Altered cortical processing of sensory input in Huntington disease mouse models.

Huntington disease (HD), a hereditary neurodegenerative disorder, manifests as progressively impaired movement and cognition. Although early abnormalities of neuronal activity in striatum are well established in HD models, there are fewer in vivo studies of the cortex. Here, we record local field potentials (LFPs) in YAC128 HD model mice versus wild-type mice. In multiple cortical areas, limb sensory stimulation evokes a greater change in LFP power in YAC128 mice. Mesoscopic imaging using voltage-sensitive dyes reveals more extensive spread of evoked sensory signals across the cortical surface in YAC128 mice. YAC128 layer 2/3 sensory cortical neurons ex vivo show increased excitatory events, which could contribute to enhanced sensory responses in vivo. Cortical LFP responses to limb stimulation, visual and auditory input are also significantly increased in zQ175 HD mice. Results presented here extend knowledge of HD beyond ex vivo studies of individual neurons to the intact cortical network.

Podocalyxin is required for maintaining blood-brain barrier function during acute inflammation.

Podocalyxin (Podxl) is broadly expressed on the luminal face of most blood vessels in adult vertebrates, yet its function on these cells is poorly defined. In the present study, we identified specific functions for Podxl in maintaining endothelial barrier function. Using electrical cell substrate impedance sensing and live imaging, we found that, in the absence of Podxl, human umbilical vein endothelial cells fail to form an efficient barrier when plated on several extracellular matrix substrates. In addition, these monolayers lack adherens junctions and focal adhesions and display a disorganized cortical actin cytoskeleton. Thus, Podxl has a key role in promoting the appropriate endothelial morphogenesis required to form functional barriers. This conclusion is further supported by analyses of mutant mice in which we conditionally deleted a floxed allele of Podxl in vascular endothelial cells (vECs) using Tie2Cre mice (PodxlΔTie2Cre). Although we did not detect substantially altered permeability in naïve mice, systemic priming with lipopolysaccharide (LPS) selectively disrupted the blood-brain barrier (BBB) in PodxlΔTie2Cre mice. To study the potential consequence of this BBB breach, we used a selective agonist (TFLLR-NH2) of the protease-activated receptor-1 (PAR-1), a thrombin receptor expressed by vECs, neuronal cells, and glial cells. In response to systemic administration of TFLLR-NH2, LPS-primed PodxlΔTie2Cre mice become completely immobilized for a 5-min period, coinciding with severely dampened neuroelectric activity. We conclude that Podxl expression by CNS tissue vECs is essential for BBB maintenance under inflammatory conditions.

Characterizing observable COVID-19 controls in Pacific Northwest grocery stores.

Utilizing a longitudinal, observational study, grocery store health and safety controls implemented during the COVID-19 pandemic across stores in two cities were characterized. Sixteen stores between Seattle, WA (n = 9) and Portland, OR (n = 7) were visited monthly by the study team from May 2020 to January 2021, and observations of controls were recorded using a standardized checklist in REDCap. The checklist included questions on the presence or absence of controls such as physical barriers, social distancing markers, required masking of customers, cleaning of check-out stands, and closures of store areas. Descriptive analyses were conducted to determine the proportion of stores with a certain control each month. Mixed-effects logistic regression was used to explore how controls changed over time, and whether differences were observed between cities or by income of the area the store serves. Source control (e.g., mask requirements) and engineering controls (e.g., physical barriers at checkout) were the most common and consistent controls observed across stores and over the study period. Controls such as having special hours for vulnerable populations, demarcations on aisles for directionality, and cleaning check-out stands between customers varied significantly over time (p < 0.05 in the mixed-effects model). Having an employee present to clean baskets and carts, as well as physical barriers between self-checkouts, were significantly more common in stores in areas above the median income (p < 0.05 in the mixed-effects model). To best protect workers and shoppers from infectious agents, controls should be evidence-based, consistently implemented across grocery stores, and coupled with administrative practices and policies to promote worker wellbeing.

Strongly Coordinating Ligands To Form Weakly Coordinating Yet Functional Organometallic Anions.

