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Search indexed bioRxiv preprints in genomics, neuroscience, cell biology and bioinformatics. Read source abstracts and check manuscript versions; preprints are not peer reviewed.

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At least 361 records · Page 20Linked to original sources

Engineering a Multilayer Microfluidic Airway-On-A-Chip with Tunable GelMA Hydrogel for Physiologically Relevant Aerosol Exposure Studies

Climate change-driven increases in forest fires pose a major global health risk due to exposure to smoke containing hazardous gases and fine particulates, emphasizing the need for physiologically relevant in vitro airway models for studying smoke-induced responses. Microfluidic lung-on-a-chip technologies provide a strong foundation for in vitro airway modeling and ongoing developments are expanding their ability to incorporate multicellular organization, extracellular matrix complexity, and physiologically relevant exposure methods. This work presents the optimization and integration of a photopolymerizable gelatin methacrylate (GelMA)-based hydrogel into a microfluidic airway-on-a-chip that models the human small conducting airways and supports controlled aerosol exposure to wood smoke. The GelMA hydrogel was optimized to support fibroblast encapsulation, endothelial, and epithelial adhesion and robust mechanical stability. The device combines the hydrogel with a compartmentalized microchannel layout, and sacrificial molding to create a 3D organotypic airway culture featuring a multilayer architecture, 3D stromal matrix, and a perfusable vasculature-like lumen. Coupling the platform with a custom aerosol exposure system enables precise, biomimetic exposure to whole wood smoke. Proof-of-concept studies using transforming growth factor beta1 (TGF-{beta}1) and whole wood smoke elicited expected inflammatory and fibrotic responses, validating the platforms physiological relevance for inhalation studies and investigating smoke-induced airway remodeling and inflammation.

bioengineering↗

Trade-offs between surviving and thriving: A careful balance of physiological limitations and reproductive effort under thermal stress

Balancing survival and reproduction presents a fundamental evolutionary challenge, especially in extreme and unpredictable environments. Thermoregulatory behavior in particular imposes a costly trade-off, as time spent maintaining optimal body temperature precludes other essential activities and forces individuals to balance competing selective pressures. By combining field observations, behavioral assays, and gene expression profiling of red-sided garter snakes (Thamnophis sirtalis parietalis), we describe how an ectothermic vertebrate navigates this trade-off in an extreme thermal environment through a combination of finely tuned mechanisms of behavioral and physiological plasticity. Snakes demonstrated remarkably consistent critical thermal limits, with minimal individual variation, suggesting a hard physiological constraint. Importantly, behavioral temperature thresholds for voluntary withdrawal and courtship cessation occurred within 5 {degrees}C of lethal, demonstrating that males routinely operate perilously close to lethal body temperatures in order to maximize reproductive opportunities. Context-dependent plasticity allows males to prioritize mating opportunities over thermoregulation until survival becomes an immediate threat. Molecular analyses of multiple tissues revealed rapid, tissue-specific transcriptomic responses activated within one hour of thermal challenge, well before critical limits were reached. Heat shock proteins were universally upregulated across all tissues under both heat and cold stress, suggesting anticipatory protective mechanisms that support risky behavioral decisions rather than simply responding to thermal damage. Tissue-specific patterns of gene expression reflected functional priorities with liver showing extensive metabolic flexibility, heart maintaining conservative stability, while brain and testis appeared to balance critical functions with stress responses. These findings demonstrate precise thermal reaction norms that integrate behavioral and molecular plasticity to maximize reproductive effort without compromising survival. The narrow margin between behavioral thresholds and lethal limits suggests this system may already be approaching evolutionary constraints. While phenotypic plasticity currently buffers populations against extreme thermal variability, it may paradoxically limit long-term adaptive potential as increasing environmental temperatures further narrow safety margins. Precise thermal decision-making, rapid physiological responses, and context-dependent behavioral switching suggest an evolved solution to the problem of balancing competing selective pressures.

zoology↗

Bioengineered 3D hPSC-cholangiocyte ducts with physiological signals for biliary disease modelling

