[Calciphylaxis: an enigma to the nephrologist]. / Calcifilassi: un enigma per il nefrologo.

Calcific uremic arteriopathy (CUA), also known as calciphylaxis, is a rare condition occurring in patients with moderate to severe chronic kidney disease. It is a serious, debilitating and potentially fatal clinical disorder affecting 1-4% of the dialysis population and is associated with a high mortality rate (60-80%). The clinical picture is characterized by painful skin lesions tending to necrotic or gangrenous ulceration ultimately necessitating amputation. Severe infectious complications leading to sepsis and death are frequent. The pathogenesis of CUA is still unknown and several pathogenetic hypotheses have been put forward; this makes its treatment difficult and often empirical. The current paper presents a systematic review of recent findings on the pathogenesis, diagnosis and treatment of CUA.

A miniature fluorescence microscope for multi-plane imaging.

Miniature fluorescence microscopes are becoming an increasingly established tool to investigate neural circuits in freely moving animals. In this work we present a lightweight one-photon microscope capable of imaging at different focal depths. The focal plane can be changed dynamically by modulating the pulse width of the control signal to a variable focus liquid lens, which is synchronized to the image sensor to enable changing focal plane between frames. The system was tested by imaging GCaMP7f expressing neurons in the mouse medial prefrontal cortex (mPFC) in vivo during open field test. Results showed that with the proposed design it is possible to image neurons across an axial scan of ~ 60 µm, resulting in a ~ 40% increase of total neurons imaged compared to single plane imaging.

Striatal direct pathway neurons play leading roles in accelerating rotarod motor skill learning.

Dorsal striatum is important for movement control and motor skill learning. However, it remains unclear how the spatially and temporally distributed striatal medium spiny neuron (MSN) activity in the direct and indirect pathways (D1 and D2 MSNs, respectively) encodes motor skill learning. Combining miniature fluorescence microscopy with an accelerating rotarod procedure, we identified two distinct MSN subpopulations involved in accelerating rotarod learning. In both D1 and D2 MSNs, we observed neurons that displayed activity tuned to acceleration during early stages of trials, as well as movement speed during late stages of trials. We found a distinct evolution trajectory for early-stage neurons during motor skill learning, with the evolution of D1 MSNs correlating strongly with performance improvement. Importantly, optogenetic inhibition of the early-stage neural activity in D1 MSNs, but not D2 MSNs, impaired accelerating rotarod learning. Together, this study provides insight into striatal D1 and D2 MSNs encoding motor skill learning.

Aberrant neural activity in prefrontal pyramidal neurons lacking TDP-43 precedes neuron loss.

Mislocalization of TAR DNA binding protein 43 kDa (TARDBP, or TDP-43) is a principal pathological hallmark identified in cases of neurodegenerative disorders such as amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). As an RNA binding protein, TDP-43 serves in the nuclear compartment to repress non-conserved cryptic exons to ensure the normal transcriptome. Multiple lines of evidence from animal models and human studies support the view that loss of TDP-43 leads to neuron loss, independent of its cytosolic aggregation. However, the underlying pathogenic pathways driven by the loss-of-function mechanism are still poorly defined. We employed a genetic approach to determine the impact of TDP-43 loss in pyramidal neurons of the prefrontal cortex (PFC). Using a custom-built miniscope imaging system, we performed repetitive in vivo calcium imaging from freely behaving mice for up to 7 months. By comparing calcium activity in PFC pyramidal neurons between TDP-43 depleted and TDP-43 intact mice, we demonstrated remarkably increased numbers of pyramidal neurons exhibiting hyperactive calcium activity after short-term TDP-43 depletion, followed by rapid activity declines prior to neuron loss. Our results suggest aberrant neural activity driven by loss of TDP-43 as the pathogenic pathway at early stage in ALS and FTD.

Detailed mapping of behavior reveals the formation of prelimbic neural ensembles across operant learning.

The prelimbic cortex (PrL) is involved in the organization of operant behaviors, but the relationship between longitudinal PrL neural activity and operant learning and performance is unknown. Here, we developed deep behavior mapping (DBM) to identify behavioral microstates in video recordings. We combined DBM with longitudinal calcium imaging to quantify behavioral tuning in PrL neurons as mice learned an operant task. We found that a subset of PrL neurons were strongly tuned to highly specific behavioral microstates, both task and non-task related. Overlapping neural ensembles were tiled across consecutive microstates in the response-reinforcer sequence, forming a continuous map. As mice learned the operant task, weakly tuned neurons were recruited into new ensembles, with a bias toward behaviors similar to their initial tuning. In summary, our data suggest that the PrL contains neural ensembles that jointly encode a map of behavioral states that is fine grained, is continuous, and grows during operant learning.

