Biology subjects

Saxena, R.

Publications and source records attributed to Saxena, R..

At least 19 recordsLinked to original sources

bioRxiv2026

Multi-region two-photon calcium imaging in freely moving animals

Complex behaviors emerge from the coordinated activity of neural circuits distributed across widely separated brain regions. Existing optical approaches for monitoring neural activity lack the combined capability to record from anatomically distributed regions with high spatiotemporal resolution in freely moving animals. Here we introduce MINI2P-II, a 1.1-g 2P miniscope that achieves large-scale calcium imaging with high spatial resolution, deep imaging depth and dual-color imaging capability. Together with a dual-scope imaging system and a novel implantable periscope lens assembly, MINI2P-II enables simultaneous 2P imaging in widely separated or anatomically adjacent brain regions. In mice, we demonstrate inter-hemisphere coordination in bilateral motor cortex during climbing, egocentric boundary coding in simultaneous retrosplenial and posterior parietal cortex recordings, and in zebra finches, we showed large-scale behavior-dependent activity in both juvenile and adult individuals. MINI2P-II opens the door to multi-region 2P imaging across species and developmental stages in freely moving animals.

neuroscience↗

bioRxiv2026

Circadian rhythms regulate refractive development across species

Myopia is a rapidly escalating global public health challenge, yet the biological mechanisms linking modern lifestyles to abnormal eye growth remain unclear. Circadian rhythms have been implicated in refractive development, but causal evidence is limited. Here, we integrate population-scale human data with an experimental animal model to determine whether circadian misalignment contributes to myopia. In >265,000 individuals from the Estonian and UK Biobanks, late chronotype was consistently associated with myopia. To assess causality, we experimentally disrupted the alignment between behavioural and environmental rhythms in mice by housing them in non-24-hour light-dark schedules. Exposure to a lengthened cycle (T26) induced a myopic shift that was, notably, reversible in early adulthood. Retinal transcriptomics revealed enrichment of mitochondrial and hypoxia-related plasticity pathways, with transcriptional changes distributed across multiple retinal cell classes. Together, these findings identify circadian misalignment as a conserved and modifiable driver of myopia, highlighting opportunities for novel preventive and therapeutic approaches.

neuroscience↗

bioRxiv2025

Dentate spikes comprise a continuum of relative input strength from the lateral and medial entorhinal cortex.

Hippocampal dentate spikes (DS) are synchronous population events generated in the dentate gyrus thought to support memory processing. DS are classified into type 1 (DS1) and type 2 (DS2) based on current sinks in the outer or middle molecular layer, where the axons of the lateral or medial entorhinal cortex terminate, respectively. This widely used classification method constrains DS into a bimodal distribution without properly testing whether such constraint is appropriate. Here we utilized silicon probe recordings with high spatial resolution spanning all layers of the dentate gyrus and discovered that the contribution of LEC/MEC inputs to DS follows a continuous distribution. We introduce a third type (DS3) that captures a previously unreported component of the distribution: simultaneous current sinks in the outer/middle molecular layer, suggestive of synchronized activation from the LEC/MEC which could facilitate binding. We verify DS3 in several data sets from independent laboratories and characterize their brain state dependence and neural recruitment.

neuroscience↗

bioRxiv2025

Enriched experience increases reciprocal synaptic connectivity and coding sparsity in higher-order cortex

The integration of new information during sleep reshapes cortical representations that support categorical knowledge. Auto-associative attractor network theories predict that reciprocal excitatory connections help form stable categorical attractors, but direct evidence is missing. We tested this using ten weeks of enriched experience (ENR) in mice as a model for knowledge accumulation and recorded single-unit activity across hippocampus and neocortex. ENR induced significant remodeling in high- but not low-level neocortex, with a shift from unidirectional to bidirectional excitatory-excitatory connections, suggestive of increased cell assemblies. This was accompanied by increased inhibitory-to-excitatory connections and sparser, more orthogonal population activity during awake rest and slow-wave sleep, particularly in deep layers. Thus, ENR reorganizes cortical circuits into a symmetric, inhibition-balanced network that improves coding efficiency, supporting long-standing attractor network predictions.

neuroscience↗

bioRxiv2025

Cognitive enrichment improves spatial memory and alters hippocampal synaptic connectivity in a mouse model for early-life stress

