Neurobiology of Pain

Chronic pain is a major risk factor for depression, yet the molecular mechanisms underlying this comorbidity remain poorly understood, particularly in women. To address this gap, we systematically investigated sex differences in the epigenomic adaptations associated with chronic pain-induced depressive-like behaviors. Neuropathic pain was induced in the mouse using the sciatic nerve cuff model, and molecular analyses were performed in the anterior cingulate cortex (ACC), a key brain region implicated in both pain and affective processing. We profiled genome-wide DNA methylation, three histone modifications (H3K27ac, H3K4me1, and H3K27me3), and gene expression using EM-seq, Cut&Tag sequencing, and RNA-seq, respectively. Differential analyses were conducted for each molecular layer and integrated through gene co-expression network analysis. We found that chronic pain induced extensive remodeling of DNA methylation and histone modification landscapes in both sexes. Strikingly, these changes occurred at largely distinct genomic loci in males and females, revealing pronounced sex-specific epigenetic responses. Despite this divergence, the affected regions displayed similar regulatory organization, including enrichment at shared genic features, transcription factor binding sites, and chromatin profiles. Importantly, these adaptations converged on partly overlapping genes, biological pathways, and co-expression modules across sexes. The most affected gene modules were predominantly associated with synapse-related processes, consistent with previous knowledge, and were closely connected to modules enriched for epigenetic regulatory functions. Together, these findings indicate that chronic pain engages sex-specific epigenetic mechanisms that ultimately converge on common functional outcomes. Such convergence highlights the potential value of targeting sex-specific epigenetic substrates in future therapeutic strategies.

Myofascial pain syndrome (MPS) is a major cause of chronic neck pain, with tissue ischemia implicated as a contributing factor. This prospective, single-arm interventional study evaluated the analgesic effect of ultrasound-guided fascia hydrorelease (US-FHR) performed around arteries supplying the neck in patients with chronic neck MPS. Thirteen adults (median age 53.0 years; 38.5% female) underwent US-FHR targeting the perivascular fascia of either the transverse cervical or dorsal scapular artery using 2 mL of normal saline. Pain intensity was assessed by visual analog scale (VAS) at rest and during movement; disability by the 5-item Pain Disability Index, Japanese version (PDI-5-J); and arterial blood flow volume before and after the procedure. The primary outcome, pain VAS during movement, decreased from 49.0 mm (interquartile range [IQR], 44.5-64.0) at baseline to 22.0 mm (IQR, 14.5-31.5) at 15 min and 22.0 mm (IQR, 14.0-34.0) at 1 week (Hodges&-Lehmann median difference, 30.5 mm [95% CI, 24.5 to 36.5] and 28.5 mm [95% CI, 18.5 to 37.0]; both P < 0.001). Pain VAS at rest improved from 21.0 mm (IQR, 13.0-43.5) to 8.0 mm at 15 min and 1 week (median difference, 14.5 mm [95% CI, 9.0 to 24.0; P = 0.001] and 13.5 mm [95% CI, 6.0 to 21.0; P = 0.007]). PDI-5-J decreased from 17.0 (IQR, 10.5-23.0) to 13.0 (IQR, 4.0-17.5) at 1 week (median difference, 5 [95% CI, 2 to 8; P = 0.004]). Blood flow volume increased from 11.2 mL/min (IQR, 4.5-14.4) to 17.2 mL/min (IQR, 6.1-23.7) immediately after US-FHR (median difference, +4.1 mL/min [95% CI, +2.5 to +8.9; P = 0.001]), although transient. One patient experienced transient bleeding that was promptly controlled. In this single-arm feasibility study, US-FHR around the target artery was simple and safe to perform and was associated with reduced neck pain. Because the study lacked a control group, these preliminary findings should be regarded as hypothesis-generating and require confirmation in controlled trials; they may also inform the future evaluation of MPS in other anatomical regions. Trial registration: UMIN Clinical Trials Registry, UMIN000053612.