The progression of intrahepatic biliary diseases remains poorly understood, underscoring the urgent need to develop physiologically relevant human intrahepatic cholangiocyte disease models. Current approaches lack the complexity and throughput to capture how diverse biliary microenvironmental signals shape cholangiocyte behavior. To address this gap, we created a fully epithelialized and perfusible 3D bile duct from human pluripotent stem cell-derived cholangiocytes characterized by robust primary ciliation, apical-basal polarity and CFTR-mediated chloride conductance. Our results revealed physiologically relevant fluid flow and biliary stroma cells as essential components for sustaining cholangiocyte epithelial barrier integrity and ciliation. From here, we interrogated a broad spectrum of biliary signals to study bile acid toxicity and cytokine-driven injury, offering an unprecedented view into intrahepatic cholangiocyte stress responses and potential pathological mechanisms. By integrating physiologically relevant signals, fidelity and functional resolution, this platform provides the foundation needed to precisely decode pathogenic drivers and accelerate therapeutic development for devastating cholangiopathies. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=198 HEIGHT=200 SRC="FIGDIR/small/686855v1_ufig1.gif" ALT="Figure 1"> View larger version (59K): org.highwire.dtl.DTLVardef@11c1255org.highwire.dtl.DTLVardef@1f78d11org.highwire.dtl.DTLVardef@14e9398org.highwire.dtl.DTLVardef@1bc08d5_HPS_FORMAT_FIGEXP M_FIG C_FIG

cell biology↗

Immediate effects of psychosocial stress on attention depend on subjective experience and not directly on stress-related physiological changes

Acute psychosocial stress is associated with physiological, subjective and cognitive changes. In particular, attention, which is considered one of the main processes driving cognition, has been related to different stress outcomes, such as anxiety, cortisol levels and autonomic responses, individually. Nonetheless, their specific contributions to and association with attention is still not fully understood. To study this association, 42 male participants were asked to perform an attentional task just before and immediately after being exposed to either an experimental treatment designed to induce psychosocial stress using the Trier Social Stress Test (TSST) or a matching stress-free control condition. The salivary cortisol concentration, heart rate, and self-reported anxiety were measured to assess the physiological response to stress and the subjective experience during the protocol. As expected, psychosocial stress induced increases in heart rate, salivary cortisol levels and anxiety. The behavioral analysis revealed that members of the control group performed better on the attentional task after the protocol, while members of the TSST group showed no changes. Moreover, after dividing the stress group into sub-groups of participants with high and low anxiety, we observed that participants in the high-anxiety group not only failed to perform better but also performed worse. Finally, after testing several single-level mediation models, we found that anxiety is sufficient to explain the changes in attention and that it mediates the effects between heart rate and cortisol levels on attention. Our results suggest that the immediate effects of acute psychosocial stress on attention are highly dependent on the participants subjective experience, which, in turn, is affected and can mediate stress-related physiological changes.

neuroscience↗

Seasonal physiological responses to heat in an alpine range-restricted bird: the Cape Rockjumper

Hot, dry summer conditions impose physiological stress on endotherms, yet we have a poor understanding of how endotherms seasonally adjust their costs of thermoregulation under hot conditions. We determined whether seasonal phenotypic plasticity in evaporative cooling capacity at high temperatures explained how the range-restricted Cape Rockjumper (Chaetops frenatus; hereafter \"Rockjumper\"), copes with hot and dry summer temperatures of the temperate mountain peaks of southwest South Africa. We measured evaporative water loss (EWL), resting metabolic rate (RMR), and body temperature (Tb) at high air temperatures (30 to 42{degrees}C) of individuals from a wild population of Rockjumpers during winter and summer (n = 11 winter, 4 females, 7 males; n = 10 summer, 6 females, 4 males). We found Rockjumper evaporative cooling in summer imposes higher EWL (i.e. greater water costs) compared to winter, although an accompanying lack of change in RMR resulted in increased summer cooling efficiency. These patterns are similar to those observed in species that inhabit regions where summer temperatures are routinely hot but not water stressed. Our findings show that avian seasonal physiological adjustments to heat can be diverse. Further seasonal studies on thermoregulation in the heat will greatly improve our knowledge of the functional value traits such as evaporative cooling efficiency and heat tolerance hold and how they contribute to the physiological stress organisms experience in heterogenous environments.

