GABAA receptor subunit composition regulates circadian rhythms in rest-wake and synchrony among cells in the suprachiasmatic nucleus.

The mammalian circadian clock located in the suprachiasmatic nucleus (SCN) produces robust daily rhythms including rest-wake. SCN neurons synthesize and respond to γ-aminobutyric acid (GABA), but its role remains unresolved. We tested the hypothesis that γ2- and δ-subunits of the GABAA receptor in the SCN differ in their regulation of synchrony among circadian cells. We used two approaches: 1) shRNA to knock-down (KD) the expression of either γ2 or δ subunits in the SCN or 2) knock-in mice harboring a point mutation in the M2 domains of the endogenous GABAA γ2 or δ subunits. KD of either γ2 or δ subunits in the SCN increased daytime running and reduced nocturnal running by reducing their circadian amplitude by a third. Similarly, δ subunit knock-in mice showed decreased circadian amplitude, increased duration of daily activity, and decreased total daily activity. Reduction, or mutation of either γ2 or δ subunits halved the synchrony among, and amplitude of, circadian SCN cells as measured by firing rate or expression of the PERIOD2 protein, in vitro. Surprisingly, overexpression of the γ2 subunit rescued these phenotypes following KD or mutation of the δ subunit, and overexpression of the δ subunit rescued deficiencies due to γ2 subunit KD or mutation. We conclude that γ2 and δ GABAA receptor subunits play similar roles in maintaining circadian synchrony in the SCN and amplitude of daily rest-wake rhythms, but that modulation of their relative densities can change the duration and amplitude of daily activities.

Global siRNA Screen Reveals Critical Human Host Factors of SARS-CoV-2 Multicycle Replication.

Defining the subset of cellular factors governing SARS-CoV-2 replication can provide critical insights into viral pathogenesis and identify targets for host-directed antiviral therapies. While a number of genetic screens have previously reported SARS-CoV-2 host dependency factors, these approaches relied on utilizing pooled genome-scale CRISPR libraries, which are biased towards the discovery of host proteins impacting early stages of viral replication. To identify host factors involved throughout the SARS-CoV-2 infectious cycle, we conducted an arrayed genome-scale siRNA screen. Resulting data were integrated with published datasets to reveal pathways supported by orthogonal datasets, including transcriptional regulation, epigenetic modifications, and MAPK signalling. The identified proviral host factors were mapped into the SARS-CoV-2 infectious cycle, including 27 proteins that were determined to impact assembly and release. Additionally, a subset of proteins were tested across other coronaviruses revealing 17 potential pan-coronavirus targets. Further studies illuminated a role for the heparan sulfate proteoglycan perlecan in SARS-CoV-2 viral entry, and found that inhibition of the non-canonical NF-kB pathway through targeting of BIRC2 restricts SARS-CoV-2 replication both in vitro and in vivo. These studies provide critical insight into the landscape of virus-host interactions driving SARS-CoV-2 replication as well as valuable targets for host-directed antivirals.

Inversion recovery and saturation recovery pulmonary vein MR angiography using an image based navigator fluoro trigger and variable-density 3D cartesian sampling with spiral-like order.

Contrast enhanced pulmonary vein magnetic resonance angiography (PV CE-MRA) has value in atrial ablation pre-procedural planning. We aimed to provide high fidelity, ECG gated PV CE-MRA accelerated by variable density Cartesian sampling (VD-CASPR) with image navigator (iNAV) respiratory motion correction acquired in under 4 min. We describe its use in part during the global iodinated contrast shortage. VD-CASPR/iNAV framework was applied to ECG-gated inversion and saturation recovery gradient recalled echo PV CE-MRA in 65 patients (66 exams) using .15 mmol/kg Gadobutrol. Image quality was assessed by three physicians, and anatomical segmentation quality by two technologists. Left atrial SNR and left atrial/myocardial CNR were measured. 12 patients had CTA within 6 months of MRA. Two readers assessed PV ostial measurements versus CTA for intermodality/interobserver agreement. Inter-rater/intermodality reliability, reproducibility of ostial measurements, SNR/CNR, image, and anatomical segmentation quality was compared. The mean acquisition time was 3.58 ± 0.60 min. Of 35 PV pre-ablation datasets (34 patients), mean anatomical segmentation quality score was 3.66 ± 0.54 and 3.63 ± 0.55 as rated by technologists 1 and 2, respectively (p = 0.7113). Good/excellent anatomical segmentation quality (grade 3/4) was seen in 97% of exams. Each rated one exam as moderate quality (grade 2). 95% received a majority image quality score of good/excellent by three physicians. Ostial PV measurements correlated moderate to excellently with CTA (ICCs range 0.52-0.86). No difference in SNR was observed between IR and SR. High quality PV CE-MRA is possible in under 4 min using iNAV bolus timing/motion correction and VD-CASPR.