Weakly coordinating anions (WCAs) are generally tailored to act as spectators with little or no function. Here we describe the implementation of strongly coordinating dianionic carboranyl N-heterocyclic carbenes (NHCs) to create organometallic -ate complexes of Au(I) that serve both as WCAs and functional catalysts. These organometallic WCAs can be utilized to form both heterobimetallic (Au(I)-/Ag(I)+; Au(I)-/Ir(I)+) and organometallic/main group ion pairs (Au(I)-/(CPh3+ or SiEt3+). Because parent unfunctionalized dianionic carboranyl NHC complex 3 is unstable in most solvents when paired with CPh3+, novel synthesis methodology was devised to create polyhalogenated carboranyl NHCs, which show superior stability toward electrophilic substitution and cyclometalation chemistry. Additionally, the WCAs containing polyhalogenated carboranyl NHCs are among the most active catalysts reported for the hydroamination of alkynes. This investigation has also produced the first examples of a low-coordinate Au(III) center with two cis accessible coordination sites and the first true dianionic carbene. These studies pave the way for the design of functional ion pairs that have the potential to participate in tandem or cooperative small-molecule activation and catalysis.

LFP clustering in cortex reveals a taxonomy of Up states and near-millisecond, ordered phase-locking in cortical neurons.

During slow-wave sleep and anesthesia, mammalian cortex exhibits a synchronized state during which neurons shift from a largely nonfiring to a firing state, known as an Up-state transition. Up-state transitions may constitute the default activity pattern of the entire cortex (Neske GT. Front Neural Circuits 9: 88, 2016) and could be critical to understanding cortical function, yet the genesis of such transitions and their interaction with single neurons is not well understood. It was recently shown that neurons firing at rates >2 Hz fire spikes in a stereotyped order during Up-state transitions (Luczak A, McNaughton BL, Harris KD. Nat Rev Neurosci 16: 745-755, 2015), yet it is still unknown if Up states are homogeneous and whether spiking order is present in neurons with rates <2 Hz (the majority). Using extracellular recordings from anesthetized cats and mice and from naturally sleeping rats, we show for the first time that Up-state transitions can be classified into several types based on the shape of the local field potential (LFP) during each transition. Individual LFP events could be localized in time to within 1-4 ms, more than an order of magnitude less than in previous studies. The majority of recorded neurons synchronized their firing to within ±5-15 ms relative to each Up-state transition. Simultaneous electrophysiology and wide-field imaging in mouse confirmed that LFP event clusters are cortex-wide phenomena. Our findings show that Up states are of different types and point to the potential importance of temporal order and millisecond-scale signaling by cortical neurons.NEW & NOTEWORTHY During cortical Up-state transitions in sleep and anesthesia, neurons undergo brief periods of increased firing in an order similar to that occurring in awake states. We show that these transitions can be classified into distinct types based on the shape of the local field potential. Transition times can be defined to <5 ms. Most neurons synchronize their firing to within ±5-15 ms of the transitions and fire in a consistent order.

Status of inflammation in relation to health related quality of life in hepatocellular carcinoma patients.

PURPOSE: Both Inflammation and health-related quality of life (HRQoL) are independent prognosticators in HCC patients. We hypothesized that inflammation can cause impairment in HRQoL and investigated the correlation between inflammatory status and HRQoL in HCC patients. METHODS: Clinical, laboratory and HRQoL (using EORTC QLQ-C30, QLQ-HCC18, C30 and HCC18 index-scores) data were prospectively collected from HCC patients at diagnosis. Correlation analyses were performed between HRQoL and inflammation-based markers including C-reactive protein (CRP), CRP/albumin ratio (CRP/alb), Glasgow Prognostic Score (GPS), Inflammation-Based Index (IBI) and Prognostic Index (PI). RESULTS: Among 445 HCC patients, higher inflammatory states were significantly correlated with worse HRQoL. For CRP and CRP/alb ratio, the HRQoL factors with higher correlations included C30 and HCC18 index-scores, certain QLQ-C30 domains and items ('physical functioning', 'role functioning', 'fatigue', 'pain', 'appetite loss') and QLQ-HCC18 items ('fatigue', 'body image', 'nutrition' and 'abdominal swelling'), where the Pearson's correlation coefficients were up to 0.416. Multivariate analyses indicated that worse HRQoL factors were significantly correlated with worse scores in GPS, IBI and PI. CONCLUSION: In HCC patients, inflammatory status correlates with HRQoL at presentation. In particular, relatively stronger correlations with CRP-based markers have been observed in HRQoL scales that assess constitutional symptoms (QLQ-C30 'physical functioning', 'role functioning', 'fatigue', 'appetite loss' and QLQ-HCC18 'fatigue' and 'nutrition') and tumor burden (QLQ-C30 'pain' and QLQ-HCC18 'abdominal swelling' and 'body image'). Future studies are warranted to evaluate whether intervention that reduces inflammation could improve HRQoL in HCC patients.

