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At least 1,351 records · Page 75Linked to original sources

bioRxiv2017

A subset of sweet-sensing neurons identified by IR56d are necessary and sufficient for fatty acid taste

Fat represents a calorically potent food source that yields approximately twice the amount of energy as carbohydrates or proteins per unit of mass. The highly palatable taste of FAs promotes food consumption, activates reward circuitry, and is thought to contribute to hedonic feeding underlying many metabolism-related disorders. Despite a role in the etiology of metabolic diseases, little is known about how dietary fats are detected by the gustatory system to promote feeding. Previously, we showed that a broad population of sugar-sensitive taste neurons expressing Gustatory Receptor 64f (Gr64f), is required for reflexive feeding responses to both FAs and sugars. Here, we report a genetic silencing screen to identify specific populations of taste neurons that mediate fatty acid taste. We find neurons identified by expression of Ionotropic Receptor 56d (IR56d) are necessary and sufficient for reflexive feeding response to FAs. Functional imaging reveals that IR56d-expressing neurons are responsive to short- and medium-chain FAs. Silencing IR56d neurons selectively abolishes fatty acid taste, while their activation is sufficient to drive feeding responses. Analysis of co-expression with Gr64f identifies two subpopulations of IR56d-expressing neurons. While physiological imaging reveals that both populations are responsive to FAs, IR56d/Gr64f neurons are activated by medium-chain FAs and are sufficient for reflexive feeding responses to FA. Moreover, flies can discriminate between sugar and FAs in an aversive taste memory assay, indicating that FA taste is a unique modality in Drosophila. Taken together, these findings localize fatty acid taste within the Drosophila gustatory center and provide an opportunity to investigate discrimination between different categories of appetitive tastants.\n\nAuthors SummaryFat represents a calorically potent food source that yields approximately twice the amount of energy as carbohydrates or proteins per unit of mass. Dietary lipids are comprised of both triacylglycerides and FAs, and growing evidence suggests that it is the free FAs that are detected by the gustatory system. The highly palatable taste of FAs promotes food consumption, activates reward centers in mammals, and is thought to contribute to hedonic feeding that underlies many metabolism-related disorders. Despite a role in the etiology of metabolic diseases, little is known about how dietary fats are detected by the gustatory system to promote feeding. We have identified a subset of sugar-sensing neurons in the fly that also respond to medium-chain FAs and are necessary and sufficient for behavioral responses to FAs. Further, we find that despite being sensed by shared neuron populations, flies can differentiate between the taste of sugar and FAs, fortifying the notion that FAs and sugar represent distinct taste modalities in flies.

neuroscience↗

bioRxiv2017

Regressions Fit for Purpose: Models of Locust Phase State Must Not Conflate Morphology With Behaviour

Locusts are defined by their capacity to transform between two very distinct integrated phenotypes or phases in response to changes in population density: a solitarious phase, which occurs when densities are low, and a gregarious phase, which arises as a consequence of crowding and can form very large and economically damaging swarms. The two phases differ fundamentally in their behaviour, physiology and morphology. A large body of work on the mechanistic basis of behavioural phase transitions has relied on multivariate logistic regression (LR) models to estimate the probability of behavioural gregariousness from multiple behavioural variables. Martin-Blazquez and Bakkali (2017, Entomologia Experimentalis et Applicata 163, 9-25) have recently proposed standardised LR models for estimating an overall gregariousness level from a combination of behavioural and, unusually, morphometric variables. Here I develop a detailed argument to demonstrate that the premise of such an overall gregariousness level is fundamentally flawed. Since locust phase transformations intrinsically entail a decoupling of behaviour and morphology, phase state cannot meaningfully be conflated onto a single axis. LR models that do so are therefore of very limited value for any analysis of phase transitions. I furthermore show why behavioural predictor variables should not be adjusted by measures of body size that themselves differ between phases. I discuss the models fitted by Martin-Blazquez and Bakkali (2017) to highlight potential pitfalls in statistical methodology that must be avoided when applying LR to the analysis of behavioural phase state. Finally, I reject the idea that standardised models provide a valid shortcut to estimating phase state across different developmental stages, strains or species. The points addressed here are pertinent to any research on transitions between complex phenotypes and behavioural syndromes.

