Biology subjects

Pereira, T.

Publications and source records attributed to Pereira, T..

At least 19 recordsLinked to original sources

bioRxiv2026

A community benchmark enables high-resolution, cross-lab quantification of mouse social behavior

Social behavior is a sensitive readout of brain state, but quantifying rodent social behavior at scale has relied on siloed, lab-specific tools. We assembled Days of our Mice (DooM), a benchmark of manually annotated video and pose tracking spanning 15 laboratories and 33 social and solitary behaviors, and hosted a community behavior classification challenge. We built on challenge solutions to develop the High-Dimensional Rodent Annotator (HiDRA), a behavior classification tool that generalizes to labs unseen during training and learns cross-lab movement features. Applied without retraining to experimental data, HiDRA resolves line-specific and state-dependent behavioral fingerprints with strong generalizability and reproducibility, establishing high-resolution, shareable social phenotyping as a scalable readout of brain and internal state. We release the DooM dataset and HiDRA classifiers as open resources.

neuroscience↗

bioRxiv2026

BOWIe: Wireless Headstage for Optical Stimulation in Closed-Loop Optogenetics Experiments in Freely-moving Rodents

Optogenetics is an invaluable technique in neuroscience to study brain circuits, but conventional optogenetic systems use tethered approaches to deliver light to the brain which can hinder natural behavior. Optoelectronic advances led to the development of wireless optical stimulators for optogenetics experiments, but most reported devices have low widespread use due to complex builds and reduced versatility, which has also limited their integration in closed-loop systems. Here, we developed BOWIe, a battery-powered, low-weight, wireless headstage for photostimulation in optogenetics experiments with rodents, especially designed for simple integration in multiple behavioral setups, including those requiring closed-loop operation. BOWIe is based on a Bluetooth-Low Energy system-on-a-chip that allows simultaneous control and communication capabilities with a reduced number of components, and can be paired with an Arduino-based transmitter for simple integration in any behavioral paradigm using analogue or digital triggers. Two modes of optical stimulation are possible, either with ferrule-coupled optical fibers or LED neural probes, which permit adapting the headstage according to material availability or experimental demands. A user-friendly app was also developed to simplify headstage pairing, definition of stimulation parameters, and protocol design. BOWIe was validated in two behavioral paradigms, including in a closed-loop experiment using computer vision, Bonsai RX, and Deeplabcut Live to track behavioral parameters online and remotely trigger photostimulation upon detected events. BOWIe's modular design and flexible integration with behavioral setups make it a practical tool for in vivo experiments with optogenetics.

bioengineering↗

bioRxiv2026

Regulation of PL fimbriae (plf) and role of PlfB as a negative regulator during avian pathogenic Escherichia coli infection in turkeys

Little is currently known about the regulation of PL fimbriae produced by some pathogenic E. coli strains. We identified PlfB, a transcriptional regulator of the PapB family, as a negative regulator of PL fimbriae (Plf). Deletion of plfB significantly increased plfA transcription and Plf production. PlfB bound specifically to the plfA promoter region. Using a PplfA-lux reporter system, the deletion of plfB caused sustained and increased expression across serial passages. Global regulatory genes such as lrp, argR, and hns were also implicated in regulation of plf expression. Loss of lrp or argR decreased plf expression, suggesting these regulators positively regulate PL fimbriae. By contrast, loss of hns increased levels of plf expression, supporting its role as a negative regulator. Interestingly, loss of plfB significantly increased bacterial colonization in the lung of 6-day-old turkey poults. Further, complementation of plfB restored plf fimbrial expression and colonization of turkey lungs to wild-type levels. This further supports an important role for PL fimbriae for colonization of turkey lungs, since loss of plfB which resulted in increased plf gene expression in turkey lungs also correlated with higher bacterial numbers in the lungs. Taken together, these results indicate that PlfB acts as a negative regulator to reduce levels of PL fimbrial expression, under some conditions, but that it is not required for plf expression. This contrasts with the role the PapB regulator, which has been shown to play an essential role in expression of P fimbriae in uropathogenic E. coli strains.

microbiology↗

bioRxiv2026

Wdr62 and Patronin cooperate to organize distinct acentrosomal microtubule networks during Drosophila oogenesis

