a fibro-inflammatory condition that can affect multiple organs. We previously demonstrated that TLR7-transgenic C57BL/6 mice showed elevated serum IgG1 levels and inflammation with fibrosis in the salivary glands (SGs), lungs, and pancreas. Moreover, we observed extensive Toll-like receptor 7 (TLR-7)-positive CD163+ M2 macrophage infiltration in SGs from IgG4-RD patients. We undertook this study to examine the fibrotic mechanism via the TLR-7 pathway. METHODS: Gene expression in SGs from human TLR7-transgenic mice and IgG4-RD patients was analyzed using DNA microarrays. We extracted the common up-regulated TLR-7-related genes in SGs from TLR7-transgenic mice and IgG4-RD patients. Finally, we investigated the interaction between CD163+ M2 macrophages and fibroblasts before and after stimulation with the TLR-7 agonist loxoribine. RESULTS: In TLR7-transgenic mice and IgG4-RD patients, IRAK3 and IRAK4 were significantly overexpressed. Real-time polymerase chain reaction validated the up-regulation of only IRAK4 in IgG4-RD patients compared with the other groups (P < 0.05). Interleukin-1 receptor-associated kinase 4 (IRAK4) was strongly detected in and around germinal centers in SGs from patients with IgG4-related dacryoadenitis and sialadenitis alone. Double immunofluorescence staining showed that IRAK4-positive cells were mainly colocalized with CD163+ M2 macrophages in SGs (P < 0.05). After stimulation with loxoribine, CD163+ M2 macrophages exhibited significantly enhanced expression of IRAK4 and NF-κB and increased supernatant concentrations of fibrotic cytokines. Finally, we confirmed that the number of fibroblasts was increased by culture with the supernatant of CD163+ M2 macrophages following stimulation with loxoribine (P < 0.05). CONCLUSION: CD163+ M2 macrophages promote fibrosis in IgG4-RD by increasing the production of fibrotic cytokines via TLR-7/IRAK4/NF-κB signaling.

Oral Squamous Cell Carcinoma Contributes to Differentiation of Monocyte-Derived Tumor-Associated Macrophages via PAI-1 and IL-8 Production.

Tumor-associated macrophages (TAMs) promote cancer cell proliferation and metastasis, as well as anti-tumor immune suppression. Recent studies have shown that tumors enhance the recruitment and differentiation of TAMs, but the detailed mechanisms have not been clarified. We thus examined the influence of cancer cells on the differentiation of monocytes to TAM subsets, including CD163+, CD204+, and CD206+ cells, in oral squamous cell carcinoma (OSCC) using immunohistochemistry, flow cytometry, and a cytokine array. Furthermore, we investigated the effect of OSCC cells (HSC-2, SQUU-A, and SQUU-B cells) on the differentiation of purified CD14+ cells to TAM subsets. The localization patterns of CD163+, CD204+, and CD206+ in OSCC sections were quite different. The expression of CD206 on CD14+ cells was significantly increased after the co-culture with OSCC cell lines, while the expressions of CD163 and CD204 on CD14+ cells showed no change. High concentrations of plasminogen activator inhibitor-1 (PAI-1) and interleukin-8 (IL-8) were detected in the conditioned medium of OSCC cell lines. PAI-1 and IL-8 stimulated CD14+ cells to express CD206. Moreover, there were positive correlations among the numbers of CD206+, PAI-1+, and IL-8+ cells in OSCC sections. These results suggest that PAI-1 and IL-8 produced by OSCC contribute to the differentiation of monocytes to CD206+ TAMs.

Adaptations during Maturation in an Identified Honeybee Interneuron Responsive to Waggle Dance Vibration Signals.

Honeybees are social insects, and individual bees take on different social roles as they mature, performing a multitude of tasks that involve multi-modal sensory integration. Several activities vital for foraging, like flight and waggle dance communication, involve sensing air vibrations through their antennae. We investigated changes in the identified vibration-sensitive interneuron DL-Int-1 in the honeybee Apis mellifera during maturation by comparing properties of neurons from newly emerged adult and forager honeybees. Although comparison of morphological reconstructions of the neurons revealed no significant changes in gross dendritic features, consistent and region-dependent changes were found in dendritic density. Comparison of electrophysiological properties showed an increase in the firing rate differences between stimulus and nonstimulus periods in foragers compared with newly emerged adult bees. The observed differences in neurons of foragers compared with newly emerged adult honeybees suggest refined connectivity, improved signal propagation, and enhancement of response features possibly important for the network processing of air vibration signals relevant for the waggle dance communication of honeybees.

A Segmentation Scheme for Complex Neuronal Arbors and Application to Vibration Sensitive Neurons in the Honeybee Brain.

The morphology of a neuron is strongly related to its physiological properties, application of logical product and thus to information processing functions. Optical microscope images are widely used for extracting the structure of neurons. Although several approaches have been proposed to trace and extract complex neuronal structures from microscopy images, available methods remain prone to errors. In this study, we present a practical scheme for processing confocal microscope images and reconstructing neuronal structures. We evaluated this scheme using image data samples and associated "gold standard" reconstructions from the BigNeuron Project. In these samples, dendritic arbors belonging to multiple projection branches of the same neuron overlapped in space, making it difficult to automatically and accurately trace their structural connectivity. Our proposed scheme, which combines several software tools for image masking and filtering with an existing tool for dendritic segmentation and tracing, outperformed state-of-the-art automatic methods in reconstructing such neuron structures. For evaluating our scheme, we applied it to a honeybee local interneuron, DL-Int-1, which has complex arbors and is considered to be a critical neuron for encoding the distance information indicated in the waggle dance of the honeybee.