Early life stress (ELS) and enrichment often have opposing effects on long-term cognitive abilities. Deprivation, such as institutionalized care during early childhood neurodevelopmental periods, results in lifelong working memory and recall deficits. In contrast, enrichment facilitates new learning and slows cognitive decline due to aging and neurodegenerative diseases. Similarly, in rodent models, enrichment facilitates learning whereas ELS induces prominent spatial memory deficits. Environmental enrichment (EE) and ELS can cause opposing changes in hippocampal structure (e.g. shifts in synaptic density) that largely depend on experimental conditions. However, it remains untested whether EE can rescue the behavioral disruptions caused by ELS and how this would impact the hippocampus at advanced ages. To address this, we conducted a longitudinal study on ELS mice, extensively training them on a cognitive enrichment track (ET) or an exercise alone control track (CT). After this, the mice underwent repeated memory testing followed by brain extraction for anatomical analysis of their hippocampus. We found that ET reversed spatial memory deficits at 6, 13 and 20 months and reduced the number of dentate gyrus (DG) to CA3 synapses. Surprisingly, this reduction occurred at excitatory MF synapses surrounding CA3 somas in the stratum pyramidale--a layer not typically associated with MF terminals. Collectively, these findings suggest that cognitive enrichment during early adulthood may reverse ELS-induced spatial memory deficits by adjusting synaptic connectivity between the DG and CA3.

animal behavior and cognition↗

bioRxiv2025

ADCY3 Ser107Pro links difficulty awakening in the morning to adiposity through circadian regulation of adipose thermogenesis

Modern lifestyles often disturb circadian rhythms, yet the genetic circuits that convert this stress into metabolic dysfunction remain poorly defined. Here, we identify a missense variant in ADCY3 (rs11676272; Ser107Pro) as a pleiotropic regulator of circadian preference and adiposity. Using genome-wide pleiotropy analysis in [~]480,000 UK Biobank participants, we show that the G risk allele (Pro107) increases morningness, BMI, and fat mass in European (N=451,324) and African (N=8,738) ancestry groups, with behavioral amplification by morning difficulty awakening observed in Europeans, power limited interaction modeling in other populations. Structural modeling and transcriptomic analysis suggest this allele destabilizes ADCY3 and alters adipose-specific splicing and expression. In mice, Adcy3 is rhythmically expressed in adipose tissue, with the conserved Pro107 site showing BMAL1 binding and cold-inducible activation. Human adipose ADCY3 expression also increases after weight loss. Together, these findings reveal a genotype-dependent, behaviorally modifiable axis linking difficulty awakening to adipose thermogenesis and obesity risk.

genomics↗

bioRxiv2025

Interleaved Replay of Novel and Familiar Memory Traces During Slow-Wave Sleep Prevents Catastrophic Forgetting

Humans and animals can learn continuously, acquiring new knowledge while integrating it into a lifelong memory pool. In contrast, artificial neural networks (ANNs) suffer from catastrophic forgetting, where new training disrupts existing memories. This issue can be alleviated in ANNs by interleaving training on new tasks with past data; however, whether the brain uses a similar strategy is unknown. In this work, we show that slow-wave sleep interleaves replay of familiar and novel (i.e. hippocampal-dependent) memory traces within individual slow waves, allowing new memories to integrate into the existing cortical pool without interference. This study presents a novel theory for how memory traces acquired across an animals life are organized within the cortical-hippocampal system to support continual learning and suggests novel principles for a broad range of continual learning AI.

neuroscience↗

bioRxiv2025

Drosophila Modeling of Insomnia-Associated Genes Reveals Diverse Underlying Sleep Phenotypes

Insomnia is a prevalent sleep disorder with highly heterogenous manifestations. While data-driven approaches to insomnia subtyping have revealed potential differences between proposed insomnia subtypes and their impacts on overall health, little is known about the genetic factors that underly and differentiate these potential insomnia subtypes. We utilize a human-genetics driven approach to Drosophila modeling to identify the range of sleep traits regulated by insomnia-associated genes. Modeling pan-neuronal loss of Drosophila orthologs of a set of insomnia genes reveals a broad range of sleep phenotypes. Through systematic characterization of traits related to sleep quantity, timing, and quality, we identify genetic factors that co-regulate aspects of the insomnia-associated phenotypic landscape. Out of the 75 insomnia-associated genes identified, only 1/3 had at least one Drosophila ortholog that regulated overall sleep quantity. In contrast, 1/3 of the insomnia-associated genes had at least one Drosophila ortholog that regulated either sleep timing or sleep quality, without impacting sleep quantity. Together this work, in Drosophila, provides support for a genetic influence on the differences between insomnia subtypes.