Acentrosomal microtubule (MT) organization is essential in many differentiated cells, yet the mechanisms that generate and maintain these networks remain poorly understood. Here, we identify Wdr62 as a conserved regulator of acentrosomal MT organization during Drosophila oogenesis. Wdr62 localizes to cortical non-centrosomal microtubule-organizing centers (ncMTOCs) and is required for proper localization of the conserved components Shot and Patronin. Loss of Wdr62 disrupts MT organization and oocyte polarity and reduces MT growth events during mid-oogenesis. Genetic analyses further reveal that Wdr62 functionally cooperates with Patronin and the MT-severing enzyme Katanin 80 to organize the acentrosomal MT network. Our data support a model in which Wdr62 promotes stabilization of MT minus ends through Patronin while Katanin-mediated severing generates new MT seeds that amplify the MT network. Moreover, Wdr62 and Patronin function together during assembly of the acentrosomal meiotic spindle, and their loss compromises fertility. Given that WDR62 mutations are associated with human disease in cell types that rely on acentrosomal MT organization, such as neurons, these findings provide insight into how disruption of WDR62-dependent MT organization may contribute to disease beyond female infertility.

developmental biology↗

bioRxiv2024

Predicting Future Development of Stress-Induced Anhedonia From Cortical Dynamics and Facial Expression

The current state of mental health treatment for individuals diagnosed with major depressive disorder leaves billions of individuals with first-line therapies that are ineffective or burdened with undesirable side effects. One major obstacle is that distinct pathologies may currently be diagnosed as the same disease and prescribed the same treatments. The key to developing antidepressants with ubiquitous efficacy is to first identify a strategy to differentiate between heterogeneous conditions. Major depression is characterized by hallmark features such as anhedonia and a loss of motivation (1, 2), and it has been recognized that even among inbred mice raised under identical housing conditions, we observe heterogeneity in their susceptibility and resilience to stress (3). Anhedonia, a condition identified in multiple neuropsychiatric disorders, is described as the inability to experience pleasure and is linked to anomalous medial prefrontal cortex (mPFC) activity (4). The mPFC is responsible for higher order functions (5-8), such as valence encoding; however, it remains unknown how mPFC valence-specific neuronal population activity is affected during anhedonic conditions. To test this, we implemented the unpredictable chronic mild stress (CMS) protocol (9-11) in mice and examined hedonic behaviors following stress and ketamine treatment. We used unsupervised clustering to delineate individual variability in hedonic behavior in response to stress. We then performed in vivo 2-photon calcium imaging to longitudinally track mPFC valence-specific neuronal population dynamics during a Pavlovian discrimination task. Chronic mild stress mice exhibited a blunted effect in the ratio of mPFC neural population responses to rewards relative to punishments after stress that rebounds following ketamine treatment. Also, a linear classifier revealed that we can decode susceptibility to chronic mild stress based on mPFC valence-encoding properties prior to stress-exposure and behavioral expression of susceptibility. Lastly, we used a markerless pose tracking computer vision tool, SLEAP (31), to predict whether a mouse would become resilient or susceptible based on facial expressions during a Pavlovian discrimination task. These results indicate that mPFC valence encoding properties and behavior are predictive of anhedonic states. Altogether, these experiments point to the need for increased granularity in the measurement of both behavior and neural activity, as these factors can predict the predisposition to stress-induced anhedonia.

neuroscience↗

bioRxiv2024

Small form factor implantable neural probe with efficient flip chip micro LED for in vivo optogenetics

Optogenetics is a widely used tool to dissect neural circuits with optical stimulation, but it requires that light is delivered to photosensitive neurons inside the brain. Implantable neural probes with microscale LEDs (LEDs) are an emerging approach to delivering light to the brain with superior light output control. However, approaches to integrate LEDs in neural probes depend on complex fabrication processes. Here, we developed an implantable small form factor neural probe that integrates highly efficient commercial flip chip LEDs using only standard lithography processes in silicon and a custom automated LED mounting approach with custom 3D-printed tools on a pick-and-place machine. The probe has a cross-sectional area under 0.013 mm2 but can output up to 2.5 mW of optical power with an irradiance of 175 mW/mm2. Due to the high plug efficiency of the LED, the neural probe can perform stimulation protocols up to 20 Hz and 80% duty cycles without surpassing estimated hotspot temperature elevations above 1 {o}C. The neural probes were validated in vivo, with brain activity in the motor cortex of transgenic mice being reliably modulated by pulsed light emitted from the probe.

