An integrated workflow for 2D and 3D posture analysis during vestibular system testing in mice

IntroductionPosture extraction from videos is fundamental to many real-world applications, including health screenings. In this study, we extend the utility and specificity of a well-established protocol, the balance beam, for examining balance and active motor coordination in adult mice of both sexes.ObjectivesThe primary objective of this study is to design a workflow for analyzing the postures of mice walking on a balance beam.MethodsWe developed new tools and scripts based on the FluoRender architecture, which can interact with DeepLabCut (DLC) through Python code. Notably, twenty input videos were divided into four feature point groups (head, body, tail, and feet), based on camera positions relative to the balance beam (left and right), and viewing angles (90° and 45° from the beam). We determined key feature points on the mouse to track posture in a still video frame. We extracted a standard walk cycle (SWC) by focusing on foot movements, which were computed by a weighted average of the extracted walk cycles. The correlation of each walk cycle to the SWC was used as the weight.ResultsWe learned that positions of the camera angles significantly improved the performance of 2D pose estimation (90°) and 3D (45°). Comparing the SWCs from age-matched mice, we found a consistent pattern of supporting feet on the beam. Two feet were consistently on the beam followed by three feet and another three feet in a 2-3-3 pattern. However, this pattern can be mirrored among individual subjects. A subtle phase shift of foot movement was also observed from the SWCs. Furthermore, we compared the SWCs with speed values to reveal anomalies in mouse walk postures. Some anomalies can be explained as the start or finish of the traversal, while others may be correlated to the distractions of the test environment, which will need further investigation.ConclusionOur posture analysis workflow improves the classical behavioral testing and analysis, allowing the detection of subtle, but significant differences in vestibular function and motor coordination

A Latent Propriospinal Network Can Restore Diaphragm Function After High Cervical Spinal Cord Injury

Spinal cord injury (SCI) above cervical level 4 disrupts descending axons from the medulla that innervate phrenic motor neurons, causing permanent paralysis of the diaphragm. Using an ex vivo preparation in neonatal mice, we have identified an excitatory spinal network that can direct phrenic motor bursting in the absence of medullary input. After complete cervical SCI, blockade of fast inhibitory synaptic transmission caused spontaneous, bilaterally coordinated phrenic bursting. Here, spinal cord glutamatergic neurons were both sufficient and necessary for the induction of phrenic bursts. Direct stimulation of phrenic motor neurons was insufficient to evoke burst activity. Transection and pharmacological manipulations showed that this spinal network acts independently of medullary circuits that normally generate inspiration, suggesting a distinct non-respiratory function. We further show that this “latent” network can be harnessed to restore diaphragm function after high cervical SCI in adult mice and rats

The Feasibility of Spectral-Domain Optical Coherence Tomography Grading of Anterior Chamber Inflammation in a Rabbit Model of Anterior Uveitis

PURPOSE. To determine the feasibility and accuracy of spectral-domain optical coherence tomography (SD-OCT) based grading of anterior chamber cell, using aqueous sampling as a standard, in a rabbit model of anterior uveitis. METHODS. Adult Dutch-belted rabbits were preimmunized with M. tuberculosis (Tb) H37RA antigen, 1 week prior to induction of anterior uveitis with an intracameral injection of Tb antigen. The anterior chamber was imaged with SD-OCT, followed by a slit lamp examination. Two independent, trained graders recorded their estimate of anterior chamber cell count using the Standardization of Uveitis Nomenclature (SUN) scores for each eye prior to performing an anterior chamber tap to determine the aqueous cell density using a hemocytometer. Using the aqueous cell density as a standard, correlation with SD-OCT counts were compared to those with SUN scores. RESULTS. Overall, SD-OCT correlated well with the hemocytometer counts (Spearman coefficient ¼ 0.53, P < 0.001) compared with SUN grading and hemocytometer counts (Spearman coefficient ¼ 0.02, P ¼ 0.88). The correlation improved to 0.65 (P < 0.001) when we excluded eyes with corneal thickness ‡ 470 lm. Eyes with corneal thickness ‡ 470 lm exhibited the greatest degree of ocular inflammation and corneal opacity. CONCLUSIONS. In our rabbit model, SD-OCT grading of anterior chamber cell correlated significantly better with aqueous cell counts, compared to traditional slit lamp grading. Spectral-domain optical coherence tomography grading of anterior chamber cell may be a good alternative to SUN grading. Although SUN grading remains the clinical gold standard, alternative quantitative methods to assess ocular inflammation could provide insight into disease mechanism and aid in measuring treatment response