Low-dose radiation: interagency collaboration on planning research could improve information on health effects.

In September 2017, the US Government Accountability Office (GAO) issued a report on the scientific basis for federal protections against the harmful effects of ionizing radiation-particularly low-dose radiation, or below about 100 millisieverts (10 rem). The report provided examples of how federal agencies developed and applied radiation protection requirements and guidance for workers and the public and examined the extent to which federal agencies funded epidemiological and radiobiological research on the health effects of low-dose radiation and their efforts at coordination of this research. The report found that agencies collaborated on individual projects on radiation's health effects but did not have a mechanism to coordinate research priorities. Consequently, GAO recommended that the Department of Energy (DOE) lead the development of a mechanism for interagency collaboration related to research on the health effects of low-dose radiation.

Mesoscale Mapping of Mouse Cortex Reveals Frequency-Dependent Cycling between Distinct Macroscale Functional Modules.

Connectivity mapping based on resting-state activity in mice has revealed functional motifs of correlated activity. However, the rules by which motifs organize into larger functional modules that lead to hemisphere wide spatial-temporal activity sequences is not clear. We explore cortical activity parcellation in head-fixed, quiet awake GCaMP6 mice from both sexes by using mesoscopic calcium imaging. Spectral decomposition of spontaneous cortical activity revealed the presence of two dominant frequency modes (<1 and ∼3 Hz), each of them associated with a unique spatial signature of cortical macro-parcellation not predicted by classical cytoarchitectonic definitions of cortical areas. Based on assessment of 0.1-1 Hz activity, we define two macro-organizing principles: the first being a rotating polymodal-association pinwheel structure around which activity flows sequentially from visual to barrel then to hindlimb somatosensory; the second principle is correlated activity symmetry planes that exist on many levels within a single domain such as intrahemispheric reflections of sensory and motor cortices. In contrast, higher frequency activity >1 Hz yielded two larger clusters of coactivated areas with an enlarged default mode network-like posterior region. We suggest that the apparent constrained structure for intra-areal cortical activity flow could be exploited in future efforts to normalize activity in diseases of the nervous system.SIGNIFICANCE STATEMENT Increasingly, functional connectivity mapping of spontaneous activity is being used to reveal the organization of the brain. However, because the brain operates across multiple space and time domains a more detailed understanding of this organization is necessary. We used in vivo wide-field calcium imaging of the indicator GCaMP6 in head-fixed, awake mice to characterize the organization of spontaneous cortical activity at different spatiotemporal scales. Correlation analysis defines the presence of two to three superclusters of activity that span traditionally defined functional territories and were frequency dependent. This work helps define the rules for how different cortical areas interact in time and space. We provide a framework necessary for future studies that explore functional reorganization of brain circuits in disease models.

Neocortical Rebound Depolarization Enhances Visual Perception.

Animals are constantly exposed to the time-varying visual world. Because visual perception is modulated by immediately prior visual experience, visual cortical neurons may register recent visual history into a specific form of offline activity and link it to later visual input. To examine how preceding visual inputs interact with upcoming information at the single neuron level, we designed a simple stimulation protocol in which a brief, orientated flashing stimulus was subsequently coupled to visual stimuli with identical or different features. Using in vivo whole-cell patch-clamp recording and functional two-photon calcium imaging from the primary visual cortex (V1) of awake mice, we discovered that a flash of sinusoidal grating per se induces an early, transient activation as well as a long-delayed reactivation in V1 neurons. This late response, which started hundreds of milliseconds after the flash and persisted for approximately 2 s, was also observed in human V1 electroencephalogram. When another drifting grating stimulus arrived during the late response, the V1 neurons exhibited a sublinear, but apparently increased response, especially to the same grating orientation. In behavioral tests of mice and humans, the flashing stimulation enhanced the detection power of the identically orientated visual stimulation only when the second stimulation was presented during the time window of the late response. Therefore, V1 late responses likely provide a neural basis for admixing temporally separated stimuli and extracting identical features in time-varying visual environments.

Cortical functional hyperconnectivity in a mouse model of depression and selective network effects of ketamine.