bioengineering↗

bioRxiv2024

Dachsous is a key player in epithelial wound closure by modulating cell shape changes and cytoskeleton dynamics

Epithelia are vital tissues in multicellular organisms, acting as barriers between external and internal environments. Simple epithelia, such as those in embryos and the adult gut, have the remarkable ability to repair wounds efficiently, making them ideal for studying epithelial repair mechanisms. In these tissues, wound closure involves the coordinated action of a contractile actomyosin cable at the wound edge and collective cell movements around the wound. However, the dynamics of cell-cell interactions during this process remain poorly understood. Here, we demonstrate that Dachsous (Ds), an atypical cadherin associated with Planar Cell Polarity, is crucial for efficient epithelial repair in the Drosophila embryonic epidermis. We show that the absence of Ds leads to delayed wound closure, impaired actomyosin cable formation, and altered cell shape changes. Additionally, we reveal that Occluding Junctions are necessary for the proper apical localization of Ds, suggesting an unanticipated interaction between these two molecular complexes. This study identifies Ds as a novel player in epithelial repair and highlights the need for further investigating the molecular mechanisms by which Ds modulates cell shape and tissue morphogenesis. Summary statementThis study shows that the atypical cadherin Dachsous is essential for epithelial wound closure, influencing cytoskeletal dynamics and cell morphology, with Occluding Junctions regulating its subcellular localization.

cell biology↗

bioRxiv2023

Social isolation recruits amygdala-cortical circuitry to escalate alcohol drinking

Social isolation profoundly alters motivation and increases vulnerability to alcohol misuse in humans, yet the underlying neural mechanisms remain unclear. Here we show that isolation escalates alcohol drinking in male mice but suppresses it in females. Whole-cell recordings revealed that neurons in the basolateral amygdala projecting to the medial prefrontal cortex (BLA-mPFC) track alcohol intake in both sexes. Isolation increased BLA-mPFC excitability in males but decreased it in females, mirroring their opposite behavioral adaptations. Given this divergence, we focused subsequent mechanistic studies on males to isolate neural pathway-level drivers of escalated alcohol intake. Cellular-resolution calcium imaging showed that activity in BLA-mPFC neurons encodes and predicts alcohol drinking, and optogenetic activation of this pathway increased alcohol intake. Simultaneous optogenetics and calcium imaging revealed that BLA-mPFC stimulation enhanced mPFC neuronal responses to alcohol, mimicking isolation-induced activity patterns, while photoinhibition reduced drinking in isolated mice. Together, these findings identify a BLA-mPFC pathway mechanism through which social isolation reconfigures prefrontal processing to promote alcohol intake.

neuroscience↗

bioRxiv2023

Keypoint-MoSeq: parsing behavior by linking point tracking to pose dynamics

Keypoint tracking algorithms have revolutionized the analysis of animal behavior, enabling investigators to flexibly quantify behavioral dynamics from conventional video recordings obtained in a wide variety of settings. However, it remains unclear how to parse continuous keypoint data into the modules out of which behavior is organized. This challenge is particularly acute because keypoint data is susceptible to high frequency jitter that clustering algorithms can mistake for transitions between behavioral modules. Here we present keypoint-MoSeq, a machine learning-based platform for identifying behavioral modules ("syllables") from keypoint data without human supervision. Keypoint-MoSeq uses a generative model to distinguish keypoint noise from behavior, enabling it to effectively identify syllables whose boundaries correspond to natural sub-second discontinuities inherent to mouse behavior. Keypoint-MoSeq outperforms commonly used alternative clustering methods at identifying these transitions, at capturing correlations between neural activity and behavior, and at classifying either solitary or social behaviors in accordance with human annotations. Keypoint-MoSeq therefore renders behavioral syllables and grammar accessible to the many researchers who use standard video to capture animal behavior.