Kv12-encoded K+ channels drive the day-night switch in the repetitive firing rates of SCN neurons.

Considerable evidence suggests that day-night rhythms in the functional expression of subthreshold potassium (K+) channels regulate daily oscillations in the spontaneous firing rates of neurons in the suprachiasmatic nucleus (SCN), the master circadian pacemaker in mammals. The K+ conductance(s) driving these daily rhythms in the repetitive firing rates of SCN neurons, however, have not been identified. To test the hypothesis that subthreshold Kv12.1/Kv12.2-encoded K+ channels play a role, we obtained current-clamp recordings from SCN neurons in slices prepared from adult mice harboring targeted disruptions in the Kcnh8 (Kv12.1-/-) or Kcnh3 (Kv12.2-/-) locus. We found that mean nighttime repetitive firing rates were higher in Kv12.1-/- and Kv12.2-/- than in wild type (WT), SCN neurons. In marked contrast, mean daytime repetitive firing rates were similar in Kv12.1-/-, Kv12.2-/-, and WT SCN neurons, and the day-night difference in mean repetitive firing rates, a hallmark feature of WT SCN neurons, was eliminated in Kv12.1-/- and Kv12.2-/- SCN neurons. Similar results were obtained with in vivo shRNA-mediated acute knockdown of Kv12.1 or Kv12.2 in adult SCN neurons. Voltage-clamp experiments revealed that Kv12-encoded current densities in WT SCN neurons are higher at night than during the day. In addition, the pharmacological block of Kv12-encoded currents increased the mean repetitive firing rate of nighttime, but not daytime, in WT SCN neurons. Dynamic clamp-mediated subtraction of modeled Kv12-encoded currents also selectively increased the mean repetitive firing rates of nighttime WT SCN neurons. Despite the elimination of the nighttime decrease in the mean repetitive firing rates of SCN neurons, however, locomotor (wheel-running) activity remained rhythmic in Kv12.1-/-, Kv12.2-/-, and Kv12.1-targeted shRNA-expressing, and Kv12.2-targeted shRNA-expressing animals.

Kv12-Encoded K + Channels Drive the Day-Night Switch in the Repetitive Firing Rates of SCN Neurons.

Considerable evidence suggests that day-night rhythms in the functional expression of subthreshold potassium (K + ) channels regulate daily oscillations in the rates of spontaneous action potential firing of neurons in the suprachiasmatic nucleus (SCN), the master circadian pacemaker in mammals. The K + conductance(s) driving these daily rhythms in repetitive firing rates, however, have not been identified. To test the hypothesis that subthreshold Kv12.1/Kv12.2-encoded K + channels play a role, we obtained current-clamp recordings from SCN neurons in slices prepared from adult mice harboring targeted disruptions in the Kcnh8 (Kv12.1 -/- ) or Kcnh3 (Kv12.2 -/- ) locus. We found that mean nighttime repetitive firing rates were higher in Kv12.1 -/- and Kv12.2 -/- , than in wild type (WT), SCN neurons. In marked contrast, mean daytime repetitive firing rates were similar in Kv12.1 -/- , Kv12.2 -/- and WT SCN neurons, and the day-night difference in mean repetitive firing rates, a hallmark feature of WT SCN neurons, was eliminated in Kv12.1 -/- and Kv12.2 -/- SCN neurons. Similar results were obtained with in vivo shRNA-mediated acute knockdown of Kv12.1 or Kv12.2 in adult SCN neurons. Voltage-clamp experiments revealed that Kv12-encoded current densities in WT SCN neurons are higher at night than during the day. In addition, pharmacological block of Kv12-encoded currents increased the mean repetitive firing rate of nighttime, but not daytime, in WT SCN neurons. Dynamic clamp-mediated subtraction of modeled Kv12-encoded currents also selectively increased the mean repetitive firing rates of nighttime WT SCN neurons. Despite the elimination of nighttime decrease in the mean repetitive firing rates of SCN neurons, however, locomotor (wheel-running) activity remained rhythmic in Kv12.1 -/- , Kv12.2 -/- , Kv12.1-targeted shRNA-expressing, and Kv12.2-targeted shRNA-expressing animals.