Circuit Investigation of Social Interaction and Substance Use Disorder Using Miniscopes.

Substance use disorder (SUD) is comorbid with devastating health issues, social withdrawal, and isolation. Successful clinical treatments for SUD have used social interventions. Neurons can encode drug cues, and drug cues can trigger relapse. It is important to study how the activity in circuits and embedded cell types that encode drug cues develop in SUD. Exploring shared neurobiology between social interaction (SI) and SUD may explain why humans with access to social treatments still experience relapse. However, circuitry remains poorly characterized due to technical challenges in studying the complicated nature of SI and SUD. To understand the neural correlates of SI and SUD, it is important to: (1) identify cell types and circuits associated with SI and SUD, (2) record and manipulate neural activity encoding drug and social rewards over time, (3) monitor unrestrained animal behavior that allows reliable drug self-administration (SA) and SI. Miniaturized fluorescence microscopes (miniscopes) are ideally suited to meet these requirements. They can be used with gradient index (GRIN) lenses to image from deep brain structures implicated in SUD. Miniscopes can be combined with genetically encoded reporters to extract cell-type specific information. In this mini-review, we explore how miniscopes can be leveraged to uncover neural components of SI and SUD and advance potential therapeutic interventions.

An open source motorized swivel for in vivo neural and behavioral recordings.

In this work we propose an open source, cost-effective motorized swivel for behavioral and neural recordings in small rodents, offering a flexible solution for managing cable twisting and tangling in a variety of experimental settings with minimal human supervision.•The device operates independently of the data acquisition system, and it can be controlled through any popular platform such as Arduino or Raspberry Pi.•All mechanical parts are 3D-printed, allowing to customize the design to fit specific experimental needs, and electromechanical components can be sourced from all major distributors, keeping the cost for the entire system under $500.•The proposed commutator is compatible with commercial or custom data acquisition systems supporting up to 10 data lines (2 for LVDS signals) and 2 power lines.

An open-source capacitive touch sensing device for three chamber social behavior test.

A common feature of many neuropsychiatric disorders is deficit in social behavior. In order to study mouse models for such disorders, several behavioral tests involving social interaction with other mice have been developed. While a precise annotation of rodent behavioral state is necessary for these types of experiments, manual annotation of rodent social behavior is time-consuming and subjective. Therefore, an automated system that can instantly and independently quantify the animal's social exploration is desirable. We developed a capacitive touch device for automated detection of direct social-exploration in a modified three-chamber social behavior test. In this device, capacitive sensors can readily detect nose-pokes and other direct physical touches from the rodent under investigation. In addition, a conductive barrier makes mouse behavioral output immediately available for real-time use, by sending data to a host computer via a custom Field-Programmable Gate Array (FPGA) platform. Our capacitive touch sensing device produced similar results to the manually annotated data, demonstrating the ability to instantly and independently analyze direct social-exploration of animals in a social behavior test. Compared to the manual annotation method, this capacitive touch sensing system can be used to instantaneously quantify direct social-exploration, saving significant amount of time of post-hoc video scoring. Furthermore, this low-cost method enhances the objectivity of data by reducing experimenter involvement in analysis.

A wireless miniScope for deep brain imaging in freely moving mice.

BACKGROUND: The increasing interest in the study of neuronal activities at the microcircuit level is motivating neuroscientists and engineers to push the limits in developing miniature in vivo imaging systems. This inter-disciplinary effort led to an increasingly widespread use of wearable miniature microscopes, constantly improving in size, cost, spatial and temporal resolutions, and signal to noise ratio. NEW METHOD: Here we developed a miniature wireless fluorescence microscope (miniScope) that allows recording of brain neural activities at single cell resolution. The wireless miniScope has onboard field-programmable gate array (FPGA) and Micro SD Card storage, and is powered by a battery backpack. RESULTS: Using this wireless miniScope, we simultaneously recorded activities from hundreds of medium spiny neurons (MSNs) in the dorsal striatum of two freely moving mice interacting with each other in an open field, with excellent spatial and temporal resolutions. COMPARISON WITH EXISTING METHODS: Existing miniaturized microscope systems have connecting cables between the microscope sensor and the data acquisition system, consequently limiting the recording to one animal at a time. The wireless miniScope allows simultaneous recording of multiple mice in a group, and could also be applied to freely behaving small primates in the future. CONCLUSION: The wireless miniScope expands the realm of possible behavioral experiments, both by minimizing the repercussions of the cable from the imaging device on the rodent's behavior and by enabling simultaneous in vivo imaging from multiple animals.