animal behavior and cognition↗

bioRxiv2017

Biophysical modeling of neural plasticity induced by transcranial magnetic stimulation

Transcranial magnetic stimulation (TMS) is a widely used noninvasive brain stimulation method capable of inducing plastic reorganisation of cortical circuits in humans. Changes in neural activity following TMS are often attributed to synaptic plasticity (e.g long-term potentiation and depression; LTP/LTD). However, the precise way in which synaptic processes such as LTP/LTD modulate the activity of large populations of neurons, as stimulated en masse by TMS, are unclear. The recent development of biophysically-informed models, which capture the physiological properties of TMS-induced plasticity using mathematics, provide an excellent framework for reconciling synaptic and macroscopic plasticity. In this article, we overview the TMS paradigms used to induce plasticity, and their limitations. We then describe the development of biophysically-based numerical models of the mechanisms underlying LTP/LTD on population-level neuronal activity, and the application of these models to TMS plasticity paradigms, including theta burst and paired associative stimulation. Finally, we outline how modeling can complement experiment to improve mechanistic understandings and optimize outcomes of TMS-induced plasticity.\n\nAbbreviations

neuroscience↗

bioRxiv2017

Network inference reveals novel connections in pathways regulating growth and defense in the yeast salt response

Cells respond to stressful conditions by coordinating a complex, multi-faceted response that spans many levels of physiology. Much of the response is coordinated by changes in protein phosphorylation. Although the regulators of transcriptome changes during stress are well characterized in Saccharomyces cerevisiae, the upstream regulatory network controlling protein phosphorylation is less well dissected. Here, we developed a computational approach to infer the signaling network that regulates phosphorylation changes in response to salt stress. The method uses integer linear programming (ILP) to integrate stress-responsive phospho-proteome responses in wild-type and mutant strains, predicted phosphorylation motifs on groups of coregulated peptides, and published protein interaction data. A key advance is that by grouping peptides into submodules before inference, the method can overcome missing protein interactions in published datasets to predict novel, stress-dependent protein interactions and phosphorylation events. The network we inferred predicted new regulatory connections between stress-activated and growth-regulating pathways and suggested mechanisms coordinating metabolism, cell-cycle progression, and growth during stress. We confirmed several network predictions with co-immunoprecipitations coupled with mass-spectrometry protein identification and mutant phospho-proteomic analysis. Results show that the cAMP-phosphodiesterase Pde2 physically interacts with many stress-regulated transcription factors targeted by PKA, and that reduced phosphorylation of those factors during stress requires the Rck2 kinase that we show physically interacts with Pde2. Together, our work shows how a high-quality computational network model can facilitate discovery of new pathway interactions during osmotic stress.

systems biology↗

bioRxiv2017

Breaking the Deadlock of Molecular Chaperones

Protein folding in the cell requires ATP-driven chaperone machines. It is poorly understood, however, how these machines fold proteins. Here we propose that the conserved Hsp70 and Hsp90 chaperones support formation of the folding nucleus by providing a gradient of decreasing hydrophobicity. Early on the folding pathway Hsp70 uses its highly hydrophobic binding pocket to recover a stalled, unproductive folding intermediate. The aggressive nature of Hsp70 action, however, blocks productive folding by grabbing hydrophobic, core-forming segments. This precludes on-pathway nucleation at high, physiological Hsp70 levels. Transfer to the less hydrophobic Hsp90 enables the intermediate to resume forming its folding nucleus. Subsequently, the protein enters a spontaneous folding trajectory towards its native state, independent of the ATPase activities of both Hsp70 and Hsp90. Our findings provide a general mechanistic concept for chaperoned protein folding.