Spontaneous activity in primary sensory neurons has been implicated in neuropathic symptoms, yet its earliest origins and immediate functional consequences remain incompletely understood. This gap is especially consequential in chemotherapy-induced peripheral neuropathy (CIPN), where sensory toxicities commonly limit effective cancer treatment. Using in vivo recordings in rats, we show that a single dose of oxaliplatin (OX) induces spontaneous firing within 24 h across touch and proprioceptive low-threshold mechanoreceptor (LTMR) afferents. Spontaneous firing consistently originated distally in peripheral axons and was accompanied by enhanced responses to mechanical stimulation, identifying LTMR sensory endings as the earliest source of spontaneous firing and a common site for spontaneous and stimulus-evoked hyperexcitability. OX also induced early structural abnormalities at sensory endings; however, SF+ LTMRs retained mechanosensory response profiles, indicating that spontaneous firing can emerge within otherwise functional sensory endings. Although coincident spontaneous and stimulus-evoked activity distorted encoding in individual LTMRs, these effects had little impact on population LTMR responses or motor behavior relying on mechanosensory feedback. Together, these findings identify sensory endings as an early target of OX neurotoxicity and demonstrate that spontaneous firing spanning multiple tactile and proprioceptive LTMR submodalities can coexist with largely preserved sensory function, indicating that even broad engagement across mechanosensory pathways is insufficient to disrupt all LTMR-dependent functions. These observations indicate that abnormal afferent activity initiated at sensory endings may be sufficient to engage sensory pathways underlying some paresthetic symptoms while leaving others largely unaffected, whereas progression to chronic neuropathic symptoms may require subsequent recruitment of the dorsal root ganglion.

Early-life adversity durably alters neural development through complex mother-offspring interactions whose underlying mechanisms remain poorly understood. We investigated how neonatal maternal separation (NMS) affects the large repertoire of maternal behaviors and subsequently influences spinal nociceptive circuit development and pain responses in rat offspring. Rat dams underwent NMS from postnatal day 2 (P2) to P12, 3h/day, and maternal behaviors were assessed before and after the separation period. These behaviors were compared to those of control (non-separated) dams. Offspring spinal cord and dorsal root ganglia were analyzed at P14 and P24 for several neurotrophic, glutamatergic, and GABAergic gene expression patterns. Offspring nociceptive sensitivity was also assessed at P24. NMS induced increased maternal behaviors (including longer arched-back nursing, higher nest occupancy, and better pup retrieval efficiency), alongside reduced self-care behaviors. These behavioral adaptations were correlated with spinal gene reprogramming in offspring, characterized by a biphasic developmental pattern. At P14, we observed elevated neurotrophic signaling alongside increased GABAergic and glutamatergic markers. By P24, neurotrophic factors decreased while compensatory changes emerged, yet persistent excitatory-inhibitory imbalances remained evident. Parallel to these results, NMS rats also showed mechanical and thermal hot hypersensitivity at P24. These findings reveal that despite apparent maternal behavioral compensation following NMS, offspring exhibit neurotrophic-driven developmental dysregulation resulting in persistent spinal circuit alterations. The disconnect between maternal behavioral normalization and sustained molecular changes suggests that early separation stress triggers enduring neurobiological cascades independent of ongoing maternal care quantity, with long-term consequences for sensory processing and pain sensitivity.