zoology↗

When metabolism meets physiology: Harvey and Harvetta

Precision medicine is an emerging paradigm that requires realistic, mechanistic models capturing the complexity of the human body. We present two comprehensive molecular to physiological-level, gender-specific whole-body metabolism (WBM) reconstructions, named Harvey, in recognition of William Harvey, and Harvetta. These validated, knowledge-based WBM reconstructions capture the metabolism of 20 organs, six sex organs, six blood cells, the gastrointestinal lumen, systemic blood circulation, and the blood-brain barrier. They represent 99% of the human body weight, when excluding the weight of the skeleton. Harvey and Harvetta can be parameterized based on physiological, dietary, and omics data. They correctly predict inter-organ metabolic cycles, basal metabolic rates, and energy use. We demonstrate the integration of microbiome data thereby allowing the assessment of individual-specific, organ-level modulation of host metabolism by the gut microbiota. The WBM reconstructions and the individual organ reconstructions are available under http://vmh.life. Harvey and Harvetta represent a pivotal step towards virtual physiological humans.

systems biology↗

Physiology and lipid accumulation capacity of different Yarrowia lipolytica and Rhodosporidium toruloides strains on glycerol

ObjectiveTo compare physiological and process parameters, as well as lipid accumulation capacity, of six strains of Yarrowia lipolytica and two strains of Rhodosporidium toruloides in media containing glycerol as the main carbon and energy source.\n\nResultsThe strains Y. lipolytica IMUFRJ 50678, Po1g, W29 and CCT 5443 displayed very similar physiological parameters, with {micro}max, [Formula] and [Formula] average values of 0.31 h-1, 0.53 (g DW/g S), 0.60 (g S/g DW.h) and 0.46 (g DW/g S), respectively. The two strains of R. toruloides presented physiological and process parameters with no significant difference, with average values of 0.084 h-1, 0.53 (g DW/g S), 0.17 (g S/g DW.h) and 0.44 (g DW/g S). Among all Y. lipolytica strains, Y. lipolytica CCT 5443 strain presented the highest YLip/S, 0.054 (g Lip/g S), and PLipof 0.040 (g Lip/l.h). Among all investigated strains (Y. lipolytica and R. toruloides), the yeast R. toruloides CCT 7815 displayed the highest lipid accumulation capacity, with YLip/Sequal to 0.11 (g Lip/g S) and PLipequal to 0.10 (g Lip/l.h).\n\nConclusionAmong all strains investigated in our study, the yeast strain R. toruloides CCT 7815 presents the most promising characteristics for industrial single cell oil production.\n\nList of abbreviations

microbiology↗

Physiological Characteristics of Viable-but-nonculturable Vibrio parahaemolyticus upon Prolonged Exposure to the Refrigerator Temperature

Although it has been reported that viable-but-nonculturable (VBNC) cells of Vibrio parahaemolyticus can be developed by a prolonged duration of cold-starvation there are restricted cellular characteristics available on understanding the exact mechanisms governing the entry of pathogens into the VBNC state. Therefore, this research was aimed at determining the cellular profile of VBNC cells of V. parahaemolyticus upon exposure to the refrigerator temperature. Strains of V. parahaemolyticus were incubated in artificial sea water (ASW) microcosms (pH 6) added with different amounts of NaCl at 4{degrees}C until these pathogens entered into such a dormant state. At a regular time-interval, both culturability and viability of these bacteria were enumerated, and then cellular profiling were carried out in terms of cellular membrane permeability, enzymatic activity, hydrophobicity, fatty acid composition, and morphological changes after cells of V. parahaemolyticus became the VBNC state. Three strains of V. parahaemolyticus used in this study showed that VBNC cells retained the strong virulent properties to Vero and CACO-2 cell lines, re-gained the cytotoxicity even after resuscitation, became permeabilized in terms of the outer membrane, showed lower levels of enzymatic (catalase and glutathione-S-transferase) activities, exerted the increasing hydrophobicity, and then exhibited increasing amounts of saturated fatty acids.\n\nIMPORTANCETo the current best knowledge, there are restricted information available on understanding the physiological characterization of viable-but-nonculturable cells. Most previous studies are still making a degree of efforts in discovering the causative effector causing microorganisms to be induced into the VBNC state. Herein, the present study showed that pathogenic V. parahaemolyticus can enter into the VBNC state when challenged by a certain environmental stress where higher amounts of NaCl combined with acidic pHs was artificially controlled. Importantly, it was indicated that VBNC V. parahaemolyticus maintained peculiarly different physiological characteristics. Furthermore, this study proposed a novel approach on the transient/stepwise conversion of the bacteria into the VBNC state. Specific alternative tools for measuring and controlling the incidence of VBNC pathogens on food are not established until now. In this aspect, results obtained from this study will used to provide an effective insight in determining physiological properties of viable-but-nonculturable V. parahaemolyticus.