biochemistry↗

bioRxiv2017

Hydrogen peroxide formation by Nox4 limits malignant transformation

Reactive oxygen species (ROS) can cause cellular damage and are thought to promote cancer-development. Nevertheless, under physiological conditions, all cells constantly produce ROS, either as chemical by-products or for signaling purpose. During differentiation cells induce the NADPH oxidase Nox4, which constitutively produces low amounts of H2O2. We infer that this constitutive H2O2 is unlikely to be carcinogenic and may rather maintain basal activity of cellular surveillance systems.\n\nUtilizing two different murine tumor models we demonstrate that Nox4 prevents malignant transformation and facilitated the recognition of DNA-damage. Upon DNA-damage repair is initiated as consequence of phosphorylation of H2AX ({gamma}H2AX). Repair only occurs if nuclear activity of the {gamma}H2AX-dephosphorylating phosphatase PP2A is kept sufficiently low, a task fulfilled by Nox4: Nox4 continuously oxidizes AKT, and once oxidized AKT captures PP2A in the cytosol. Absence of Nox4 facilitates nuclear PP2A translocation and dephosphorylation of {gamma}H2AX. Simultaneously the proportion of active, phosphorylated AKT is increased. Thus, DNA-damage is not recognized and the increase in AKT activity promotes proliferation. The combination of both events resulted in genomic instability and tumor initiation.\n\nWith the identification of the first cancer-protective source of reactive oxygen species, Nox4, the paradigm of reactive-oxygen species-induced initiation of malignancies should be revised.\n\nSignificanceThe stereotype of ROS produced by NADPH oxidases as cause of malignant diseases persists generalized since decades. We demonstrate that the NADPH oxidase Nox4, as constitutive source of ROS, prevents malignant transformation and that its pharmacological inhibition as currently aspired by several companies will potentially increase the risk of malignant cell transformation and eventually tumor formation.\n\nPrecisBy oxidizing AKT and keeping PP2A in the cytosol, the NADPH oxidase Nox4 allows proper DNA damage repair and averts cancer development.

cancer biology↗

bioRxiv2017

Expression of lamina proteins Lamin Dm0 and Kugelkern suppresses stem cell proliferation

The nuclear lamina is involved in numerous cellular functions, such as gene expression, nuclear organization, nuclear stability, and cell proliferation. The mechanism underlying the involvement of lamina is often not clear, especially in physiological contexts. Here we investigate how the farnesylated lamina proteins Lamin Dm0 and Kugelkern are linked to proliferation control of intestinal stem cells (ISCs) in adult Drosophila flies by loss-of-function and gain-of-function experiments. We found that ISCs mutant for Lamin Dm0 or Kugelkern proliferate, whereas overexpression of Lamin Dm0 or Kugelkern strongly suppressed proliferation. The anti-proliferative activity is, at least in part, due to suppression of Jak/Stat but not Delta/Notch signalling. Lamin Dm0 expression suppresses Jak/Stat signalling by normalization of about 50% of the Stat target genes in ISCs.\n\nAuthor summaryThe nuclear lamina is a protein meshwork that lies beneath the inner side of the nuclear membrane and interacts with nuclear pores, chromatin and the cytoskeleton. Changes in proteins of the nuclear lamina cause a wide range of diseases which are often not well understood. It is hypothesized that impairment of stem cell function, as a result of lamina changes, might play a key role in some of those diseases. Here we use the well characterized Drosophila midgut as a system to investigate the role of lamina proteins Lamin Dm0 and Kugelkern on stem cell proliferation.

developmental biology↗

bioRxiv2017

Oxytocin Signaling in the Medial Amygdala is required for Sex Discrimination of Social Cues

The neural control of social behaviors in rodents requires the encoding of pheromonal cues by the vomeronasal system. Here we show that the typical preference of male mice for females is eliminated in mutants lacking oxytocin, a neuropeptide modulating social behaviors in many species. Ablation of the oxytocin receptor in aromatase expressing neurons of the medial amygdala (MeA) fully recapitulates the elimination of female preference in males. Further, single unit recording in the MeA uncovered significant changes in the sensory representation of conspecific cues in the absence of oxytocin signaling. Finally, acute manipulation of oxytocin signaling in adults is sufficient to alter social interaction preferences in males as well as responses of MeA neurons to chemosensory cues. These results uncover the critical role of oxytocin signaling in a molecularly defined neuronal population in order to modulate the behavioral and physiological responses of male mice to females on a moment-to-moment basis.