Exogenous opioids that activate mu-opioid receptors (MORs) in nociceptive circuits mediate transient pain relief lasting minutes to hours but have more limited utility for treating chronic pain. By comparison, electrical or magnetic stimulation of the motor cortex can induce pain relief lasting weeks, for which the underlying mechanisms have remained unclear. Here we report an unconventional role for endogenous opioidergic signaling in the rapid induction of long-lasting analgesia from motor cortical stimulation, which triggers opioid-peptide-dependent neural plasticity in the rostral ventromedial medulla (RVM), a key node in the brain's descending pain control pathways. To dissect the circuit and cellular bases for these effects, we created a miniaturized, millimeter-sized device allowing focal, non-invasive transcranial magnetic stimulation (TMS) of the mouse motor cortex. In mice with chronic neuropathic pain, reflexive and affective pain behaviors diminished for 1-2 weeks after one session of TMS treatment. Chemogenetic and optogenetic manipulations showed that motor cortical layer 5 pyramidal neurons with axonal projections to the RVM mediated TMS-induced pain relief. High-density electrophysiological recordings revealed that TMS treatment shifted the balance of RVM activity between pain-ON and pain-OFF neurons to a state promoting greater suppression of pain. Genetic and neuropharmacological manipulations revealed that NMDA-receptor-dependent signaling and MOR activation by endogenous opioid peptides in the RVM jointly mediate the long-lasting analgesia induced by a transient bout of TMS. Strikingly, enkephalinase inhibition in the RVM during TMS treatment enhanced the amplitude and duration of analgesia, showing that transiently boosting endogenous opioidergic signaling during TMS increases analgesia-conferring plasticity. In accord, re-analyses of data from human subjects with chronic pain support the idea that opioid administration amplifies analgesia from motor cortical TMS. Overall, our results showcase miniaturized TMS devices as versatile tools for basic and translational neuroscience and detail a hybrid, long-range neural network and NMDA- and opioid-receptor-dependent plasticity mechanism for durable pain relief. These findings point the way to mechanistically grounded, synergistic neurostimulation and drug therapies for brain diseases and disorders that jointly target neural circuit and molecular signaling pathways.

Background: Chronic pain is a multidimensional condition that often persists despite conventional treatment and adversely affects multiple domains of daily life. Music listening has emerged as a promising non-pharmacological intervention, with accumulating evidence supporting its beneficial effects on pain and associated psychological outcomes. However, despite growing evidence of efficacy, the translation of music listening into routine clinical practice remains limited, partly because intervention reporting has received comparatively little attention. Objective: To evaluate the effectiveness of music listening interventions for chronic pain and systematically assess the methodological quality and completeness of intervention reporting to identify barriers to reproducibility and clinical implementation. Methods: Systematic searches were conducted in PubMed, Cochrane Library, CINAHL, and Web of Science through June 2025, with no date restrictions on publication. Randomized controlled trials involving adults with chronic pain receiving music listening interventions were included. Two independent reviewers screened studies, extracted data, and assessed risk of bias. Intervention reporting was evaluated using the TIDieR checklist, and a random-effects meta-analysis was performed for pain intensity outcomes. Results: Ten RCTs involving 538 participants were included. Music listening interventions varied substantially in delivery, duration, and music selection procedures, reflecting considerable heterogeneity in intervention design. Most studies reported significant improvements in pain and psychological outcomes. Meta-analysis of eight trials (10 effect estimates), demonstrated a moderate reduction in pain intensity (SMD = -0.53, 95% CI: -0.96 to -0.11, p = 0.014; I2 = 76.2%). Although intervention rationale and procedures were generally well described, reporting of intervention modifications, treatment fidelity, and adherence was frequently incomplete. These reporting deficiencies may compromise reproducibility and limit translation into clinical practice. Conclusions: Music listening appears to be a safe, accessible, and scalable non-pharmacological intervention for chronic pain management, with benefits extending beyond pain reduction to psychological wellbeing, quality of life, and functioning. However, incomplete reporting of key intervention components may limit reproducibility and hinder clinical implementation. Future trials should adopt standardized and transparent reporting standards to facilitate implementation into clinical practice.