neuroscience↗

bioRxiv2022

Steroidogenesis and androgen/estrogen signaling pathways are altered in in vitro matured testicular tissues of prepubertal mice

Children undergoing cancer treatments are at risk for impaired fertility. Cryopreserved prepubertal testicular biopsies could theoretically be later matured in vitro to produce spermatozoa for assisted reproductive technology. A complete in vitro spermatogenesis has been obtained from mouse prepubertal testicular tissue, although with low efficiency. Steroid hormones being essential for the progression of spermatogenesis, the aim of this study was to investigate steroidogenesis and steroid signaling in organotypic cultures. Histological, RT-qPCR, western blot analyses and steroid hormone measurements were performed on in vitro cultured mouse prepubertal testicular tissues and age-matched in vivo controls. Despite a conserved density of Leydig cells after 30 days of culture (D30), transcript levels of adult Leydig cell and steroidogenic markers were decreased. Increased amounts of progesterone and estradiol and reduced androstenedione levels were observed at D30, together with decreased transcript levels of steroid metabolizing genes and steroid target genes. hCG was insufficient to facilitate Leydig cell differentiation, restore steroidogenesis and improve sperm yield. In conclusion, this study reports the failure of adult Leydig cell development and altered steroid production and signaling in tissue cultures. The organotypic culture system will need to be further improved before it can be translated in clinics for childhood cancer survivors.

developmental biology↗

bioRxiv2022

Zebrafish caudal fin amputation induces a metabolic switch necessary for cell identity transitions and cell cycle re-entry to support blastema formation and bone regeneration

Regeneration depends on the ability of mature cells at the injury site to respond to injury, generating tissue-specific progenitors that incorporate the blastema and proliferate to reconstitute the original organ architecture. The metabolic microenvironment has been tightly connected to cell function and identity during development and tumorigenesis. Yet, the link between metabolism and cell identity at the mechanistic level in a regenerative context remains unclear. The adult zebrafish caudal fin, and bone cells specifically, have been crucial for the understanding of mature cell contribution to tissue regeneration. Here, we use this model to explore the relevance of glucose metabolism for the cell fate transitions preceding new osteoblast formation and blastema assembly. We show that injury triggers a shift in the metabolic profile at early stages of regeneration, enhancing glycolysis at the expense of mitochondrial oxidation. This metabolic switch mediates transcriptional changes that make mature osteoblast amenable to be reprogramed into pre-osteoblasts and induces cell cycle re-entry and progression. Manipulation of the metabolic profile led to severe reduction of the pre-osteoblast pool, diminishing their capacity to generate new osteoblasts, and to a complete abrogation of blastema formation. Overall, our data indicate that metabolic alterations have a powerful instructive role in regulating genetic programs that dictate fate decisions and stimulate proliferation, thereby providing a deeper understanding on the mechanisms regulating blastema formation and bone regeneration.

developmental biology↗

bioRxiv2021

Motor dysfunction in Drosophila melanogaster as a biomarker for developmental neurotoxicity

Humans interact with numerous chemical compounds with direct health implications, with several able to induce developmental neurotoxicity (DNT), which bear developmental, behavioral, and cognitive consequences from a young age. Current guidelines for DNT testing are notably costly, time consuming, and unsuitable for testing large numbers of chemicals. Therefore, there is a need for adequate alternatives to conventional animal testing for neurotoxicity and DNT. Here we show that detailed kinematic analysis can provide a strong indicator for DNT, using known (chlorpyrifos, CPS) or putative ({beta}-N-methylamino-L-alanine, BMAA) neurotoxic compounds. We exposed Drosophila melanogaster to these compounds during development and evaluated for common general toxicity -- notably developmental survival and pupal positioning, together with the FlyWalker system, a detailed adult kinematics evaluation method. At concentrations that do not induce general toxicity, the solvent DMSO had a significant effect on kinematic parameters. Nonetheless, CPS not only induced developmental lethality but also significantly impaired coordination in comparison to DMSO, altering 16 motor parameters, validating the usefulness of our kinematic approach. Interestingly, BMAA, although not lethal during development, induced a dose-dependent motor decay, targeting most parameters in young adult animals, phenotypically resembling normally aged, non-exposed flies. This effect was subsequently attenuated during ageing, indicating an adaptive response. Furthermore, BMAA induced an abnormal terminal differentiation of leg motor neurons, without inducing degeneration, underpinning the observed altered mobility phenotype. Overall, our results support our kinematic approach as a novel, highly sensitive and reliable tool to assess potential DNT of chemical compounds.