neuroscience↗

bioRxiv2017

The Anti-Epileptic Ketogenic Diet Alters Hippocampal Transporter Levels and Reduces Adiposity in Aged Rats

Nutritional ketosis is induced by high fat/low carbohydrate dietary regimens, which produce high levels of circulating ketone bodies, shifting metabolism away from glucose utilization. While ketogenic diets (KD) were initially introduced to suppress seizures, they are garnering attention for their potential to treat a myriad of neurodegenerative and metabolic disorders that are associated with advanced age. The feasibility and physiological impact of implementing a long-term KD in old animals, however, has not been systematically examined. In this study, young and aged rats consumed a calorically- and nutritionally-matched KD or control diet for 12 weeks. All KD-fed rats maintained higher levels of {beta}-hydroxybutyrate (BHB) and lower levels of glucose relative to controls. However, it took the aged rats longer to reach asymptotic levels of BHB compared to young animals. Moreover, KD-fed rats had significantly less visceral white and brown adipose tissue than controls without a loss of lean mass. Interestingly, the KD lead to significant alterations in protein levels of hippocampal transporters for monocarboxylates, glucose and vesicular glutamate and GABA. Most notably, the age-related decline in vesicular glutamate transporter expression was reversed by the KD. These data demonstrate the feasibility and potential benefits of KDs for treating age-associated neural dysfunction.

neuroscience↗

bioRxiv2017

Brain state and polarity dependent modulation of brain networks by transcranial direct current stimulation

Transcranial direct current stimulation (TDCS) has been widely used to improve cognitive function. However, current deficiencies in mechanistic understanding hinders wider applicability. To clarify its physiological effects, we acquired fMRI whilst simultaneously acquiring TDCS to the right inferior frontal gyrus (rIFG) of healthy human participants, a region involved in coordinating activity within brain networks. TDCS caused widespread modulation of network activity depending on brain state ( rest or choice reaction time task) and polarity (anodal or cathodal). During task, TDCS increased salience network activation and default mode network deactivation, but had the opposite effect during rest. Furthermore, there was an interaction between brain state and TDCS polarity, with cathodal effects more pronounced during task performance and anodal effects more pronounced during rest. Overall, we show that rIFG TDCS produces brain state and polarity dependent effects within large-scale cognitive networks, in a manner that goes beyond predictions from the current literature.

neuroscience↗

bioRxiv2017

Transcriptional Architecture of Synaptic Communication Delineates Cortical GABAergic Neuron Identity

Understanding the organizational logic of neural circuits requires deciphering the biological basis of neuron type diversity and identity, but there is no consensus on defining a neuron type. We analyzed single cell transcriptomes of anatomically and physiologically characterized cortical ground truth populations and conducted a computational genomic screen for transcription profiles that distinguish them. We discovered that cardinal GABAergic neuron types are delineated by a transcriptional architecture that encodes their synaptic communication patterns. This architecture comprises 6 categories of ~40 gene families including cell adhesion molecules, transmitter-modulator receptors, ion channels, signaling proteins, neuropeptides and vesicular release components, and transcription factors. Combinatorial expression of select members across families shapes a multi-layered molecular scaffold along cell membrane that may customize synaptic connectivity patterns and input-output signaling properties. This molecular genetic framework of neuronal identity integrates cell phenotypes along multiple axes and provides a foundation for discovering and classifying neuron types.