Background: Chronic and non-healing wounds remain a major clinical challenge with limited therapeutic options. Angiogenesis and inflammation are central to tissue repair, and mesenchymal stem cells (MSC) contribute to these processes through their trophic and immunomodulatory secretome. Cannabidiol (CBD) exhibits antioxidant and immunomodulatory properties. However, whether CBD-rich Cannabis sativa extract stimulate MSC toward a pro-angiogenic secretome remains unclear. Purpose: This study aims to determine whether purified CBD or a phytochemically CBD-rich full spectrum extract stimulate umbilical cord-derived human MSC (UC-hMSC) to secrete pro-angiogenic factors and enhance endothelial responses relevant to wound healing. Methods: UC-hMSC were preconditioned with either purified CBD or a CBD-rich full-spectrum extract. Transcriptional changes were assessed by qPCR. The functional impact of the resulting secretome was evaluated in vitro using HUVEC-based proliferation and tube formation assays, and in vivo through the chick chorioallantoic membrane assay. To explore underlying mechanisms, we examined HIF-1 stabilization and VEGFA release in UC-hMSC, and VEGFR-2/ERK signaling in HUVEC. Results: Purified CBD and full-spectrum CBD extract preconditioned UC-hMSC secretomes, increased HUVEC proliferation, tube formation, and enhanced vascular branching in the CAM assay. Mechanistic analyses indicated activation of the HIF-1/VEGF axis in UC-hMSC, and ERK1/2 activation in HUVEC that was sensitive to VEGFR-2 blockade. Conclusion: Purified CBD and CBD-rich full-spectrum extract prime UC-hMSC toward a pro-angiogenic secretome that promotes endothelial activation and neovascularization. These findings suggest that cannabinoid-based preconditioning of UC-hMSC involves the HIF-1/VEGF axis and VEGFR-2/ERK signaling pathways in endothelial cells, supporting further investigation of this approach in wound healing and regenerative therapies.

Fibroproliferative diseases such as systemic sclerosis are complex conditions characterized by chronic skin inflammation and progressive fibrosis, with fibroblast activation as a central feature. While Transforming Growth Factor Beta (TGF-{beta}) signaling is a well-established driver of fibrosis in SSc, inflammatory pathways such as Nuclear Factor Kappa B (NF-{kappa}B) also contribute substantially to disease morbidity. We previously identified CD109 as a TGF-{beta} co-receptor and negative regulator of fibrotic signaling; however, its role in inflammatory signaling remains unknown. Here, we investigate the function of CD109 in regulating inflammatory signaling in skin fibroblasts. We show that, CD109 co-localizes and associates with Toll-like receptors (TLR2, TLR4) and tumor necrosis factor receptors (TNFRI, TNFRII), and that loss of CD109 enhances TNF--induced NF-{kappa}B activation and reprograms cytokine production in human dermal fibroblasts. Furthermore, both global and fibroblast-specific CD109 knockout mice exhibit increased immune cell infiltration and skin inflammation. In parallel, single-cell transcriptomic analyses across a pan-disease fibroblast atlas show that CD109 expression is preferentially maintained in structural and homeostatic fibroblast subtypes, whereas immune-interacting fibroblast subsets consistently display decreased CD109 levels. Pathway-level analyses of fibroblast pseudobulk samples reveal altered activity of canonical inflammatory pathways in SSc compared to healthy skin. Together, these findings identify CD109 as a fibroblast-intrinsic negative regulator of inflammatory signaling and suggest a broader role for CD109 in modulating inflammatory responses in systemic sclerosis. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=102 SRC="FIGDIR/small/736423v1_ufig1.gif" ALT="Figure 1"> View larger version (53K): org.highwire.dtl.DTLVardef@be9e08org.highwire.dtl.DTLVardef@794173org.highwire.dtl.DTLVardef@b81eb5org.highwire.dtl.DTLVardef@1e811f5_HPS_FORMAT_FIGEXP M_FIG Graphical Abstract: CD109 Restrains Fibroblast-Driven Inflammation by Modulating NF-{kappa}B Signaling. Generated using FigureLabs.ai and edited using Adobe Photoshop. C_FIG

The neuromuscular junction (NMJ) is a specialised synapse between motor neurons and skeletal muscle, and its progressive deterioration contributes to age-related and metabolic disease-associated declines in muscle function. Advanced glycation end-products (AGEs) accumulate in tissues during ageing, diabetes, and chronic metabolic dysfunction and have been implicated in neuromuscular degeneration, yet their effects on the intact NMJ have not previously been examined in a human model system. This study employed a fully human, serum-free, and neural growth factor-free NMJ co-culture system, combining neural progenitor cells with immortalised human myoblasts derived from an 83-year-old donor, to investigate the effects of AGE exposure on neuromuscular integrity across structural, metabolic, functional, and secretory outcomes. AGE exposure induced significant reductions in motor neuron axonal length, myotube remodelling with centralised nuclear positioning, mitochondrial membrane depolarisation, elevated mitochondrial superoxide production, mitochondrial uncoupling, and reductions in spontaneous contraction intensity and frequency. Neurotrophic and myogenic growth factor signalling was significantly downregulated in AGE-treated co-cultures. These findings identify the NMJ as a sensitive target of glycation stress and establish this fully human co-culture platform as a physiologically relevant model for investigating glycation-related neuromuscular pathology and evaluating candidate therapeutic interventions.