See Huang and Liston (doi:10.1093/awx166) for a scientific commentary on this article.Human depression is associated with glutamatergic dysfunction and alterations in resting state network activity. However, the indirect nature of human in vivo glutamate and activity assessments obscures mechanistic details. Using the chronic social defeat mouse model of depression, we determine how mesoscale glutamatergic networks are altered after chronic stress, and in response to the rapid acting antidepressant, ketamine. Transgenic mice (Ai85) expressing iGluSnFR (a recombinant protein sensor) permitted real-time in vivo selective characterization of extracellular glutamate and longitudinal imaging of mesoscale cortical glutamatergic functional circuits. Mice underwent chronic social defeat or a control condition, while spontaneous cortical activity was longitudinally sampled. After chronic social defeat, we observed network-wide glutamate functional hyperconnectivity in defeated animals, which was confirmed with voltage sensitive dye imaging in an independent cohort. Subanaesthetic ketamine has unique effects in defeated animals. Acutely, subanaesthetic ketamine induces large global cortical glutamate transients in defeated animals, and an elevated subanaesthetic dose resulted in sustained global increase in cortical glutamate. Local cortical inhibition of glutamate transporters in naïve mice given ketamine produced a similar extracellular glutamate phenotype, with both glutamate transients and a dose-dependent accumulation of glutamate. Twenty-four hours after ketamine, normalization of depressive-like behaviour in defeated animals was accompanied by reduced glutamate functional connectivity strength. Altered glutamate functional connectivity in this animal model confirms the central role of glutamate dynamics as well as network-wide changes after chronic stress and in response to ketamine.

Resolution of High-Frequency Mesoscale Intracortical Maps Using the Genetically Encoded Glutamate Sensor iGluSnFR.

Wide-field-of-view mesoscopic cortical imaging with genetically encoded sensors enables decoding of regional activity and connectivity in anesthetized and behaving mice; however, the kinetics of most genetically encoded sensors can be suboptimal for in vivo characterization of frequency bands higher than 1-3 Hz. Furthermore, existing sensors, in particular those that measure calcium (genetically encoded calcium indicators; GECIs), largely monitor suprathreshold activity. Using a genetically encoded sensor of extracellular glutamate and in vivo mesoscopic imaging, we demonstrate rapid kinetics of virally transduced or transgenically expressed glutamate-sensing fluorescent reporter iGluSnFR. In both awake and anesthetized mice, we imaged an 8 × 8 mm field of view through an intact transparent skull preparation. iGluSnFR revealed cortical representation of sensory stimuli with rapid kinetics that were also reflected in correlation maps of spontaneous cortical activities at frequencies up to the alpha band (8-12 Hz). iGluSnFR resolved temporal features of sensory processing such as an intracortical reverberation during the processing of visual stimuli. The kinetics of iGluSnFR for reporting regional cortical signals were more rapid than those for Emx-GCaMP3 and GCaMP6s and comparable to the temporal responses seen with RH1692 voltage sensitive dye (VSD), with similar signal amplitude. Regional cortical connectivity detected by iGluSnFR in spontaneous brain activity identified functional circuits consistent with maps generated from GCaMP3 mice, GCaMP6s mice, or VSD sensors. Viral and transgenic iGluSnFR tools have potential utility in normal physiology, as well as neurologic and psychiatric pathologies in which abnormalities in glutamatergic signaling are implicated. SIGNIFICANCE STATEMENT: We have characterized the usage of virally transduced or transgenically expressed extracellular glutamate sensor iGluSnFR to perform wide-field-of-view mesoscopic imaging of cortex in both anesthetized and awake mice. Probes for neurotransmitter concentration enable monitoring of brain activity and provide a more direct measure of regional functional activity that is less dependent on nonlinearities associated with voltage-gated ion channels. We demonstrate functional maps of extracellular glutamate concentration and that this sensor has rapid kinetics that enable reporting high-frequency signaling. This imaging strategy has utility in normal physiology and pathologies in which altered glutamatergic signaling is observed. Moreover, we provide comparisons between iGluSnFR and genetically encoded calcium indicators and voltage-sensitive dyes.

Systematic evaluation of circulating inflammatory markers for hepatocellular carcinoma.