Uncovering Voltage-sensing Mechanisms of Neuronal KCNQ Channels and Related Pathological Variants

Neuronal KCNQ channels mediate the M-current, a key regulator of membrane excitability in the nervous system. Mutations in KCNQ2 and KCNQ3 channels cause severe neurodevelopmental disorders, including epileptic encephalopathies, and more recently, autism spectrum disorder (ASD). However, the impact that different mutations have on channel function remains poorly defined, largely because of our limited understanding of the voltage-sensing mechanisms that trigger channel gating. Here, I define the parameters of voltage sensor movements in wt-KCNQ2, wt-KCNQ3, and channels bearing epilepsy- and ASD- associated mutations using cysteine accessibility and voltage clamp fluorometry (VCF). Cysteine modification reveals eight to ten amino acids in the S4 becomes exposed upon voltage sensing domain activation of KCNQ2 and KCNQ3 channels. VCF shows that the voltage dependence and the time course of S4 movement and channel opening/closing closely correlate. VCF reveals different mechanisms by which disease-causing&nbsp;mutations affect KCNQ2 channel voltage-dependent gating, while also revealing the mechanisms behind S4 activation mutated channels. Further, polyunsaturated fatty acid compounds have been shown to have therapeutic benefits for cardiovascular disease,&nbsp;and recently&nbsp;neurological disorders, because of their ability to modulate voltage-gated ion channels. Polyunsaturated fatty acids are introduced to the body through diet, and infiltrate&nbsp;the cardiovascular system and&nbsp;the central and peripheral nervous system. Here, I screen a variety of PUFAs on KCNQ3 channels to find candidates that modulate KCNQ3 activity and could correct neurological disease-associated mutations. These results show that N-arachidonoyl Taurine and Arachidonic Amine could be promising candidates to correct for different mutational defects in ion channel functionality.&nbsp;This study provides novel insight into KCNQ2 and KCNQ3 channel function in physiological and pathophysiological conditions, which will further improve therapies for devasting neurological channelopathies.</p

Distinctive mechanisms of epilepsy-causing mutants discovered by measuring S4 movement in KCNQ2 channels

Neuronal KCNQ channels mediate the M-current, a key regulator of membrane excitability in the central and peripheral nervous systems. Mutations in KCNQ2 channels cause severe neurodevelopmental disorders, including epileptic encephalopathies. However, the impact that different mutations have on channel function remains poorly defined, largely because of our limited understanding of the voltage-sensing mechanisms that trigger channel gating. Here, we define the parameters of voltage sensor movements in wt-KCNQ2 and channels bearing epilepsy-associated mutations using cysteine accessibility and voltage clamp fluorometry (VCF). Cysteine modification reveals that a stretch of eight to nine amino acids in the S4 becomes exposed upon voltage sensing domain activation of KCNQ2 channels. VCF shows that the voltage dependence and the time course of S4 movement and channel opening/closing closely correlate. VCF reveals different mechanisms by which different epilepsy-associated mutations affect KCNQ2 channel voltage-dependent gating. This study provides insight into KCNQ2 channel function, which will aid in uncovering the mechanisms underlying channelopathies

Progress on Open Science: Towards a Shared Research Knowledge System. Final Report of the Open Science Policy Platform

This final report of the EU Open Science Policy Platform (OSPP) provides a brief overview of its four-year mandate from 2016 to 2020, followed by an update on progress by each stakeholder group over the past two years since the publication of the OSPP’s recommendations across the European Commission’s eight ambitions on Open Science, (OSPP-REC1). This summary of Practical Commitments for Implementation with specific examples of progress by each stakeholder community across Europe (see Annex A) is followed by a perspective from each group on the major outstanding blockers to progress and possible next steps. The group of 25 key stakeholder representatives have then come together to propose a vision for moving beyond Open Science to create a shared research knowledge system by 2030

Improved electro-destruction of bacterial biofilms by coating conductive surfaces with polymers

International audienc

Influence of the apo A5 gene on fat-soluble vitamins

Workshops W6 Regulation and role of APOAV in lipoprotein metabolism : Workshop W06-P-005International audienc