Cav1.2 is an L-type voltage-gated Ca2+ channel (VGCC) that supports Ca2+ influx in response to membrane depolarization. Ca2+ entering via Cav1.2 alters gene expression, activates Ca2+-dependent enzymes and has been implicated in synaptic plasticity. ORL-1 is a Gi/o-coupled G protein-coupled receptor (GPCR) that is expressed in the peripheral and central nervous systems. Both Cav1.2 and ORL-1 are expressed in the hippocampus, where they have been implicated in learning and memory. It is well-documented that ORL-1 interacts with another VGCC, Cav2.2. However, less is known about potential interactions between Cav1.2 and ORL-1. Here, we examine the interplay between Cav1.2 (Cav1c, Cav2{delta}-1, Cav{beta}1) and ORL-1 co-expressed in tsA-201 cells by using biochemical, electrophysiological and confocal imaging analysis. Co-immunoprecipitations revealed that ORL-1 independently interacts with Cav1c and Cav2{delta}-1 subunits of the Cav1.2 channel complex. Electrophysiological recordings revealed that co-expression with ORL-1 reduced Cav1.2 peak current density without altering its biophysical properties. Acute perfusion with the ORL-1 receptor agonist nociceptin (1 M) did not alter Cav1.2 current density. Confocal imaging experiments revealed that ORL-1 significantly decreases Cav1.2 plasma membrane expression by disrupting forward trafficking. Interestingly, ORL-1 did not affect Cav1.2 endocytosis. Overall, our results demonstrate a previously unrecognized interaction between ORL-1 and Cav1.2 that alters Cav1.2 membrane expression without affecting biophysical properties.

An autocrine pro-adhesive/pro-contractile signaling loop, through the mechanosensitive transcriptional cofactor YAP, promotes fibrosis. The CCN family of matricellular proteins modify adhesive signaling. Of these, CCN3 is antifibrotic. We show that BLR-200, a CCN3-derived peptide, has anti-fibrotic properties in the bleomycin-induced model of scleroderma skin fibrosis. In vitro, BLR-200 delayed, but did not abolish, fibroblast adhesion to collagen and nuclear YAP localization. In vivo, BLR-200 prevented/treated bleomycin-induced skin fibrosis, and reduced bleomycin-induced expression of profibrotic genes including alpha-smooth muscle actin, CCN1 and CCN2. Lineage tracing and scRNA-seq analyses revealed that the myofibroblasts in this model were quantitatively derived from collagen-lineage Pi16+/Col15+ve fibroblasts. BLR-200 prevented myofibroblast differentiation in this model and trajectory of fibroblasts toward a Sfrp2-positive subset, a cell type associated with poor clinical outcome. BLR-200 impairs YAP activation in vitro and appearance of translationally-relevant fibroblast subtypes in vivo and is a novel anti-fibrotic agent for SSc skin fibrosis.

The intestinal mesentery has been recently classified as a 'new' organ and contains various cell types including adipocytes, preadipocytes, endothelial cells, and immune cells. In addition, neuronal cell bodies are found in the small intestinal mesentery and are situated either individually or clustered together with glial cells in small ganglion structures close to the gut wall. However, little is known about the origin or function of these extra-intestinal mesenteric neurons. The aim of this study was to better these characterize mesenteric neurons and to examine their connectivity with the ENS using calcium imaging in adult mouse ileum with the mesentery attached. Here we show that neurons in the mesentery express typical ENS neurochemical markers, respond to 5-HT, ATP and the nicotinic agonist DMPP, and receive nicotinic synaptic inputs. Furthermore, using labeling with the neuronal tracer DiI, some mesenteric neurons were found to project into the gut wall and can provide functional excitatory inputs to myenteric neurons. By contrast, we did not find evidence for mesenteric neurons providing inputs to other extrinsic neuronal targets, suggesting that they preferentially interact with the ENS. We also demonstrate that mesenteric neurons can be activated by intestinal distension and that the mesentery provides a source of inhibition to the myenteric plexus. Taken together, we show that the ENS not only interacts with vagal and spinal afferents, and sympathetic and parasympathetic nerves, but also neurons situated in the mesentery. Finally, our data suggest that these neurons may provide a form of negative feedback to the myenteric plexus such as in the event of intestinal distension. These findings have important implications for the regulation of intestinal motility in physiological and pathophysiological conditions.