microbiology↗

Protein nitration is a physiological regulator of cardiac lactate dehydrogenase active site loop mobility and activity

Protein tyrosine nitration is a hallmark of oxidative stress related disease states, commonly detected as anti-nitrotyrosine immunoreactivity. The precise reactive oxygen sources, mechanisms of nitration as well as the modified target proteins and functional consequences, however, remain often unclear. Here we explore protein tyrosine nitration under basal conditions and find surprisingly physiologically nitrated proteins. Upon purifying a prominent physiologically nitrotyrosine immunopositive in hearts from mouse, rat and pig, we identify it as lactate dehydrogenase (LDH). Mechanistically, LDHs degree of basal nitration depended on two canonical sources, NO synthase (NOS) and myeloperoxidase (MPO), respectively. When validating the nitrated amino acid by MALDI-TOF mass spectrometry, we, surprisingly, located LDH nitration not to a tyrosine but the C-terminal tryptophan, Trp324. Molecular dynamics simulations suggested that Trp324 nitration restricts the interaction of the active site loop with the C-terminal -helix essential for activity. This prediction was confirmed by enzyme kinetics revealing an apparent lower Vmax of nitrated LDH, although yet unidentified concurrent oxidative modifications may contribute. Protein nitration is, thus, not a by definition disease marker but reflects also physiological signaling by eNOS/NO, MPO/nitrite and possibly other pathways. The commonly used assay of anti-nitrotyrosine immunoreactivity is apparently cross-reactive to nitrotryptophan requiring a reevaluation of the protein nitration literature. In the case of LDH, nitration of Trp324 is aggravated under cardiac metabolic stress conditions and functionally limits maximal enzyme activity. Trp324-nitrated LDH may serve both as a previously not recognized disease biomarker and possibly mechanistic lead to understand the metabolic changes under these conditions.

biochemistry↗

Cognitive self-regulation influences pain-related physiology

Cognitive self-regulation can shape pain experience, but its effects on autonomic responses to painful events are unclear. In this study, participants (N = 41) deployed a cognitive strategy based on reappraisal and imagination to regulate pain up or down on different trials while skin conductance responses (SCR) and electrocardiogram (ECG) activity were recorded. Using a machine learning approach, we first developed stimulus-locked SCR and ECG physiological markers predictive of pain ratings. The physiological markers demonstrated high sensitivity and moderate specificity in predicting pain across two datasets, including an independent test dataset (N = 84). When we tested the markers on the cognitive self-regulation data, we found that cognitive self-regulation had significant impacts on both pain ratings and pain-related physiology in accordance with regulatory goals. These findings suggest that self-regulation can impact autonomic nervous system responses to painful stimuli and provide pain-related autonomic profiles for future studies.

neuroscience↗

Noninvasive Vagus Nerve Stimulation Alters Neural Response and Physiological Autonomic Tone to Noxious Thermal Challenge

The mechanisms by which noninvasive vagal nerve stimulation (nVNS) affect central and peripheral neural circuits that subserve pain and autonomic physiology are not clear, and thus remain an area of intense investigation. Effects of nVNS vs sham stimulation on subject responses to five noxious thermal stimuli (applied to left lower extremity), were measured in 30 healthy subjects (n=15 sham and n=15 nVNS), with fMRI and physiological galvanic skin response (GSR). With repeated noxious thermal stimuli a group x time analysis showed a significantly (p < .001) decreased response with nVNS in bilateral primary and secondary somatosensory cortices (SI and SII), left dorsoposterior insular cortex, bilateral paracentral lobule, bilateral medial dorsal thalamus, right anterior cingulate cortex, and right orbitofrontal cortex. A group x time x GSR analysis showed a significantly decreased response in nVNS group (p < .0005) in bilaterally in SI, lower and mid medullary brainstem, and inferior occipital cortex. Finally, nVNS treatment showed decreased activity in pronociceptive brainstem nuclei (e.g. the reticular nucleus and rostral ventromedial medulla) and key autonomic integration nuclei (e.g. the rostroventrolateral medulla, nucleus ambiguous, and dorsal motor nucleus of the vagus nerve). In aggregate, noninvasive vagal nerve stimulation reduced the physiological response to noxious thermal stimuli and impacted neural circuits important for pain processing and autonomic output.