BACKGROUNDS & AIMS: A number of circulating inflammatory factors are implicated in the pathogenesis and prognostication of hepatocellular carcinoma (HCC). We aim to evaluate the prognostication of multiple serum inflammatory factors simultaneously and develop an objective inflammatory score for HCC. METHODS: A prospective cohort of 555 patients with HCC with paired serum samples was accrued from 2009 to 2012. The blood levels of conventional inflammatory markers, namely C-reactive protein (CRP), albumin, neutrophils, lymphocytes and platelet, were determined, and 41 other exploratory markers were measured by a multiplex assay. The prognostication and interaction of markers were determined by univariate and multivarite analyses. RESULTS: The cohort was randomly divided into training cohort (n=139) and validation cohort (n=416). There were no differences in baseline characteristics between the two cohorts. In the training cohort, independent prognostic factors for overall survival included CRP (hazard ratio [HR] 1.107; P=.003), albumin (HR 0.953; P=.032) and interleukin-8 (HR=5.816; P<.001). We have modified the existing inflammation-based index (IBI) by adding serum interleukin-8 level. The modified IBI could stratify patients into four groups with distinct overall survival (P<.001). The results were also validated in the validation cohort. When compared with IBI and other conventional inflammatory markers, the modified IBI had better prognostic performance with higher c-index and homogeneity likelihood ratio chi-square. CONCLUSIONS: Among the conventional and exploratory circulating inflammatory markers, higher CRP, lower albumin and higher interleukin-8 were independent prognosticators. By combining these factors, a simple and accurate inflammatory index could be constructed.

The Underlying Neurobiological Mechanisms of Psychosis: Focus on Neurotransmission Dysregulation, Neuroinflammation, Oxidative Stress, and Mitochondrial Dysfunction.

Psychosis, defined as a set of symptoms that results in a distorted sense of reality, is observed in several psychiatric disorders in addition to schizophrenia. This paper reviews the literature relevant to the underlying neurobiology of psychosis. The dopamine hypothesis has been a major influence in the study of the neurochemistry of psychosis and in development of antipsychotic drugs. However, it became clear early on that other factors must be involved in the dysfunction involved in psychosis. In the current review, it is reported how several of these factors, namely dysregulation of neurotransmitters [dopamine, serotonin, glutamate, and γ-aminobutyric acid (GABA)], neuroinflammation, glia (microglia, astrocytes, and oligodendrocytes), the hypothalamic-pituitary-adrenal axis, the gut microbiome, oxidative stress, and mitochondrial dysfunction contribute to psychosis and interact with one another. Research on psychosis has increased knowledge of the complexity of psychotic disorders. Potential new pharmacotherapies, including combinations of drugs (with pre- and probiotics in some cases) affecting several of the factors mentioned above, have been suggested. Similarly, several putative biomarkers, particularly those related to the immune system, have been proposed. Future research on both pharmacotherapy and biomarkers will require better-designed studies conducted on an all stages of psychotic disorders and must consider confounders such as sex differences and comorbidity.

Alterations in brain network connectivity and subjective experience induced by psychedelics: a scoping review.

Intense interest surrounds current research on psychedelics, particularly regarding their potential in treating mental health disorders. Various studies suggest a link between the subjective effects produced by psychedelics and their therapeutic efficacy. Neuroimaging evidence indicates an association of changes in brain functional connectivity with the subjective effects of psychedelics. We conducted a review focusing on psychedelics and brain functional connectivity. The review focused on four psychedelic drugs: ayahuasca, psilocybin and LSD, and the entactogen MDMA. We conducted searches in databases of MEDLINE, Embase, APA PsycInfo and Scopus from inception to Jun 2023 by keywords related to functional connectivity and psychedelics. Using the PRISMA framework, we selected 24 articles from an initial pool of 492 for analysis. This scoping review and analysis investigated the effects of psychedelics on subjective experiences and brain functional connectivity in healthy individuals. The studies quantified subjective effects through psychometric scales, revealing significant experiences of altered consciousness, mood elevation, and mystical experiences induced by psychedelics. Neuroimaging results indicated alterations in the functional connectivity of psychedelics, with consistent findings across substances of decreased connectivity within the default mode network and increased sensory and thalamocortical connectivity. Correlations between these neurophysiological changes and subjective experiences were noted, suggesting a brain network basis of the psychedelics' neuropsychological impact. While the result of the review provides a potential neural mechanism of the subjective effects of psychedelics, direct clinical evidence is needed to advance their clinical outcomes. Our research serves as a foundation for further exploration of the therapeutic potential of psychedelics.

Observation of room temperature B-Cl activation of the HCB11Cl11(-) anion and isolation of a stable anionic carboranyl phosphazide.