Spinal cord injury (SCI) leads to the formation of a chronic scar composed of glial and fibrotic components that severely restrict neural regeneration and functional recovery. While the scar composition has been widely studied, the spatiotemporal evolution of tissue mechanics and the role it plays in regulating the post-injury responses remain poorly understood. Here we present an integrated mechanobiological and multi-omics analysis of spinal cord remodeling following a severe thoracic contusion injury. Using nanoindentation and viscoelasticity measurements taken via atomic force microscopy (AFM), we demonstrate that SCI induces a dynamic mechanical response characterized by rapid tissue softening during the acute phase reaching a minimum at one-month post-injury, followed by progressive stiffening associated with chronic scar maturation at six months. Bulk RNA sequencing reveals that early mechanical softening coincides with strong activation of inflammatory and matrix-degrading pathways whereas chronic stiffening correlates with upregulation of collagen synthesis, extracellular matrix (ECM) organization and fibrotic remodeling pathways. Concurrently, mechanotransduction regulators exhibit temporally coordinated activation, indicating that cells dynamically sense and respond to evolving mechanical cues. Viscoelastic analysis further shows that chronic scar tissue exhibited increased stiffness and prolonged relaxation dynamics, reflecting dense collagen deposition and proteoglycan accumulation that reinforces a mechanically restrictive microenvironment. Together, these findings establish that the post-injury scar represents a dynamic mechanobiological system in which the evolving tissue mechanics, viscoelasticity and mechanotransduction collectively regulate ECM remodeling, resulting in regenerative failure. This study provides a comprehensive mechanobiological framework for SCI progression and highlights the opportunities for mechanically informed therapeutic strategies aimed at modulating scar mechanics to promote tissue repair.

We showed before that the endogenous peptide Nociceptin blocks the premature differentiation of oligodendrocytes (OLGs), preventing untimely precocious myelination in the developing brain. Consistent with this early function, Nociceptin brain expression is developmentally regulated, sharply decreasing with the initiation and progression of myelination. However, we now found that at difference with controls and relapsing-remitting multiple sclerosis (RRMS), Nociceptin levels are highly elevated in cerebrospinal fluid from patients with the most severe progressive MS (PMS) forms. This questioned whether Nociceptin early developmental effects could be latter recapitulated, interfering with remyelination in PMS. This possibility was tested by inducing experimental autoimmune encephalomyelitis in older mice, at an age equivalent to that with increased risk of RRMS transition into PMS. Older animals develop persistently highly debilitating clinical symptoms, and display both brain and spinal cord demyelination. Importantly, these mice exhibit elevated brain Nociceptin levels, and their treatment with an antagonist of the Nociceptin receptor (NOR) elicits a regression of clinical scoring that is accompanied by higher ratios of OLGs/OLG progenitor cells, increased myelination, and reduction of reactive astrocytes. These findings suggest that Nociceptin may be a crucial player in the age-related progression of MS; interfering with OLG maturation and remyelination, and perhaps further exacerbating neurological dysfunction by targeting astrocyte populations. The upregulation of Nociceptin secretion by human astrocytes in response to proinflammatory cytokines, also points to this peptide as a mediator of microglia-astrocyte interactions supporting MS progression with aging. NOR may offer a novel pharmacological target for ameliorating the devastating effects of MS progression.