Quantifying organellar ultrastructure in cryo-electron tomography using a surface morphometrics pipeline.

Cellular cryo-electron tomography (cryo-ET) enables three-dimensional reconstructions of organelles in their native cellular environment at subnanometer resolution. However, quantifying ultrastructural features of pleomorphic organelles in three dimensions is challenging, as is defining the significance of observed changes induced by specific cellular perturbations. To address this challenge, we established a semiautomated workflow to segment organellar membranes and reconstruct their underlying surface geometry in cryo-ET. To complement this workflow, we developed an open-source suite of ultrastructural quantifications, integrated into a single pipeline called the surface morphometrics pipeline. This pipeline enables rapid modeling of complex membrane structures and allows detailed mapping of inter- and intramembrane spacing, curvedness, and orientation onto reconstructed membrane meshes, highlighting subtle organellar features that are challenging to detect in three dimensions and allowing for statistical comparison across many organelles. To demonstrate the advantages of this approach, we combine cryo-ET with cryo-fluorescence microscopy to correlate bulk mitochondrial network morphology (i.e., elongated versus fragmented) with membrane ultrastructure of individual mitochondria in the presence and absence of endoplasmic reticulum (ER) stress. Using our pipeline, we demonstrate ER stress promotes adaptive remodeling of ultrastructural features of mitochondria including spacing between the inner and outer membranes, local curvedness of the inner membrane, and spacing between mitochondrial cristae. We show that differences in membrane ultrastructure correlate to mitochondrial network morphologies, suggesting that these two remodeling events are coupled. Our pipeline offers opportunities for quantifying changes in membrane ultrastructure on a single-cell level using cryo-ET, opening new opportunities to define changes in ultrastructural features induced by diverse types of cellular perturbations.

Hyperammonaemia disrupts daily rhythms reversibly by elevating glutamate in the central circadian pacemaker.

Patients with cirrhosis exhibit features of circadian disruption. Hyperammonaemia has been suggested to impair both homeostatic and circadian sleep regulation. Here, we tested if hyperammonaemia directly disrupts circadian rhythm generation in the central pacemaker, the suprachiasmatic nuclei (SCN) of the hypothalamus. Wheel-running activity was recorded from mice fed with a hyperammonaemic or normal diet for ~35 days in a 12:12 light-dark (LD) cycle followed by ~15 days in constant darkness (DD). The expression of the clock protein PERIOD2 (PER2) was recorded from SCN explants before, during and after ammonia exposure, ±glutamate receptor antagonists. In LD, hyperammonaemic mice advanced their daily activity onset time by ~1 h (16.8 ± 0.3 vs. 18.1 ± 0.04 h, p = .009) and decreased their total activity, concentrating it during the first half of the night. In DD, hyperammonaemia reduced the amplitude of daily activity (551.5 ± 27.7 vs. 724.9 ± 59 counts, p = .007), with no changes in circadian period. Ammonia (≥0.01 mM) rapidly and significantly reduced PER2 amplitude, and slightly increased circadian period. The decrease in PER2 amplitude correlated with decreased synchrony among circadian cells in the SCN and increased extracellular glutamate, which was rescued by AMPA glutamate receptor antagonists. These data suggest that hyperammonaemia affects circadian regulation of rest-activity behaviour by increasing extracellular glutamate in the SCN.