The perchlorinated carba-closo-dodecaborate anion is typically inert toward B-Cl functionalization. We present here the observation of two competing reactions that occur with this anion at ambient temperature. When this molecule is treated with n-BuLi and subsequently reacted with tosyl azide, a cycloaddition occurs and results in chloride substitution at a B-Cl vertex. The competing and dominant pathway is a substitution reaction to form the azide N3CB11Cl11(-). This rare anionic carboranyl azide reacts with PPh3 in FC6H5 to afford a stable anionic phosphazide. When dissolved in tetrahydrofuran, the phosphazide is in equilibrium with free PPh3 and N3CB11Cl11(-). Both the triazole and phosphazide are characterized by single-crystal X-ray diffraction, NMR and IR spectroscopy, and high-resolution mass spectrometry.

A fishermen-developed intervention reduced musculoskeletal load associated with commercial Dungeness crab harvesting.

This study characterized physical risk factors associated with injuries during a Dungeness crab harvesting task and evaluated the efficacy of a fishermen-developed ergonomic control (banger bar) in mitigating physical risk factors, including biomechanical loads in the low back, shoulders, and upper extremities, and postural instability. In a repeated-measures laboratory study, 25 healthy male participants performed manual crab harvesting tasks in five conditions: without any banger bar (control) and with 4 bars of differing heights or designs. The results showed that the ergonomic control reduced trunk and shoulder angles, L5/S1, and shoulder moments; muscle activities in low back, shoulders, and upper extremities; perceived exertion ratings; and postural sway measures. Moreover, these measures were lowest when the bar height was at 60 cm, indicating that the banger bar can reduce the risk of musculoskeletal injuries and postural instability, and that bar height is an important factor affecting these injury risk measures.

Testing for EGFR Variants in Pleural and Pericardial Effusion Cell-Free DNA in Patients With Non-Small Cell Lung Cancer.

Importance: Molecular testing in non-small cell lung cancer (NSCLC) is commonly limited by inadequate tumor sample. Plasma cell-free DNA (cfDNA) genotyping as a complementary test is specific but only moderately sensitive. Genotyping of cfDNA in pleural and pericardial effusion (PE-cfDNA) can further optimize molecular diagnostic yield and reduce the need for repeated biopsies. Objective: To prospectively validate droplet digital polymerase chain reaction (ddPCR) for detection of sensitizing EGFR variants and acquired Thr790Met variant (T790M) from PE-cfDNA in patients with NSCLC. Design, Setting, and Participants: This prospective diagnostic validation study was conducted between September 6, 2016, and January 21, 2021 at 2 major Hong Kong cancer centers. Patients with advanced NSCLC with both wild-type and variant EGFR status and exudative PE who underwent thoracocentesis or pericardiocentesis were randomly enrolled. Patients were either EGFR-tyrosine kinase inhibitor (TKI) naive (cohort 1) or EGFR-TKI treated but osimertinib naive (cohort 2). Enrolled patients underwent pleural- or pericardial-fluid and blood sampling for ddPCR EGFR testing. EGFR status results with ddPCR testing of PE-cfDNA and blood were compared with EGFR status in matched tumor biopsy or PE cell block samples. Main Outcomes and Measures: Specificity, sensitivity, and concordance of PE-cfDNA for detection of sensitizing EGFR variants and acquired T790M variation. Results: Among 171 patients (54% female) enrolled, there were 104 in cohort 1 and 67 in cohort 2. In cohort 1, 37% (38/102) were EGFR-variant positive; PE-cfDNA showed 97% sensitivity (95% CI, 92%-100%), 97% specificity (95% CI, 93%-100%), and 97% concordance (ĸ = 0.94, P < .001) for the detection of sensitizing EGFR variants. It was more sensitive than plasma in detecting sensitizing EGFR variants (97% vs 74%, P < .001). In cohort 2, 38% (15 of 40) were positive for the EGFR T790M variant; PE-cfDNA showed 87% sensitivity (95% CI, 69%-100%), 60% specificity (95% CI, 41%-79%), and 70% concordance (ĸ = 0.42, P = .004) for acquired T790M. The EGFR T790M variant was detected in 51% of PE-cfDNA vs 25% of PE cell block samples. Conclusions and Relevance: In this diagnostic study, EGFR variants could be accurately detected from PE-cfDNA in patients with NSCLC. More EGFR T790M was detected in PE-cfDNA than in guideline-recommended PE cell block preparations. These results suggest that PE-cfDNA can complement plasma and tumor genotyping for detecting EGFR variants in patients with advanced NSCLC.