neuroscience↗

bioRxiv2021

Autophagy modulation enhances enzyme replacement therapy response in Fabry disease

SummaryFabry disease is a lysosomal disease due to -galactosidase A (a-GalA) deficiency. Since 2001, Enzyme replacement therapy (ERT) has been used as specific treatment of Fabry disease, with variable effects depending on patient gender and affected organs. In Fabry cells, the endososomal/lysosomal system is highly altered. Consequently, the exogenous enzyme may be mistargeted and trapped into intracellular vesicles instead of reaching the lysosomes. In this study, we aimed to investigate the mechanisms underlying the processing of the exogenous enzyme. We used Fabry cells (cultured fibroblasts and podocyte cell line) to study the enzyme internalization and its effects on the catabolism of the main a-Gal A substrate, globotriaosylceramide (Gb3), upon autophagy inhibition. The exogenous enzyme reaches the early endosome in a similar timeframe in Fabry and control cells, while its targeting to lysosomes is delayed in Fabry cells. Gb3 concentration is lowered upon therapeutic enzyme addition or wortmannin treatment with a synergetic effect. These findings illustrate the positive impact of autophagy inhibition on enzyme trafficking and processing, allowing the increase of functional enzyme rate within the lysosome. Given the high cost of ERT, a better understanding of the cellular fate of the exogenous enzyme may lead to improve its targeting to the lysosome.

biochemistry↗

bioRxiv2021

Cholesteryl Hemiazelate Induces Lysosome Dysfunction and Exocytosis in Macrophages

OBJECTIVEA key event in atherogenesis is the formation of lipid-loaded macrophages, lipidotic cells, which exhibit irreversible accumulation of undigested modified low-density lipoproteins in lysosomes. This event culminates with the loss of cell homeostasis, inflammation and cell death. In this study we propose to identify the chemical etiological factors and understanding the molecular and cellular mechanisms responsible for the impairment of lysosome function in macrophages. APPROACH AND RESULTSUsing shotgun lipidomics we have discovered that a family of oxidized lipids (cholesteryl hemiesters, ChE), end products of oxidation of polyunsaturated cholesteryl esters, occurs at higher concentrations in the plasma of two cohorts of cardiovascular disease patients than in the plasma of a control cohort. Macrophages exposed to the most prevalent ChE, cholesteryl hemiazelate (ChA) exhibit lysosome enlargement, peripheral lysosomal positioning, lysosome dysfunction and lipidosis which are irreversible. The transcriptomic profile of macrophages exposed to ChA indicates that the lysosome pathway is deeply affected and is well correlated with lysosome phenotypic and functional changes. Interestingly, the dysfunctional peripheral lysosomes are more prone to fuse with the plasma membrane, secreting their undigested luminal content into the extracellular milieu with potential consequences to the pathology. CONCLUSIONWe identify ChA not only as one of the molecules involved in the etiology of irreversible lysosome dysfunction culminating with lipidosis but also as a promoter of exocytosis of the dysfunctional lysosomes. The latter event is a new mechanism that may be important in the pathogenesis of atherosclerosis.

cell biology↗

bioRxiv2020

Depletion of senescent cells improves functional recovery after spinal cord injury

Persistent senescent cells (SCs) are known to underlie ageing-related chronic disorders, but it is now recognized that SCs may be at the center of tissue remodeling events, namely during development or organ repair. Here we show that two distinct senescence profiles are induced in the context of a spinal cord injury between the regenerating zebrafish and the non-regenerating mouse. While induced-SCs in the zebrafish are progressively cleared out, they accumulate over time in mice. Depletion of SCs in spinal cord injured mice, with different senolytic drugs, improved locomotor, sensory and bladder functions. This functional recovery is associated with improved myelin sparing, reduced fibrotic scar, attenuated inflammation and increased axonal growth. Targeting SCs is a promising therapeutic strategy not only for spinal cord injuries but potentially for other organs that lack regenerative competence.