Health Economics of Selective Early Rescue Surfactant Administration and Standard Surfactant Administration for Newborns With Respiratory Distress Syndrome.

OBJECTIVE: To quantify the effect of early rescue surfactant administration techniques for preterm infants with respiratory distress syndrome (RDS) from a health care delivery system perspective. METHODS: A cost-consequence model was developed based on previously published literature to compare the health economic impact of implementing early surfactant administration strategies vs standard surfactant administration via endotracheal intubation and mechanical ventilation (MV). RESULTS: Early rescue surfactant treatment strategies are associated with a decrease in the number of patients requiring MV, cumulative MV days, and rate of neonatal complications. Total annual surfactant costs are higher than standard surfactant administration, but this is offset by savings in total hospital and complication costs. CONCLUSIONS: This cost-consequence analysis suggests selective early rescue surfactant administration strategies are associated with a lower health care burden in premature infants with RDS.

[Depression, anxiety and quality of life in pediatric patients with leukemia]. / Depresión, ansiedad y calidad de vida en pacientes pediátricos con leucemia.

Background: Leukemia is the most frequent cancer in children and adolescents, and it has a high prevalence of depression and anxiety which deteriorates the quality of life related to health. The symptoms of depression and anxiety may go unnoticed by the physician as a normal response during cancer treatment. Objective: To determine the prevalence of depression, anxiety and health-related quality of life in pediatric patients with leukemia. Material and methods: study with the participation of Mexican children and adolescents with leukemia whose depression was determined with the Childhood Depression Inventory, their anxiety with the Spence Childhood Anxiety Scale, and their health-related quality of life (HRQoL) with PedsQL 4.0. Results: 37 participants, with a median age of 11 years (8-14 years); 19 (51.4%) were male. The marital status of the parents in 25 participants (67.5%) was married, in 10 (27%) had a domestic partnership, in one (2.7%) had divorced parents and in one it was single (2.7%). The prevailing religion was Catholic in 29 (78.3%); 16 patients (43.2%) reported depression, 10% anxiety and 94.5% reported an adequate health-related quality of life, with an average of 74.2 +- 16.2. Conclusions: Depression was the most prevalent, followed by anxiety. Health-related quality of life was reported as good. The harmful impact is still prevalent in a vulnerable population, which must be attended in a comprehensive and timely manner at all levels of care.

Introducción: la leucemia es el cáncer más frecuente en niños y adolescentes, y tiene una alta prevalencia de depresión y ansiedad que deterioran la calidad de vida relacionada con la salud. Los síntomas de depresión y ansiedad pueden pasar inadvertidos por el médico al considerar que son una respuesta normal durante el tratamiento del cáncer. Objetivo: determinar la prevalencia de depresión, ansiedad y calidad de vida relacionada con la salud en pacientes pediátricos con leucemia. Material y métodos: estudio en el que participaron niños y adolescentes mexicanos con leucemia cuya depresión se estableció con el Inventario de Depresión Infantil, su ansiedad con la Escala de Ansiedad Infantil Spence y su calidad de vida relacionada con la salud (CVRS) con el PedsQL 4.0. Resultados: fueron 37 participantes, con una mediana de edad de 11 años (8-14 años); 19 (51.4%) fueron del género masculino. Los padres de 25 pacientes (67.5%) estaban casados, los de 10 (27%) en unión libre, el de uno estaba divorciado (2.7%) y el de otro soltero (2.7%). La religión prevalente fue la católica en 29 (78.3%); 16 pacientes (43.2%) reportaron depresión, 10% ansiedad y 94.5% reportó adecuada calidad de vida relacionada con la salud, con un promedio de 74.2 +- 16.2. Conclusiones: la depresión fue la más prevalente, seguida de la ansiedad; la calidad de vida relacionada con la salud se reportó como buena. El impacto nocivo aún sigue siendo prevalente en una población vulnerable, la cual se debe atender de manera integral y oportuna en todos los niveles de atención.