A small-molecule screen in C. elegans yields a new calcium channel antagonist.

Small-molecule inhibitors of protein function are powerful tools for biological analysis and can lead to the development of new drugs. However, a major bottleneck in generating useful small-molecule tools is target identification. Here we show that Caenorhabditis elegans can provide a platform for both the discovery of new bioactive compounds and target identification. We screened 14,100 small molecules for bioactivity in wild-type worms and identified 308 compounds that induce a variety of phenotypes. One compound that we named nemadipine-A induces marked defects in morphology and egg-laying. Nemadipine-A resembles a class of widely prescribed anti-hypertension drugs called the 1,4-dihydropyridines (DHPs) that antagonize the alpha1-subunit of L-type calcium channels. Through a genetic suppressor screen, we identified egl-19 as the sole candidate target of nemadipine-A, a conclusion that is supported by several additional lines of evidence. egl-19 encodes the only L-type calcium channel alpha1-subunit in the C. elegans genome. We show that nemadipine-A can also antagonize vertebrate L-type calcium channels, demonstrating that worms and vertebrates share the orthologous protein target. Conversely, FDA-approved DHPs fail to elicit robust phenotypes, making nemadipine-A a unique tool to screen for genetic interactions with this important class of drugs. Finally, we demonstrate the utility of nemadipine-A by using it to reveal redundancy among three calcium channels in the egg-laying circuit. Our study demonstrates that C. elegans enables rapid identification of new small-molecule tools and their targets.

Comparative analysis of six genome sequences of three novel picornaviruses, turdiviruses 1, 2 and 3, in dead wild birds, and proposal of two novel genera, Orthoturdivirus and Paraturdivirus, in the family Picornaviridae.

In this territory-wide molecular epidemiology study of picornaviruses, involving 6765 dead wild birds from 201 species in 50 families over a 12 month period, three novel picornaviruses, turdiviruses 1, 2 and 3 (TV1, TV2 and TV3), were identified from birds of different genera in the family Turdidae. In contrast to many other viruses in birds of the family Turdidae or viruses of the family Picornaviridae, TV1, TV2 and TV3 were found exclusively in the autumn and winter months. Two genomes each of TV1, TV2 and TV3 were sequenced. Regions P1, P2 and P3 of the three turdiviruses possessed, respectively, <40, <40 and <50 % amino acid identities with those of other picornaviruses. Moreover, P1, P2 and P3 of TV1 also possessed, respectively, <40, <40 and <50 % amino acid identities with those of TV2 and TV3. Phylogenetic analysis revealed that TV1, TV2 and TV3 were distantly related to members of the genus Kobuvirus. Among the three turdiviruses, TV2 and TV3 were always clustered together, with high bootstrap supports of 1000. The genomic features of TV2 and TV3 were also distinct from TV1, including lower G+C contents, shorter leader protein and a preference for codon sequence NNT rather than NNC for amino acids that can use either NNT or NNC as codons (P<0.001 by χ(2)-test). Based on our results we propose two novel genera, Orthoturdivirus for TV1, and Paraturdivirus for TV2 and TV3, in the family Picornaviridae. The type of internal ribosomal entry site for TV1, TV2 and TV3 remains to be determined.

Stress impacts sensory variability through cortical sensory activity motifs.

Medically unexplained symptoms in depression are common. These individual-specific complaints are often considered an 'idiom of distress', yet animal studies suggest that cortical sensory representations are flexible and influenced by spontaneous cortical activity. We hypothesized that stress would reveal activity dynamics in somatosensory cortex resulting in greater sensory-evoked response variability. Using millisecond resolution in vivo voltage sensitive dye (VSD) imaging in mouse neocortex, we characterized spontaneous regional depolarizations within limb and barrel regions of somatosensory cortex, or spontaneous sensory motifs, and their influence on sensory variability. Stress revealed an idiosyncratic increase in spontaneous sensory motifs that is normalized by selective serotonin reuptake inhibitor treatment. Spontaneous motif frequency is associated with increased variability in sensory-evoked responses, and we optogenetically demonstrate that regional depolarization in somatosensory cortex increases sensory-evoked variability for seconds. This reveals a putative circuit level target for changes in sensory processing and for unexplained physical complaints in stress-related psychopathology.