A Two-Step GRIN Lens Coating for In Vivo Brain Imaging.

The complex spatial and temporal organization of neural activity in the brain is important for information-processing that guides behavior. Hence, revealing the real-time neural dynamics in freely-moving animals is fundamental to elucidating brain function. Miniature fluorescence microscopes have been developed to fulfil this requirement. With the help of GRadient INdex (GRIN) lenses that relay optical images from deep brain regions to the surface, investigators can visualize neural activity during behavioral tasks in freely-moving animals. However, the application of GRIN lenses to deep brain imaging is severely limited by their availability. Here, we describe a protocol for GRIN lens coating that ensures successful long-term intravital imaging with commercially-available GRIN lenses.

Miniscope GRIN Lens System for Calcium Imaging of Neuronal Activity from Deep Brain Structures in Behaving Animals.

Visualizing neural activity from deep brain regions in freely behaving animals through miniature fluorescent microscope (miniscope) systems is becoming more important for understanding neural encoding mechanisms underlying cognitive functions. Here we present our custom-designed miniscope GRadient INdex (GRIN) lens system that enables simultaneously recording from hundreds of neurons for months. This article includes miniscope design, the surgical procedure for GRIN lens implantation, miniscope mounting on the head of a mouse, and data acquisition and analysis. First, a target brain region is labeled with virus expressing GCaMP6; second, a GRIN lens is implanted above the target brain region; third, following mouse surgical recovery, a miniscope is mounted on the head of the mouse above the GRIN lens; and finally, neural activity is recorded from the freely behaving mouse. This system can be applied to recording the same population of neurons longitudinally, enabling the elucidation of neural mechanisms underlying behavioral control. © 2018 by John Wiley & Sons, Inc.

Distinct and Dynamic ON and OFF Neural Ensembles in the Prefrontal Cortex Code Social Exploration.

The medial prefrontal cortex (mPFC) is important for social behavior, but the mechanisms by which mPFC neurons code real-time social exploration remain largely unknown. Here we utilized miniScopes to record calcium activities from hundreds of excitatory neurons in the mPFC while mice freely explored restrained social targets in the absence or presence of the psychedelic drug phencyclidine (PCP). We identified distinct and dynamic ON and OFF neural ensembles that displayed opposing activities to code real-time behavioral information. We further illustrated that ON and OFF ensembles tuned to social exploration carried information of salience and novelty for social targets. Finally, we showed that dysfunctions in these ensembles were associated with abnormal social exploration elicited by PCP. Our findings underscore the importance of mPFC ON and OFF neural ensembles for proper exploratory behavior, including social exploration, and pave the way for future studies elucidating neural circuit dysfunctions in psychiatric disorders.

Spatially Compact Neural Clusters in the Dorsal Striatum Encode Locomotion Relevant Information.

An influential striatal model postulates that neural activities in the striatal direct and indirect pathways promote and inhibit movement, respectively. Normal behavior requires coordinated activity in the direct pathway to facilitate intended locomotion and indirect pathway to inhibit unwanted locomotion. In this striatal model, neuronal population activity is assumed to encode locomotion relevant information. Here, we propose a novel encoding mechanism for the dorsal striatum. We identified spatially compact neural clusters in both the direct and indirect pathways. Detailed characterization revealed similar cluster organization between the direct and indirect pathways, and cluster activities from both pathways were correlated with mouse locomotion velocities. Using machine-learning algorithms, cluster activities could be used to decode locomotion relevant behavioral states and locomotion velocity. We propose that neural clusters in the dorsal striatum encode locomotion relevant information and that coordinated activities of direct and indirect pathway neural clusters are required for normal striatal controlled behavior. VIDEO ABSTRACT.

Volumetric visualization of the near- and far-field wake in flapping wings.

The flapping wings of flying animals create complex vortex wake structure; understanding its spatial and temporal distribution is fundamental to animal flight theory. In this study, we applied the volumetric 3-component velocimetry to capture both the near- and far-field flow generated by a pair of mechanical flapping wings. For the first time, the complete three-dimensional wake structure and its evolution throughout a wing stroke were quantified and presented experimentally. The general vortex wake structure maintains a quite consistent form: vortex rings in the near field and two shear layers in the far field. Vortex rings shed periodically from the wings and are linked to each other in successive strokes. In the far field, the shed vortex rings evolve into two parallel shear layers with dominant vorticity convected from tip and root vortices. The shear layers are nearly stationary in space compared to the periodic vortex rings shed in the near field. In addition, downwash passes through the centers of the vortex rings and extends downward between the two shear layers.