[Confinement and social distancing: stress, anxiety, depression in children and adolescents]. / Confinamiento y distanciamiento social: estrés, ansiedad, depresión en niños y adolescentes.

Among the strategies to avoid COVID-19 disease that affects the world population it is keeping away from contagion through confinement and social distancing, which can cause in children and adolescents a change in the psychosocial environment with a high prevalence of psychological and psychiatric disorders, such as stress, anxiety disorders, depressive disorders, risk of suicide and eating disorders. During confinement, were made school closures, shutdowns of recreational and sports spaces, cancellation of outdoor activities, which can have consequences in social habits, sleep, modifications in the eating pattern that entails alterations in anthropometric variables, neurophysiological alterations with behaviors that affect mental health, in addition to the frustration of not executing plans, fear, boredom and the uncertainty of the duration of the pandemic could lead to further stress. Social distancing favors a reduction in interaction, is related to anxiety, stress and depression, in addition to the concern of infecting the family. These scenarios can generate stress for parents, domestic violence and uncontrolled use of the internet. Children and adolescents are vulnerable populations that adapt to the environment in crisis. The objective of this work was to review the influence of confinement and social distancing on stress, anxiety and depression in children and adolescents.

Entre las estrategias para evitar la enfermedad por COVID-19 que afecta a la población mundial está evitar el contagio a partir del confinamiento y el distanciamiento social, lo que puede ocasionar en niños y adolescentes un cambio en el entorno psicosocial con alta prevalencia de alteraciones psicológicas y psiquiátricas, como estrés, transtornos de ansiedad, transtornos depresivos, riesgo de suicidio y trastornos de la conducta alimentaria. Durante el confinamiento se realizaron cierres de escuelas, de espacios recreativos y deportivos, cancelación de actividades fuera de casa, lo cual puede tener consecuencias en los hábitos sociales, del sueño, modificaciones en el patrón de alimentación que conllevan alteraciones en las variables antropométricas, alteraciones neurofisiológicas conductuales que afectan la salud mental, además de la frustración de la no ejecución de planes, miedo, aburrimiento y la incertidumbre de la duración, los cuales podrían generar mayor estrés. El distanciamiento social propicia una reducción de la interacción y se relaciona con ansiedad, estrés y depresión, además de la preocupación de contagiar a la familia. Estos escenarios pueden generar estrés en los padres, violencia intrafamiliar y uso descontrolado de internet. Los niños y adolescentes son población vulnerable que se adapta al entorno en crisis generadas por el confinamiento y distanciamiento social. El objetivo de este trabajo fue revisar la influencia del confinamiento y el distanciamiento social en el estrés, la ansiedad y la depresión de niños y adolescentes.

Intento de suicidio con metanfetamina endovenosa y chemsex / Attempted Suicide with Intravenous Methamphetamine and Chemsex

RESUMEN Introducción: El trastorno de déficit de atención e hiperactividad no diagnosticado es una condición comórbida frecuente en los trastornos por abuso de sustancias, y tiene un importante impacto en la vida de los pacientes, pues incrementa el riesgo de abuso de sustancias y el riesgo derivado de conductas impulsivas. En el momento actual no existen trabajos que estudien la relación entre el TDAH y el fenómeno emergente del chemsex, es decir, el uso de sustancias psicoactivas en el contexto de fiestas sexuales para maximizar el tiempo de práctica sexual y de las experiencias subjetivas. En estas sesiones se usan diferentes sustancias como, por ejemplo, las catinonas sintéticas como la mefedrona, que podría inducir complicaciones médicas y psiquiátricas como psicosis, agresividad e ideación suicida. Caso: Un varón de 44 arios ingresó en la unidad de agudos de psiquiatría tras un segundo intento de suicidio de alto riesgo con el uso de metanfetamina endovenosa mezclada con un ácido queratolítico, en el contexto de un episodio depresivo y práctica activa de sesiones de chemsex con una frecuencia semanal, en las que tomaba mefedrona vía transrectal, que provoca episodios puntuales de psicosis tóxica autolimatada. Se objetivó clínica compatible con TDAH no diagnosticado, y se aplicaron ASRS-V1.1 y WURS, que indicaron TDAH. Se ajustó el tratamiento antidepresivo y remitieron los síntomas depresivos y la ideación autolítica. Discusión: La búsqueda de síntomas de TDAH «ocultos¼ en el contexto del abuso de sustancias es importante para no retrasar diagnósticos con importancia esencial y prevenir complicaciones. Complicaciones médicas y psiquiátricas relacionadas con la práctica del chemsex se han comunicado en Europa y recientemente en España, donde es un fenómeno emergente que preocupa. Se describen el intento de suicidio con metanfetamina, síntomas depresivos y psicóticos, abuso de sustancias y complicaciones médicas.