neuroscience↗

bioRxiv2017

The 10,000 Immunomes Project: A resource for human immunology

New immunological assays now enable rich measurements of human immune function, but difficulty attaining enough measurements across sufficiently large and diverse cohorts has hindered describing normal human immune physiology on a large scale. Here we present the 10,000 Immunomes Project (10KIP), a diverse human immunology reference derived from over 44,000 individuals across 242 studies from ImmPort, a publicly available resource of raw immunology study data and protocols. We carefully curated datasets, aggregating subjects from healthy/control arms and harmonizing data across studies. We demonstrate 10KIPs utility by describing variations in serum cytokines and leukocytes by age, race, and sex; defining a baseline cell-cytokine network; and using 10KIP as a common control to describe immunologic changes in pregnancy. Subject-level data is available for interactive visualization and download at http://10kImmunomes.org/. We believe 10KIP can serve as a common control cohort and will accelerate hypothesis generation by clinical and basic immunologists across diverse populations.\n\nOne Sentence SummaryAn open online resource of human immunology data from more than 10,000 normal subjects including interactive data visualization and download enables a new look at immune system differences across age and sex, rapid hypothesis generation, and creation of custom control cohorts.

immunology↗

bioRxiv2017

Synthesis, secretion, and perception of abscisic acid regulates stress responses in Chlorella sorokiniana

Abscisic acid (ABA) is a phytohormone that has been extensively characterized in higher plants for its roles in seed and bud dormancy, leaf abscission, and stress responses. Genomic studies have identified orthologs for ABA-related genes throughout the Viridiplantae, including in unicellular algae; however, the role of ABA in algal physiology has not been characterized, and the existence of such a role has been a matter of dispute. In this study, we demonstrate that ABA is involved in regulating algal stress responses. Chlorella sorokiniana strain UTEX 1230 contains genes orthologous to those of higher plants which are essential for ABA biosynthesis, sensing, and degradation. RNAseq-based transcriptomic studies reveal that treatment with ABA induces dramatic changes in gene expression profiles, including the induction of a subset of genes involved in DNA replication and repair, a phenomenon which has been demonstrated in higher plants. Pretreatment of C. sorokiniana cultures with ABA exerts a protective effect on cell viability in response to ultraviolet radiation. Additionally, C. sorokiniana produces and secretes biologically relevant amounts of both ABA and the oxylipin 12-oxo-phytodienoic acid (OPDA) into the growth medium in response to abiotic stressors. Taken together, these phenomena suggest that ABA signaling evolved as an intercellular stress response signaling molecule in eukaryotic microalgae prior to the evolution of multicellularity and colonization of land.

plant biology↗

bioRxiv2017

Induction of differentiation and metabolic reprogramming in human hepatoma cells by adult human serum

Tissue culture medium routinely contains fetal bovine serum (FBS). Here we show that culturing human hepatoma cells in their native, adult serum (human serum, HS) results in the restoration of key morphological and metabolic features of normal liver cells. When moved to HS, these cells show differential transcription of 22-32% of the genes, stop proliferating, and assume a hepatocyte-like morphology. Metabolic analysis shows that the Warburg-like metabolic profile, typical for FBS-cultured cells, is replaced by a diverse metabolic profile consistent with in vivo hepatocytes. We demonstrate the formation of large lipid and glycogen stores, increased glycogenesis, increased {beta}-oxidation, increased ketogenesis, and decreased glycolysis. Finally, organ-specific functions are restored, including xenobiotics degradation and secretion of bile, very low density lipoprotein, and albumin. Thus, organ-specific functions are not necessarily lost in cell cultures, but might be merely suppressed in FBS. Together, we showed that cells that are representative of normal physiology can be produced from cancer cells simply by replacing FBS by HS in culture media. The effect of serum is often overseen in cell culture and we provide a detailed study in the changes that occur, provide insight in some of the serum components that may play a role in the establishment of the different phenotypes, and discuss how these finding might be beneficial to a variety of research fields.