neuroscience↗

bioRxiv2025

Application of spatial transcriptomics across organoids: a high-resolution spatial whole-transcriptome benchmarking dataset

Stem cell-derived organoids hold promise to model tissue-specific disease. To enable this, it is crucial to assess how transcriptional signatures, cellular organisation and composition of organoids compare to in vivo counterparts. However, technologies which elucidate regional molecular identity, like spatial transcriptomics, have been challenging to apply to organoids. This study presents the first systematic profiling of multiple stem cell derived organoid models (brain, heart muscle, heart valve, kidney, lung, cartilage, and haematopoietic) with Stereo-seq, a full transcriptome, spatial transcriptomics assay using on-chip in situ RNA capture at subcellular resolution. It describes optimisation of this assay to characterise organoids, use of multiple organoid samples on a single chip, assess differences in RNA capture efficiency compared to reference tissues and its limitations. This study introduces a bespoke analysis method that partitions samples into regions and further characterises them. These findings inform future works to characterise organoids using spatial transcriptomics, providing insights in optimising RNA capture of multiple organoids across a chip and novel methods for regional analysis.

bioinformatics↗

bioRxiv2025

Evolution of T cell responses in the tuberculin skin test reveals generalisable Mtb-reactive T cell metaclones.

T cells contribute to immune protection and pathogenesis in tuberculosis, but measurements of polyclonal responses have failed to resolve correlates of outcome. We report the first temporal evaluation of the human in vivo clonal repertoire of Mtb-reactive T cell responses, by T cell receptor (TCR) sequencing at the site of a standardised antigenic challenge. Initial recruitment of non-Mtb reactive T cells is followed by enrichment of Mtb-reactive clones arising from oligoclonal T cell proliferation. We introduce a modular computational pipeline, Metaclonotypist, to sensitively cluster distinct TCRs with shared epitope specificity, which we apply here to establish a catalogue of public Mtb-reactive HLA-restricted T cell metaclones. Although most in vivo Mtb-reactive T cells are private, 10 metaclones were sufficient to identify Mtb-T cell reactivity across our study population (N[≥]128), indicating striking population level immunodominance of specific TCR-peptide interactions that may offer novel approaches to patient stratification and vaccine development.

immunology↗

bioRxiv2025

Drosophila Modeling Identifies Increased Sleep as a Link Between Insomnia and Cardiovascular Disease

Insomnia is a common sleep disorder associated with negative long-term health outcomes, including cardiovascular disease (CVD). We selected 16 genes from 13 insomnia- and CVD- associated genetic loci and disrupted Drosophila melanogaster orthologs in neuronal or cardiac tissue to characterize their roles in regulating sleep and cardiac physiology. Neuronal disruption of four orthologs (APOB, FUR, CYP17A1, and TCF4) resulted in short-sleeping flies, and three (MRAS, HDAC9, and TDRKH) resulted in long-sleeping flies. Short-sleeping fly lines impacted cardiac physiology consistent with links between short or poor-quality sleep and increased CVD in humans. Conversely, heart-specific disruption of five orthologs (RASD1, PHACTR1, CNNM2, MRAS, and TCF4) led to defects in cardiac physiology, with varied effects on sleep. Heart-specific knockdown lines that altered fractional shortening, a measure of cardiac contractility, also influenced long and short sleep states. These findings reveal bidirectional relationships between sleep states and cardiac performance, representing a potential feedback loop linking insomnia and CVD.

neuroscience↗

bioRxiv2025

Interleukin-13-mediated alterations in esophageal epithelial mitochondria contribute to tissue remodeling in eosinophilic esophagitis