ABSTRACT Background: Undiagnosed attention deficit and hyperactivity disorder (ADHD) is frequent in the substance abuse disorder population, and has an intense repercussion in the daily life of the patients. That condition increases the risk of substance abuse disorders and risk conducts derived from impulsivity. There are not yet studies linking ADHD and the new emergent phenomenon of chemsex, that is, the action of use the intravenous injection of substances in sex parties, with the objective of maximising the practising time and sexual experience. In these sessions, different types of drug are used, for example, synthetic cathinone or mephedrone, and may lead to diverse medical and psychiatric complications like psychosis, aggressiveness and suicide ideation. Case: We report the case of a 44-year-old man admitted into a psychiatric unit, presenting with 2nd time suicidal high risk attempt using intravenous methamphetamine and a dissolvent acid in the context of a depressive episode, after practising chemsex sessions almost every weekend with psychoactive substances taken orally, intravenously and intrarectally. The patient also presented psychotic symptoms from the use of these drugs. When we interviewed the patient during his hospital stay, we observed that there were symptoms of a possible attention and hyperactivity deficit, which was confirmed by study with ASRS-V1.1 and WURS scales. After three weeks, the patient improved substantially as his depressive symptoms and autolytic ideation disappeared. Discussion: Look for "hidden" symptoms in substance abuse disorder patients. It is impor-tant not to delay important diagnostic tests and to prevent complications. Psychiatric and medical conditions related to chemsex have been reported in several European cities, and recently in Spain where it is an emergent phenomenon and a problem concerning medical and political institutions. The suicide attempt with metamphetamine and acid taken intravenously in the case we report, depressive and psychotic symptoms, addiction, suicide attempts and medical complications are described.

Attempted suicide with intravenous methamphetamine and chemsex.

BACKGROUND: Undiagnosed attention deficit and hyperactivity disorder (ADHD) is frequent in the substance abuse disorder population, and has an intense repercussion in the daily life of the patients. That condition increases the risk of substance abuse disorders and risk conducts derived from impulsivity. There are not yet studies linking ADHD and the new emergent phenomenon of chemsex, that is, the action of use the intravenous injection of substances in sex parties, with the objective of maximising the practising time and sexual experience. In these sessions, different types of drug are used, for example, synthetic cathinone or mephedrone, and may lead to diverse medical and psychiatric complications like psychosis, aggressiveness and suicide ideation. CASE: We report the case of a 44-year-old man admitted into a psychiatric unit, presenting with 2nd time suicidal high risk attempt using intravenous methamphetamine and a dissolvent acid in the context of a depressive episode, after practising chemsex sessions almost every weekend with psychoactive substances taken orally, intravenously and intrarectally. The patient also presented psychotic symptoms from the use of these drugs. When we interviewed the patient during his hospital stay, we observed that there were symptoms of a possible attention and hyperactivity deficit, which was confirmed by study with ASRS-V1.1 and WURS scales. After three weeks, the patient improved substantially as his depressive symptoms and autolytic ideation disappeared. DISCUSSION: Look for "hidden" symptoms in substance abuse disorder patients. It is important not to delay important diagnostic tests and to prevent complications. Psychiatric and medical conditions related to chemsex have been reported in several European cities, and recently in Spain where it is an emergent phenomenon and a problem concerning medical and political institutions. The suicide attempt with metamphetamine and acid taken intravenously in the case we report, depressive and psychotic symptoms, addiction, suicide attempts and medical complications are described.