cell biology↗

bioRxiv2017

Mitochondrial sirtuins sir-2.2 and sir-2.3 regulate lifespan in C. elegans

Mitochondrial sirtuins regulate biochemical pathways and are emerging drug targets for metabolic and age-related diseases such as cancer, diabetes, and neurodegeneration. Yet, their functions remain unclear. Here, we uncover a novel physiological role for the C. elegans mitochondrial sirtuins, sir-2.2 and sir-2.3, in lifespan regulation. Using a genetic approach, we demonstrate that sir-2.2 and sir-2.3 mutants live 28-30% longer than controls when fed the normal lab diet of E. coli OP50. Interestingly, this effect is diet specific and is not observed when animals are fed the strain HT115, which is typically used for RNAi experiments. While decreased consumption of food is a known mechanism for lifespan extension, this does not account for the increased lifespan in the mitochondrial sirtuin mutants. sir-2.2 and sir-2.3 mutants display altered expression of genes involved in oxidative stress response, including increased expression of the mitochondrial superoxide dismutase sod-3 and decreased levels of catalases ctl-1 and ctl-2. Like their extended lifespan phenotype, these alterations in oxidative stress gene expression are diet dependent. The mitochondrial sirtuin mutants are more resistant to the lifespan extending effects of low levels of superoxide, suggesting that their increased lifespan involves a hormetic response. Our data suggest that sir-2.2 and sir-2.3 are not completely redundant in function and may possess overlapping yet distinct mechanisms for regulating oxidative stress response and lifespan.

genetics↗

bioRxiv2017

A Methodology for Evaluating the Performance of Alerting and Detection Algorithms Running on Continuous Patient Data

ObjectivesClinicians in the intensive care unit (ICU) are presented with a large number of physiological data consisting of periodic and frequently sampled measurements, such as heart rate and blood pressure, as well as aperiodic measurements, such as noninvasive blood pressure and laboratory studies. Because this data can be overwhelming, there is considerable interest in designing algorithms that help integrate and interpret this data and assist ICU clinicians in detecting or predicting in advance patients who may be deteriorating. In order to decide whether to deploy such algorithms in a clinical trial, it is important to evaluate these algorithms using retrospective data. However, the fact that these algorithms will be running continuously, i.e., repeatedly sampling incoming patient data, presents some novel challenges for algorithm evaluation. Commonly used measures of performance such as sensitivity and positive predictive value (PPV) are easily applied to static \"snapshots\" of patient data, but can be very misleading when applied to indicators or alerting algorithms that are running on continuous data. Our objective is to create a method for evaluating algorithm performance on retrospective data with the algorithm running continuously throughout the patients stay as it would in a real ICU.\n\nMethodsWe introduce our evaluation methodology in the context of evaluating an algorithm, a Hemodynamic Instability Indicator (HII), for assisting bedside ICU clinicians with the early detection of hemodynamic instability before the onset of acute hypotension. Each patients ICU stay is divided into segments that are labelled as hemodynamically stable or unstable based on clinician interventions typically aimed at treating hemodynamic instability. These segments can be of varying length with varying degrees of exposure to potential alerts, whether true positive or false positive. Furthermore, to simulate how clinicians might interact with the alerting algorithm, we use a dynamic alert supervision mechanism which suppresses subsequent alerts unless the indicator has significantly deteriorated since the prior alert. Under these conditions determining what counts as a positive or negative instance, and calculations of sensitivity, specificity, and positive predictive value can be problematic. We introduce a methodology for consistently counting positive and negative instances. The methodology distinguishes between counts based on alerting events and counts based on sub-segments, and show how they can be applied in calculating measures of performance such as sensitivity, specificity, positive predictive value.\n\nResultsThe introduced methodology is applied to retrospective evaluation of two algorithms, HII and an alerting algorithm based on systolic blood pressure. We use a database, consisting of data from 41,707 patients from 25 US hospitals, to evaluate the algorithms. Both algorithms are evaluated running continuously throughout each patients stay as they would in a real ICU setting. We show how the introduced performance measures differ for different algorithms and for different assumptions.\n\nDiscussionThe standard measures of diagnostic tests in terms of true positives, false positives, etc. are based on certain assumptions which may not apply when used in the context of measuring the performance on an algorithm running continuously, and thus repeatedly sampling from the same patient. When such measures are being reported it is important that the underlying assumptions be made explicit; otherwise, the results can be very misleading.\n\nConclusionWe introduce a methodology for evaluating how an alerting algorithm or indicator will perform running continuously throughout every patients ICU stay, not just for a subset of patients for selected episodes.