neuroscience↗

A physiology-based model of bile acid metabolism shows altered tissue concentrations after drug administration and in specific genotype subgroups

Drug-induced liver injuries (DILI) are an important issue in drug development and patient safety and often lead to termination of drug-development programs or late withdrawals of drugs. Since DILI events are hard to diagnose in preclinical settings, a need for alternative prediction methods such as computational modeling emerges. Impairment of bile acid (BA) metabolism, known as cholestasis, is a frequent form of DILI. Being rather a systemic then a single organ related disease, whole-body physiology-based modeling is a predestined approach for cholestasis modeling. The objectives of the presented study were 1) the development of a physiology-based model for human bile acid metabolism, 2) model validation and characterization for a virtual population, and 3) prediction and quantification of the effects of genetic predispositions and drug interaction on bile acid metabolism. The developed physiology-based bile acid (PBBA) model is based on the standard PBPK model of PKSim(R) and describes the bile acid circulation in a healthy reference individual. Active processes such as the hepatic synthesis, gallbladder emptying upon meal intake, transition through the gastrointestinal tract, reabsorption into the liver, distribution within the body, and excretion are included. The kinetics of active processes for the surrogate BA glycochenodeoxycholic acid were fitted to time-concentration profiles of blood BA levels reported in literature. The robustness of our PBBA model is underlined by the comparison of simulated plasma BA concentrations in a virtual population of 1,000 healthy individuals with reported data. In addition to plasma concentrations, the PBBA model allows simulations of BA exposure in relevant tissues like the liver and can therefore enhance the mechanistic understanding of cholestasis. This feature was used to analyse the reported increased risk of cholestatic DILI in Benign Recurrent Intrahepatic Cholestasis type 2 (BRIC2) patients. Simulations of the PBBA model suggest a higher susceptibility of BRIC2 patients towards cholestatic DILI due to BA accumulation in hepatocytes. Apart from these intrinsic effects, drug-interactions and their effect on the systemic bile acid metabolism were simulated by combining the PBBA model with a drug PBPK model of cyclosporine A (CsA). The results of which confirmed the reported higher risk of developing DILI as a consequence of CsA intake. Altogether, the presented model enhances our mechanistic understanding of cholestasis, allows the identification of drug-interactions leading to altered BA levels in blood and organs, and could be used to prevent clinical cases of cholestasis and enhance patient safety.

systems biology↗

Single cell transcriptomic analysis of the adult mouse pituitary reveals a novel multi-hormone cell cluster and physiologic demand-induced lineage plasticity

The anterior pituitary gland drives a set of highly conserved physiologic processes in mammalian species. These hormonally-controlled processes are central to somatic growth, pubertal transformation, fertility, lactation, and metabolism. Current models, largely built upon candidate gene based immuno-histochemical and mRNA analyses, suggest that each of the seven hormones synthesized by the pituitary is produced by a specific and exclusive cell lineage. However, emerging evidence suggests more complex models of hormone specificity and cell plasticity. Here we have applied massively parallel single-cell RNA sequencing (scRNA-seq), in conjunction with a set of orthogonal mRNA and protein imaging studies, to systematically map the cellular composition of adult male and female mouse pituitaries at single-cell resolution and in the setting of major physiologic demands. These analyses reveal sex-specific cellular diversity associated with normal pituitary homeostasis, and identify an array of cells with complex complements of hormone-enrichment as well as a series of non-hormone producing interstitial and supporting cell lineages. These scRNA-seq studies identify a major cell population that is characterized by a unique multi-hormone gene expression profile. The detection of dynamic shifts in cellular representations and transcriptome profiles in response to two well-defined physiologic stresses suggests corresponding roles of a number of these clusters in cellular plasticity within the adult pituitary. These studies point to an unanticipated complexity and plasticity in pituitary cellular composition that expands upon current models and concepts of pituitary gene expression and hormone production.

genomics↗

Imbalanced Nucleocytoskeletal Connections Create Common Polarity Defects in Progeria and Physiological Aging