BackgroundThe significance of mitochondria in EoE pathobiology remains elusive. ObjectiveTo determine the impact of EoE inflammatory mediators upon mitochondrial biology in esophageal epithelium, the mechanisms mediating these effects, and their functional significance to EoE pathobiology. MethodsMitochondria were evaluated in human biopsies, MC903/Ovalbumin-induced murine EoE, and human esophageal keratinocytes. Esophageal keratinocytes were treated with EoE-relevant cytokines and JAK/STAT inhibitor ruxolitinib. To deplete mitochondria, 3D organoids generated from TFAMloxp/loxp mice were subjected ex vivo to Cre or siRNA against Transcription factor A, mitochondria (TFAM) was transfected into esophageal keratinocytes. Mitochondrial respiration, membrane potential, and superoxide levels were measured. ResultsWe find evidence of increased mitochondria in esophageal epithelium of patients with EoE and mice with EoE-like inflammation. In esophageal keratinocytes, IL-4 and IL-13 increase mitochondrial mass. IL-13 increases mitochondrial biogenesis in a JAK/STAT-dependent manner. In 3D organoids, IL-13 limits squamous cell differentiation (SCD), and this is blunted upon TFAM depletion. IL-13 decreases mitochondrial respiration and superoxide level, although mitochondria remain intact. IL-13-mediated suppression of superoxide was abrogated upon TFAM depletion in esophageal keratinocytes. ConclusionsWe report that increased mitochondrial mass is a feature of EoE. Among EoE-relevant cytokines, IL-13 is the primary driver of increased mitochondrial mass in esophageal keratinocytes by promoting mitochondrial biogenesis in a JAK/STAT-dependent manner. IL-13-mediated accumulation of mitochondria impairs SCD in esophageal keratinocytes and also suppresses oxidative stress, a factor that is known to induce SCD. These findings identify a novel mechanism through which IL-13 promotes EoE-associated epithelial remodeling. Clinical ImplicationThese findings further lay a foundation for exploration of level of esophageal epithelial mitochondria as a predictive biomarker for response to dupilumab. Capsule summaryIL-13 promotes mitochondrial biogenesis in esophageal epithelium, contributing to impaired squamous cell differentiation.

molecular biology↗

bioRxiv2024

Exploring the microbiome of black soil from Central India and the impacts of agricultural practices on soil microbial communities

BackgroundHealthy microbial communities in soil are indispensable for agricultural productivity. The diversity in physiochemical properties, their geological origins, and local climatic conditions lead to the evolution of diverse soil types with distinct bacterial communities. Despite a global distribution, the microbial composition of black soil remains largely unexplored. Here, we investigate the composition, diversity, and functional potential of bacterial communities in black soils from central India. We also examine how common agricultural disturbances like fertilizer application and the growth of crops impact soil communities in a three-year longitudinal field experiment involving three crops. ResultsThe black soil core microbiome was dominated by phyla Actinobacteriota and Proteobacteria members. Analysis of bacterial composition in vermicompost revealed the presence of plant growth-promoting taxa. Among the assessed cropping practices, crop growth significantly affected microbiome composition in soil. We also observed the impact of crops in selectively enhancing the microbiome in the soil, highlighting the role of plant species in shaping community structure. During this experiment, fertilizer treatment did not influence the microbiome significantly. Overall, the data showed the vulnerability and resilience of soil bacterial communities to agricultural disturbances. ConclusionThe microbial composition of black soil in central India was revealed, and the effects of agricultural management on black soil microbial composition were evaluated. The prominent impact of individual plants showed the role of plant-root microbiome interactions in shaping the composition of soil communities. Fertilizer application did not show significant effects during the field experiment, reflecting the resilience of soil communities to short-term disturbances. We also shed light on the functional metagenomic potential of black soil communities and the effect of common agricultural disturbances in a prominent soil type.

microbiology↗

bioRxiv2024

A novel membrane stress-response that blocks chromosomal replication by targeting the DnaA initiator via the ClpP protease