bioinformatics↗

bioRxiv2017

Preferential cholinergic excitation of corticopontine neurons

Pyramidal neurons in layer 5 of the neocortex comprise two broad classes of projection neurons: corticofugal neurons, including corticopontine (CPn) neurons, and intratelencephalic neurons, including commissural/callosal (COM) neurons. These non-overlapping neuron subpopulations represent discrete cortical output channels contributing to perception, decision making, and behavior. CPn and COM neurons have distinct morphological and physiological characteristics, and divergent responses to modulatory transmitters such as serotonin and acetylcholine (ACh). To better understand how ACh regulates cortical output, in slices of mouse prefrontal cortex (PFC) we compared the responsivity of CPn and COM neurons to transient exposure to exogenous or endogenous ACh. In both neuron subtypes, exogenous ACh generated qualitatively similar biphasic responses in which brief hyperpolarization was followed by longer-lasting enhancement of excitability. However, cholinergic inhibition was more pronounced in COM neurons, while excitatory responses were larger and longer lasting in CPn neurons. Similarly, optically triggered release of endogenous ACh from cholinergic terminals preferentially and persistently (for ~40 s) enhanced the excitability of CPn neurons, but had little impact on COM neurons. Cholinergic excitation of CPn neurons involved at least three distinct ionic mechanisms: activation of a calcium-sensitive but calcium-permeable nonspecific cation conductance, suppression of Kv7 channels (the \"M-current\"), and activation of the calcium-dependent nonspecific cation conductance underlying afterdepolarizations. Our results demonstrate projection-specific selectivity in cholinergic signaling in the PFC, and suggest that transient release of ACh during behavior will preferentially promote corticofugal output.

neuroscience↗

bioRxiv2017

Streptolysin-induced endoplasmic reticulum stress promotes group A streptococcal in vivo biofilm formation and necrotizing fasciitis

Group A Streptococcus (GAS) is a human pathogen that causes infections ranging from mild to fulminant and life-threatening. Biofilms have been implicated in acute GAS soft-tissue infections such as necrotizing fasciitis (NF). However, most in vitro models used to study GAS biofilms have been designed to mimic chronic infections and insufficiently recapitulate in vivo conditions and the host-pathogen interactions that might influence biofilm formation. Here we establish and characterize an in vitro model of GAS biofilm development on mammalian cells that simulates microcolony formation observed in a murine model of human NF. We show that on mammalian cells, GAS forms dense aggregates that display hallmark biofilm characteristics including a three-dimensional architecture and enhanced tolerance to antibiotics. In contrast to abiotic-grown biofilms, host-associated biofilms require the expression of secreted GAS streptolysins O and S (SLO, SLS) resulting in the release of a host-associated biofilm promoting-factor(s). Supernatants from GAS-infected mammalian cells or from cells treated with endoplasmic reticulum (ER) stressors restore biofilm formation to an SLO and SLS null mutant that is otherwise attenuated in biofilm formation on cells, together suggesting a role for streptolysin-induced ER stress in this process. In an in vivo mouse model, the streptolysin-null mutant is attenuated in both microcolony formation and bacterial spread, but pre-treatment of softtissue with an ER-stressor restores the ability of the mutant to form wild type like microcolonies that disseminate throughout the soft tissue. Taken together, we have identified a new role of streptolysin-driven ER stress in GAS biofilm formation and NF disease progression.\n\nSignificance StatementAlthough it is well-accepted that bacterial biofilms are associated with many chronic infections, little is known about the mechanisms by which group A Streptococcus (GAS) biofilms contribute to acute soft tissue-invasive diseases like necrotizing fasciitis (NF). In this study, we establish a physiologically relevant in vitro model to study GAS biofilm formation on mammalian cells and validate our findings in a mouse model that mimics human NF. This study demonstrates a novel role of GAS streptolysin-mediated ER stress in the development and spread of GAS biofilms in acute softtissue infections. We also show that biofilm formation depends on the release of a host-associated factor that promotes microcolony formation and GAS dissemination in vivo.

microbiology↗