Studies of the accelerated aging disorder Hutchinson-Gilford progeria syndrome (HGPS) can potentially reveal cellular defects associated with physiological aging. HGPS results from expression and abnormal nuclear envelope association of a farnesylated, truncated variant of prelamin A called progerin. We surveyed the diffusional mobilities of nuclear membrane proteins to identify proximal effects of progerin expression. The mobilities of three proteins were reduced in fibroblasts from children with HGPS compared to normal fibroblasts: SUN2, nesprin-2G, and emerin. These proteins function together in nuclear movement and centrosome orientation in fibroblasts polarizing for migration. Both processes were impaired in fibroblasts from children with HGPS and in NIH3T3 fibroblasts expressing progerin, but were restored by inhibiting protein farnesylation. Progerin affected both the coupling of the nucleus to actin cables and the oriented flow of the cables necessary for nuclear movement and centrosome orientation. Progerin overexpression increased levels of SUN1, which couples the nucleus to microtubules through nesprin-2G and dynein, and microtubule association with the nucleus. Reducing microtubule-nuclear connections through SUN1 depletion or dynein inhibition rescued the polarity defects. Nuclear movement and centrosome orientation were also defective in fibroblasts from normal individuals over 60 years old and both defects were rescued by reducing the increased level of SUN1 in these cells or inhibiting dynein. Our results identify imbalanced nuclear engagement of the cytoskeleton (microtubules, high; actin filaments, low) as the basis for intrinsic cell polarity defects in HGPS and physiological aging and suggest that rebalancing the connections can ameliorate the defects.\n\nSignificanceThe rare, premature aging syndrome HGPS arises from expression of a pathological prelamin A variant, termed progerin. Studies of progerin may identify treatments for HGPS and reveal novel cellular and molecular characteristics of normal aging. Here, we show that progerin selectively affects mobilities of three nuclear membrane proteins, SUN2, nesprin-2G and emerin that position the nucleus and establish cell polarity essential for migration. We find that both processes are defective in fibroblasts from children with HGPS and aged (> 60 years) individuals. The mechanism underlying these defects is excessive interaction of the nucleus with microtubules. Our work identifies nuclear-based defects in cell polarization as intrinsic factors in premature and physiological aging and suggests means for correcting them.

cell biology↗

Terminal Reproductive Investment, Physiological Trade-offs and Pleiotropic Effects: Their effects produce complex immune/reproductive interactions in the cricket Gryllus texensis

O_LIShould females increase or decrease reproduction when attacked by pathogens? Two hypotheses provide opposite predictions. Terminal reproductive investment theory predicts an increase in reproduction, but hypothesized physiological trade-offs between reproduction and immune function might be expected to produce a decrease. There is evidence for both hypotheses. What determines the choice between the two responses remains unclear. We examine the effect of age on the reproductive response to immune challenge in long-wing females of the Texas field cricket, Gryllus texensis, when fed an ecologically valid (limited) diet.\nC_LIO_LIThe limited diet reduced reproductive output. However, immune challenge had no effect on their reproductive output either in young or middle-aged crickets, which is contrary to either prediction.\nC_LIO_LIFlight muscle maintenance correlated negatively with reproductive output, suggesting a physiological trade-off between flight muscle maintenance and reproduction. Within the long-wing variant there was considerable variability in flight muscle maintenance. This variability may mask physiological trade-offs between immunity and reproduction.\nC_LIO_LIMiddle-aged crickets had higher total phenoloxidase (PO) activity in their hemolymph, compared to young females, which is contrary to the terminal investment theory. Given that PO is involved in both immunity and reproduction, the increased PO may reflect simultaneous investment in both functions.\nC_LIO_LIWe identified four proPO transcripts in a published RNA-seq dataset (transcriptome). Three of the proPO genes were expressed either in the fat body or the ovaries (supporting the hypothesis that PO is bifunctional); however, the two organs expressed different subsets. The possible bifunctionality of PO suggests that it may not be an appropriate immune measure for immune/reproductive trade-offs in some species.\nC_LIO_LIIncreasing age may not cue terminal reproductive investment prior to senescence.\nC_LI

ecology↗

An event-based paradigm for analyzing fluorescent astrocyte activity uncovers novel single-cell and population-level physiology