In Escherichia coli, membrane-stress due to interrupted lipoprotein (Lpp) maturation impairs DNA replication and arrests cell growth. How the two disparate processes of Lpp maturation and DNA replication are connected remains unclear. We demonstrate that upon membrane-stress the Rcs stress-response pathway is activated and the replication initiator DnaA is lost, which explains the replication block. However, the Lon protease, a key regulator of the Rcs pathway, is not required for the DnaA loss. We further ruled out the involvement of (p)ppGpp, one of the major mediators of stress-response in bacteria. However, upon deletion of the ClpP protease gene, DnaA was stable, replication initiated, and there was no cell-growth arrest. In wildtype cells, overexpression of DnaA was lethal even without the membrane-stress apparently from hyper-initiation. The hyper-initiation was restrained in {Delta}crp cells, and overexpression of DnaA was able to overcome the growth-arrest. {Delta}fis cells, which were earlier found resistant to the membrane-stress, showed DnaA stability and normal replication upon induction of the membrane-stress. We conclude that DnaA loss suffices to explain the growth-arrest upon the membrane-stress. The stress-response pathway described here appears novel because of its independence from Lon and (p)ppGpp, which have been implicated in other stress-responses that block DNA replication. SignificanceThe seminal observation that DNA replication-stress can block cell division in E. coli (SOS response), introduced the concept of checkpoint control in the cell cycle. Here, we describe a novel checkpoint control that functions in the opposite direction: membrane-stress causing replication block. We show how two apparently unrelated outcomes of reducing a membrane phospholipid, accumulation of a precursor lipoprotein (pLpp) and block of replication initiation, could be linked. The pLpp accumulation stresses the membrane that causes a response culminating in activating the ClpP protease that blocks replication by targeting the initiator DnaA. DnaA being vital and highly conserved, the detail understanding of the response pathway is likely to open new avenues to treat bacterial infection.

microbiology↗

bioRxiv2024

An integrative approach prioritizes the orphan GPR61 genomic region in tissue-specific regulation of chronotype

ObjectivesChronotype, a manifestation of circadian rhythms, affects morning or evening preferences and ease of getting-up. This study explores the genetic basis of morning chronotype and ease of getting-up, focusing on the G protein-coupled receptor locus, GPR61. MethodsWe analyzed the genetic correlation between chronotype and ease of getting-up using linkage disequilibrium score regression with summary statistics from the UK Biobank (n=453,379). We prioritized shared signals between chronotype and ease of getting-up using the Human Genetic Evidence (HuGE) score. We assessed the significance of GPR61 and the lead variant rs12044778 through colocalization and in-silico analyses from ENCODE, Genotype-Tissue Expression, Hi-C, and Knockout Mouse Project databases to explore potential regulatory roles of causal genes. ResultsWe identified a strong genetic correlation (Rg=0.80, P=4.9 x10324) between chronotype and ease of getting-up. Twenty-three genes, including three circadian core clock components, had high HuGE scores for both traits. Lead variant rs12044778 in GPR61 was prioritized for its high HuGE score (45) and causal pleiotropy (posterior probability=0.98). This morningness variant influenced gene expression in key tissues: decreasing GPR61 in tibial nerve, increasing AMIGO1 in subcutaneous adipose, and increasing ATXN7L2 in the cerebellum. Functional knockout models showed GPR61 knockout increased fat mass and activity, AMIGO1 knockout increased activity, and ATXN7L2 knockout reduced body weight without affecting activity. ConclusionsOur findings reveal pleiotropic genetic factors influencing chronotype and ease of getting-up, emphasizing GPR61s rs12044778 and nearby genes like AMIGO1 and ATXN7L2. These insights advance understanding of circadian preferences and suggest potential therapeutic interventions. SIGNIFICANCEThis study investigates the genetic underpinnings of chronotype preferences and ease of getting up, with a focus on the orphan G protein-coupled receptor GPR61 and the locus lead variant rs12044778. By combining genomic data with in silico functional analysis, we provide mechanistic insight into a locus for morning chronotype and ease of getting in the morning. We identified the variant rs12044778 as a key regulator of GPR61 and nearby genes AMIGO1 and ATXN7L2 influencing circadian and metabolic traits. Our findings shed light on the intricate genetic networks governing circadian rhythms, suggesting potential therapeutic targets for disorders of the circadian rhythm.