CellPAINT: Turnkey Illustration of Molecular Cell Biology.

CellPAINT is an interactive digital tool that allows non-expert users to create illustrations of the molecular structure of cells and viruses. We present a new release with several key enhancements, including the ability to generate custom ingredients from structure information in the Protein Data Bank, and interaction, grouping, and locking functions that streamline the creation of assemblies and illustration of large, complex scenes. An example of CellPAINT as a tool for hypothesis generation in the interpretation of cryoelectron tomograms is presented. CellPAINT is freely available at http://ccsb.scripps.edu/cellpaint.

Circadian neurons in the paraventricular nucleus entrain and sustain daily rhythms in glucocorticoids.

Signals from the central circadian pacemaker, the suprachiasmatic nucleus (SCN), must be decoded to generate daily rhythms in hormone release. Here, we hypothesized that the SCN entrains rhythms in the paraventricular nucleus (PVN) to time the daily release of corticosterone. In vivo recording revealed a critical circuit from SCN vasoactive intestinal peptide (SCNVIP)-producing neurons to PVN corticotropin-releasing hormone (PVNCRH)-producing neurons. PVNCRH neurons peak in clock gene expression around midday and in calcium activity about three hours later. Loss of the clock gene Bmal1 in CRH neurons results in arrhythmic PVNCRH calcium activity and dramatically reduces the amplitude and precision of daily corticosterone release. SCNVIP activation reduces (and inactivation increases) corticosterone release and PVNCRH calcium activity, and daily SCNVIP activation entrains PVN clock gene rhythms by inhibiting PVNCRH neurons. We conclude that daily corticosterone release depends on coordinated clock gene and neuronal activity rhythms in both SCNVIP and PVNCRH neurons.

SAR.IoT: Secured Augmented Reality for IoT Devices Management.

Currently, solutions based on the Internet of Things (IoT) concept are increasingly being adopted in several fields, namely, industry, agriculture, and home automation. The costs associated with this type of equipment is reasonably small, as IoT devices usually do not have output peripherals to display information about their status (e.g., a screen or a printer), although they may have informative LEDs, which is sometimes insufficient. For most IoT devices, the price of a minimalist display, to output and display the device's running status (i.e., what the device is doing), might cost much more than the actual IoT device. Occasionally, it might become necessary to visualize the IoT device output, making it necessary to find solutions to show the hardware output information in real time, without requiring extra equipment, only what the administrator usually has with them. In order to solve the above, a technological solution that allows for the visualization of IoT device information in actual time, using augmented reality and a simple smartphone, was developed and analyzed. In addition, the system created integrates a security layer, at the level of AR, to secure the shown data from unwanted eyes. The results of the tests carried out allowed us to validate the operation of the solution when accessing the information of the IoT devices, verify the operation of the security layer in AR, analyze the interaction between smartphones, the platform, and the devices, and check which AR markers are most optimized for this use case. This work results in a secure augmented reality solution, which can be used with a simple smartphone, to monitor/manage IoT devices in industrial, laboratory or research environments.

IndoorCare: Low-Cost Elderly Activity Monitoring System through Image Processing.

The Portuguese population is aging at an increasing rate, which introduces new problems, particularly in rural areas, where the population is small and widely spread throughout the territory. These people, mostly elderly, have low income and are often isolated and socially excluded. This work researches and proposes an affordable Ambient Assisted Living (AAL)-based solution to monitor the activities of elderly individuals, inside their homes, in a pervasive and non-intrusive way, while preserving their privacy. The solution uses a set of low-cost IoT sensor devices, computer vision algorithms and reasoning rules, to acquire data and recognize the activities performed by a subject inside a home. A conceptual architecture and a functional prototype were developed, the prototype being successfully tested in an environment similar to a real case scenario. The system and the underlying concept can be used as a building block for remote and distributed elderly care services, in which the elderly live autonomously in their homes, but have the attention of a caregiver when needed.