Recent work examining astrocytic physiology centers on fluorescence imaging approaches, due to development of sensitive fluorescent indicators and observation of spatiotemporally complex calcium and glutamate activity. However, the field remains hindered in fully characterizing these dynamics, both within single cells and at the population-level, because of the insufficiency of current region-of-interest-based approaches to describe activity that is often spatially unfixed, size-varying, and propagative. Here, we present a paradigm-shifting analytical framework that releases astrocyte biologists from ROI-based tools. Astrocyte Quantitative Analysis (AQuA) software enables users to take an event-based approach to accurately capture and quantify the irregular activity observed in astrocyte imaging datasets. We apply AQuA to a range of ex vivo and in vivo imaging data, and uncover previously undescribed physiological phenomena in each. Since AQuA is data-driven and based on machine learning principles, it can be applied across model organisms, fluorescent indicators, experimental modes, and imaging resolutions and speeds, enabling researchers to elucidate fundamental astrocyte physiology.

neuroscience↗

Automated high-throughput heart rate measurement in medaka and zebrafish embryos under physiological conditions

RationaleAccurate and efficient quantification of heartbeats in small fish models is an important readout to study cardiovascular biology, disease states and pharmacology at large scale. However, dependence on anesthesia, laborious sample orientation or requirement for fluorescent reporters have hampered the establishment of high-throughput heartbeat analysis. ObjectiveTo overcome these limitations, we aimed to develop a high-throughput assay with automated heart rate scoring in medaka (Oryzias latipes) and zebrafish (Danio rerio) embryos under physiological conditions designed for genetic screens and drug discovery and validation. Methods and ResultsWe established an efficient screening assay employing automated label-free heart rate determination of randomly oriented, non-anesthetized specimen in microtiter plates. Automatically acquired bright-field data feeds into an easy-to-use HeartBeat software, a MATLAB algorithm with graphical user interface developed for automated quantification of heart rate and rhythm. Sensitivity of the assay and algorithm was demonstrated by profiling heart rates during entire embryonic development. Our analysis pipeline revealed acute temperature changes triggering rapid adaption of heart rates, which has implications for standardization of experimental layout. The approach is scalable and allows scoring of multiple embryos per well resulting in a throughput of >500 embryos per 96-well plate. In a proof of principle screen for compound testing, our assay captured concentration-dependent effects of nifedipine and terfenadine over time. ConclusionA novel workflow and HeartBeat software provide efficient means for reliable and direct quantification of heart rate and rhythm of small fish in a physiological environment. Importantly, confounding factors such as anesthetics or laborious mounting are eliminated. We provide detailed profiles of embryonic heart rate dynamics in medaka and zebrafish as reference for future assay development. Ease of sample handling, automated imaging, physiological conditions and software-assisted analysis now facilitate various large-scale applications ranging from phenotypic screening, interrogation of gene functions to cardiovascular drug development pipelines.

developmental biology↗

Exploring Deep Physiological Models for Nociceptive Pain Recognition

Standard feature engineering involves manually designing and assessing measurable descriptors based on some expert knowledge in the domain of application, followed by the selection of the best performing set of designed features in order to optimize an inference model. Several studies have shown that this whole manual process can be efficiently replaced by deep learning approaches which are characterized by the integration of feature engineering, feature selection and inference model optimization into a single learning process. Such techniques have proven to be very successful in the domain of image processing and have been able to attain state-of-the-art performances while significantly outperforming traditional approaches based on hand-crafted features. In the following work, we explore deep learning approaches for the analysis of physiological signals. More precisely, deep learning architectures are designed for the assessment of measurable physiological channels in order to perform an accurate classification of different levels of artificially induced nociceptive pain. Most of the previous works related to pain intensity classification based on physiological signals rely on a carefully designed set of hand-crafted features in order to achieve a relatively good classification performance. Therefore, the current work aims at building competitive pain intensity classification models without the need of domain specific expert knowledge for the generation of relevant features. The assessment of the designed deep learning architectures is based on the BioVid Heat Pain Database (Part A) and experimental validation demonstrates that the proposed uni-modal architecture for the electrodermal activity (EDA) and the deep fusion approaches significantly outperform previous classification methods reported in the literature, with respective average performances of 85.03% and 83.76% for the binary classification experiment consisting of the discrimination between the baseline level and the pain tolerance level (T0vs.T4) in a Leave-One-Subject-Out (LOSO) cross-validation evaluation setting.

bioinformatics↗