neuroscience↗

bioRxiv2020

The Neural Basis for a Persistent Internal State in Drosophila Females

Sustained changes in mood or action require persistent changes in neural activity, but it has been difficult to identify and characterize the neural circuit mechanisms that underlie persistent activity and contribute to long-lasting changes in behavior. Here, we focus on changes in the behavioral state of Drosophila females that persist for minutes following optogenetic activation of a single class of central brain neurons termed pC1. We find that female pC1 neurons drive a variety of persistent behaviors in the presence of males, including increased receptivity, shoving, and chasing. By reconstructing cells in a volume electron microscopic image of the female brain, we classify 7 different pC1 cell types and, using cell type specific driver lines, determine that one of these, pC1-Alpha, is responsible for driving persistent female shoving and chasing. Using calcium imaging, we locate sites of minutes-long persistent neural activity in the brain, which include pC1 neurons themselves. Finally, we exhaustively reconstruct all synaptic partners of a single pC1-Alpha neuron, and find recurrent connectivity that could support the persistent neural activity. Our work thus links minutes-long persistent changes in behavior with persistent neural activity and recurrent circuit architecture in the female brain.

neuroscience↗

bioRxiv2019

Deciphering the many maps of the Xingu, an assessment of deforestation from land cover classifications at multiple scales

Remote sensing is an invaluable tool to objectively illustrate the rapid decline in habitat extents worldwide. The many operational Earth Observation platforms provide options for the generation of land cover maps, each with unique characteristics, as well as considerable semantic differences in the definition of classes. As a result, differences in baseline estimates are inevitable. Here we compare forest cover and surface water estimates over four time periods spanning three decades (1989-2018) for [~]1.3 million km2 encompassing the Xingu river basin, Brazil, from published, freely accessible remotely sensed classifications. While all datasets showed a decrease in forest extent over time, we found a large range in the total area reported by each product for all time periods. The greatest differences ranged from 9% (year 2000) to 17% of the total area (2014-2018 period). We also show the high sensitivity of forest fragmentation metrics (entropy and foreground area density) to data quality and spatial resolution, with cloud cover and sensor artefacts resulting in errors. We further show the importance of choosing surface water datasets carefully because they differ greatly in location and amount of surface water mapped between sources. In several of the datasets illustrating the land cover following operationalization of the Belo Monte dam, the large reservoirs are notably absent. Freshwater ecosystem health is influenced by the land cover surrounding water bodies (e.g. Riparian zones). Understanding differences between the many remotely sensed baselines is fundamentally important to avoid information misuse, and to objectively choose the most appropriate dataset for conservation, taxonomy or policy-making. The differences in forest cover between the datasets examined here are not a failure of the technology, but due to different interpretations of forest and characteristics of the input data (e.g. spatial resolution). Our findings demonstrate the importance of transparency in the generation of remotely sensed datasets and the need for users to familiarize themselves with the characteristics and limitations of each chosen data set.

ecology↗

bioRxiv2019

Identifying relationships between imaging phenotypes and lung cancer-related mutation status: EGFR and KRAS

EGFR and KRAS are the most frequently mutated genes in lung cancer, being active research topics in targeted therapy. Biopsy is the traditional method to genetically characterise a tumour. However, it is a risky procedure, painful for the patient, and, occasionally, the tumour might be inaccessible. This work aims to study and debate the nature of the relationships between imaging phenotypes and lung cancer-related mutation status. Until now, the literature has failed to point to new research directions, mainly consisting of results-oriented works in a field where there is still not enough available data to train clinically viable models. We intend to open a discussion about critical points and to present new possibilities for future radiogenomics studies. We conducted high-dimensional data visualisation and developed classifiers, which allowed us to analyse the results for EGFR and KRAS biological markers according to different combinations of input features. We show that EGFR mutation status might be correlated to CT scans imaging phenotypes, however, the same does not seem to hold true for KRAS mutation status. Also, the experiments suggest that the best way to approach this problem is by combining nodule-related features with features from other lung structures.

bioengineering↗