Modelling the functional roles of synaptic and extra-synaptic γ-aminobutyric acid receptor dynamics in circadian timekeeping.

In the suprachiasmatic nucleus (SCN), γ-aminobutyric acid (GABA) is a primary neurotransmitter. GABA can signal through two types of GABAA receptor subunits, often referred to as synaptic GABAA (gamma subunit) and extra-synaptic GABAA (delta subunit). To test the functional roles of these distinct GABAA in regulating circadian rhythms, we developed a multicellular SCN model where we could separately compare the effects of manipulating GABA neurotransmitter or receptor dynamics. Our model predicted that blocking GABA signalling modestly increased synchrony among circadian cells, consistent with published SCN pharmacology. Conversely, the model predicted that lowering GABAA receptor density reduced firing rate, circadian cell fraction, amplitude and synchrony among individual neurons. When we tested these predictions, we found that the knockdown of delta GABAA reduced the amplitude and synchrony of clock gene expression among cells in SCN explants. The model further predicted that increasing gamma GABAA densities could enhance synchrony, as opposed to increasing delta GABAA densities. Overall, our model reveals how blocking GABAA receptors can modestly increase synchrony, while increasing the relative density of gamma over delta subunits can dramatically increase synchrony. We hypothesize that increased gamma GABAA density in the winter could underlie the tighter phase relationships among SCN cells.

Antibody-Based Inhibition of Pathogenic New World Hemorrhagic Fever Mammarenaviruses by Steric Occlusion of the Human Transferrin Receptor 1 Apical Domain.

Pathogenic clade B New World mammarenaviruses (NWM) can cause Argentine, Venezuelan, Brazilian, and Bolivian hemorrhagic fevers. Sequence variability among NWM glycoproteins (GP) poses a challenge to the development of broadly neutralizing therapeutics against the entire clade of viruses. However, blockade of their shared binding site on the apical domain of human transferrin receptor 1 (hTfR1/CD71) presents an opportunity for the development of effective and broadly neutralizing therapeutics. Here, we demonstrate that the murine monoclonal antibody OKT9, which targets the apical domain of hTfR1, can sterically block cellular entry by viral particles presenting clade B NWM glycoproteins (GP1-GP2). OKT9 blockade is also effective against viral particles pseudotyped with glycoproteins of a recently identified pathogenic Sabia-like virus. With nanomolar affinity for hTfR1, the OKT9 antigen binding fragment (OKT9-Fab) sterically blocks clade B NWM-GP1s and reduces infectivity of an attenuated strain of Junin virus. Binding of OKT9 to the hTfR1 ectodomain in its soluble, dimeric state produces stable assemblies that are observable by negative-stain electron microscopy. A model of the OKT9-sTfR1 complex, informed by the known crystallographic structure of sTfR1 and a newly determined structure of the OKT9 antigen binding fragment (Fab), suggests that OKT9 and the Machupo virus GP1 share a binding site on the hTfR1 apical domain. The structural basis for this interaction presents a framework for the design and development of high-affinity, broadly acting agents targeting clade B NWMs. IMPORTANCE Pathogenic clade B NWMs cause grave infectious diseases, the South American hemorrhagic fevers. Their etiological agents are Junin (JUNV), Guanarito (GTOV), Sabiá (SABV), Machupo (MACV), Chapare (CHAV), and a new Sabiá-like (SABV-L) virus recently identified in Brazil. These are priority A pathogens due to their high infectivity and mortality, their potential for person-to-person transmission, and the limited availability of effective therapeutics and vaccines to curb their effects. While low homology between surface glycoproteins of NWMs foils efforts to develop broadly neutralizing therapies targeting NWMs, this work provides structural evidence that OKT9, a monoclonal antibody targeting a single NWM glycoprotein binding site on hTfR1, can efficiently prevent their entry into cells.