Spatial registration of neuron morphologies based on maximization of volume overlap.

BACKGROUND: Morphological features are widely used in the study of neuronal function and pathology. Invertebrate neurons are often structurally stereotypical, showing little variance in gross spatial features but larger variance in their fine features. Such variability can be quantified using detailed spatial analysis, which however requires the morphologies to be registered to a common frame of reference. RESULTS: We outline here new algorithms - Reg-MaxS and Reg-MaxS-N - for co-registering pairs and groups of morphologies, respectively. Reg-MaxS applies a sequence of translation, rotation and scaling transformations, estimating at each step the transformation parameters that maximize spatial overlap between the volumes occupied by the morphologies. We test this algorithm with synthetic morphologies, showing that it can account for a wide range of transformation differences and is robust to noise. Reg-MaxS-N co-registers groups of more than two morphologies by iteratively calculating an average volume and registering all morphologies to this average using Reg-MaxS. We test Reg-MaxS-N using five groups of morphologies from the Droshophila melanogaster brain and identify the cases for which it outperforms existing algorithms and produce morphologies very similar to those obtained from registration to a standard brain atlas. CONCLUSIONS: We have described and tested algorithms for co-registering pairs and groups of neuron morphologies. We have demonstrated their application to spatial comparison of stereotypic morphologies and calculation of dendritic density profiles, showing how our algorithms for registering neuron morphologies can enable new approaches in comparative morphological analyses and visualization.

Interneurons in the Honeybee Primary Auditory Center Responding to Waggle Dance-Like Vibration Pulses.

Female honeybees use the "waggle dance" to communicate the location of nectar sources to their hive mates. Distance information is encoded in the duration of the waggle phase (von Frisch, 1967). During the waggle phase, the dancer produces trains of vibration pulses, which are detected by the follower bees via Johnston's organ located on the antennae. To uncover the neural mechanisms underlying the encoding of distance information in the waggle dance follower, we investigated morphology, physiology, and immunohistochemistry of interneurons arborizing in the primary auditory center of the honeybee (Apis mellifera). We identified major interneuron types, named DL-Int-1, DL-Int-2, and bilateral DL-dSEG-LP, that responded with different spiking patterns to vibration pulses applied to the antennae. Experimental and computational analyses suggest that inhibitory connection plays a role in encoding and processing the duration of vibration pulse trains in the primary auditory center of the honeybee.SIGNIFICANCE STATEMENT The waggle dance represents a form of symbolic communication used by honeybees to convey the location of food sources via species-specific sound. The brain mechanisms used to decipher this symbolic information are unknown. We examined interneurons in the honeybee primary auditory center and identified different neuron types with specific properties. The results of our computational analyses suggest that inhibitory connection plays a role in encoding waggle dance signals. Our results are critical for understanding how the honeybee deciphers information from the sound produced by the waggle dance and provide new insights regarding how common neural mechanisms are used by different species to achieve communication.

NeuronDepot: keeping your colleagues in sync by combining modern cloud storage services, the local file system, and simple web applications.

Neuroscience today deals with a "data deluge" derived from the availability of high-throughput sensors of brain structure and brain activity, and increased computational resources for detailed simulations with complex output. We report here (1) a novel approach to data sharing between collaborating scientists that brings together file system tools and cloud technologies, (2) a service implementing this approach, called NeuronDepot, and (3) an example application of the service to a complex use case in the neurosciences. The main drivers for our approach are to facilitate collaborations with a transparent, automated data flow that shields scientists from having to learn new tools or data structuring paradigms. Using NeuronDepot is simple: one-time data assignment from the originator and cloud based syncing-thus making experimental and modeling data available across the collaboration with minimum overhead. Since data sharing is cloud based, our approach opens up the possibility of using new software developments and hardware scalabitliy which are associated with elastic cloud computing. We provide an implementation that relies on existing synchronization services and is usable from all devices via a reactive web interface. We are motivating our solution by solving the practical problems of the GinJang project, a collaboration of three universities across eight time zones with a complex workflow encompassing data from electrophysiological recordings, imaging, morphological reconstructions, and simulations.

Characterization of locomotor-related spike activity in protocerebrum of freely walking cricket.

To characterize the neural elements involved in the higher-order control of spontaneous walking in insects, we recorded extracellular spike activity in the protocerebrum of freely walking crickets (Gryllus bimaculatus). Locomotor behavior was simultaneously recorded using a newly developed motion tracking system. We focused on spike units that altered their firing patterns during walking. According to their activity patterns with reference to walking bouts, these locomotor-related spike units were classified into the following four types: continuously activated unit during walking (type 1); continuously inhibited unit during walking (type 2); transiently activated unit at the onset of walking (type 3); and transiently activated unit at the termination of walking (type 4). The type 1 unit was the most dominant group (25 out of 33 units), whereas only a few units each were recorded for types 2-4. Some of the locomotor-related units tended to change firing pattern before the onset or termination of walking bouts. Spike activity in some type 1 units was found to be closely correlated with walking speed. When firing timing was compared between unit pairs, their temporal relationships (synchronization/desynchronization) altered, depending on the behavioral state (standing/walking). Mechanical stimuli applied to the body surface elicited excitatory responses in the majority of the units. Histological observations revealed that the recorded sites were concentrated near or within the mushroom body and central complex in the protocerebrum.