genomics↗

bioRxiv2024

Genetic variants affect diurnal glucose levels throughout the day

Circadian rhythms not only coordinate the timing of wake and sleep but also regulate homeostasis within the body, including glucose metabolism. However, the genetic variants that contribute to temporal control of glucose levels have not been previously examined. Using data from 420,000 individuals from the UK Biobank and replicating our findings in 100,000 individuals from the Estonian Biobank, we show that diurnal serum glucose is under genetic control. We discover a robust temporal association of glucose levels at the Melatonin receptor 1B (MTNR1B) (rs10830963, P = 1e-22) and a canonical circadian pacemaker gene Cryptochrome 2 (CRY2) loci (rs12419690, P = 1e-16). Furthermore, we show that sleep modulates serum glucose levels and the genetic variants have a separate mechanism of diurnal control. Finally, we show that these variants independently modulate risk of type 2 diabetes. Our findings, together with earlier genetic and epidemiological evidence, show a clear connection between sleep and metabolism and highlight variation at MTNR1B and CRY2 as temporal regulators for glucose levels.

genetics↗

bioRxiv2023

Distinct rich and diverse clubs regulate coarse and fine binocular disparity processing: Evidence from stereoscopic task-based fMRI

While cortical regions involved in processing binocular disparities have been studied extensively, little is known on how the human visual system adapts to changing disparity magnitudes. Even though there is an established correlation of BOLD signal with disparity magnitudes, this correlation is not inherent and instead arises from specific causal interactions within an integrated network. Here, we investigate causal mechanisms of coarse and fine binocular disparity processing using fMRI with a clinically validated, custom anaglyph-based stimulus. Therefore, we use degree (D) and participation coefficient (PC) metrics representing rich and diverse properties of the brain network, respectively. Twenty-six healthy participants were asked to indicate hidden 3D shapes through anaglyph filters at four disparity magnitudes. Our findings reveal significant changes at different disparity magnitudes in terms of D and PC of Middle Temporal (MT), V2, V3 and Superior Parietal Lobule (SPL) across both hemispheres. Of these, MT exhibited overlapping rich and diverse club characteristics among other brain regions. Further, diverse clubs outperform rich clubs in decoding disparity magnitudes irrespective of the hemisphere, thereby reinforcing their integrative network properties. These findings imply that distinct rich and diverse clubs exist and provide functional evidence for the variability in human stereopsis.

neuroscience↗

bioRxiv2023

Identifying novel links between cardiovascular disease and insomnia by Drosophila modeling of genes from a pleiotropic GWAS locus

Insomnia symptoms have been associated with cardiovascular disease (CVD), doubling the risk of incident CVD, but specific shared pathways remain poorly understood. Recently, genome-wide association studies (GWAS) identified genetic loci significantly associated with insomnia symptoms, including one locus (near ATP5G1, UBE2Z, SNF8, IGF2BP1, and GIP) that was previously linked with CVD in an independent GWAS. To evaluate the cell-autonomous role of genes within the 17q21 insomnia and CVD locus, we used Drosophila melanogaster models to perform tissue-specific RNAi knockdown of four conserved orthologues (ATPSynC, Lsn, Bruce, and Imp) in neurons and in the heart. To identify non-cell-autonomous mechanisms, we also assessed heart function in flies with neuronal-specific knockdown and sleep in flies with heart-specific knockdown. Neuronal and cardiac-specific RNAi knockdown of several of the genes conserved in Drosophila led to compromised sleep quality and impaired cardiac performance. Neuronal-specific knockdown of ATPSynC, Imp, and Lsn led to disruptions in sleep quantity and quality. Knockdown of ATPSynC and Lsn in the heart led to significantly reduced cardiac performance without and with cardiac dilation, respectively. Furthermore, Lsn and ATPSynC-suppressed hearts showed disruption in the actin-containing myofibrillar organization and led to a significantly shortened lifespan. Non-cell-autonomous effects were seen both from neurons to heart (Imp), and heart to neurons (ATPSynC and Lsn). Specifically, Imp neuronal knockdown led to a significantly compromised cardiac function, whereas knockdown of ATPSynC and Lsn in the heart led to compromised sleep characterized by increased sleep fragmentation, both accompanied by an increase in inflammation through Upd3, an inflammatory cytokine, in the heart or head, respectively. We also demonstrate disrupted cardiac function or sleep upon cardiac-specific or neuronal-specific overexpression of Upd3, respectively, showing a direct link between cardiac dysfunction and sleep disruption through inflammation. Our study reveals tissue-specific and cross-tissue consequences of Drosophila knockdown of multiple genes at this locus, providing novel insights into potential genetic mechanisms linking CVD and insomnia. Our study also highlights the key role of these four conserved genes in both sleep and cardiac function.

physiology↗