Social media users often feel that time vanishes while scrolling, but real feeds confound novelty, rewards, social signals, and self-paced control, leaving the driver of this distortion unclear. We tested whether self-paced visual exploration is sufficient to compress subjective time by comparing active scrolling with passive, yoked viewing and a static baseline. Twenty-three adults viewed sequences of natural images under three within-subject conditions: Scrolling (self-paced mouse clicks), Watching (a passive, yoked replay of their own scrolling sequence), and a Baseline (a static image). Participants estimated the elapsed duration of each block. Subjective duration was most compressed under Scrolling (48% of elapsed time), followed by Watching (51%) and Baseline (65%). Two sources separated these effects. Adding back the empty inter-image fixations brought the image-rich conditions to within seconds of the Baseline, showing that observers barely counted the blank gaps; the Scrolling--Watching difference, by contrast, was independent of these shared gaps, isolating self-paced control as a second source of compression. Electrophysiology linked that control to anticipatory neural states and the timing of early visual responses, with no amplified encoding of individual images. The results favor an attention-weighted account of timing, on which subjective duration tracks how much attention reaches the clock, a resource that a self-paced stream and its uncounted gaps both draw away.

Recent advances in spatially resolved transcriptomics (SRT) enabled measurement of sets of pathway genes activity within tissues. However, existing gene set activity scoring methods overlook spatial dependencies among tissue locations, restricting their ability to capture region-specific pathway activities associated with disease pathology or cellular communication. Moreover, these methods lack significance-level inference for activity scores, provide limited interpretability of gene-level contribution to a pathway, and scale poorly to advanced large-size SRT datasets. To address these limitations, we present GESSO (Gene sEt activity Score analysis with Spatial lOcation), a spatially informed gene set scoring method adaptable to diverse SRT platforms. GESSO models gene set activity levels through a graph-regularized matrix decomposition algorithm, jointly inferring spatially coherent gene set activity scores (GASs) and interpretable metagene weights that capture gene-level contributions. It further implements a permutation-based local significance test and a stratified low-resolution approximation that scales to high-resolution SRT datasets such as Visium HD, Stereo-seq, and Xenium Prime. Across 13 datasets from five SRT platforms, GESSO outperformed all existing methods in accuracy, calibration, interpretability, and scalability. Applications revealed novel biological programs, including spatially confined EMT activation within tumor-stroma interfaces, developmental signaling gradients across embryonic tissues, and coordinated B-cell, T-cell, and signaling pathways within germinal centers of human lymph node tissue, revealing the spatial organization of immune function at subregional resolution.

Medical foundation models compress biomedical data into embeddings that support diverse downstream clinical tasks. However, successful model deployment is hampered by performance degradation on external data. It is recognized that embeddings capture acquisition signatures, such as hardware and technical differences, in addition to biology. Effective harmonization must remove the acquisition signature while preserving biological signals, a trade-off that current methods fail to balance adequately. Input-level normalization fails to eliminate acquisition signatures from embeddings, whereas embedding-level methods adjust features in an untargeted manner. We present FEATMAP, a harmonization approach that models acquisition signatures as geometric distortions between manifolds of similarly arranged embeddings. Using paired data that isolate the effect of acquisition signatures, FEATMAP fits a single global affine transformation per foundation model to correct acquisition signatures directly in the embedding space. This targeted, reusable correction aims to preserve biological and demographic variation while harmonizing across acquisition signatures. Across scanner and foundation-model harmonization in digital pathology and field-strength harmonization in brain MRI, FEATMAP improves cross-condition embedding similarity, reduces performance gaps without retraining, and suggests potential for the alignment of disparate embedding spaces.

Heterochromatin protein 1 (HP1 is an intrinsic component of heterochromatin domains where it is involved in a diverse set of functions including heterochromatin spreading and organization, chromatin compaction and transcriptional silencing. It has been suggested that HP1 functions through a phase separation mechanism, a process that has been observed in vitro in the presence of N-terminal phosphorylation, nucleic acids and nucleosome arrays. HP1 can also interact with numerous binding partners that contain a specific motif called an HP1 access code (HAC). HACs recognize and bind to an interface formed by the chromoshadow (CSD) domains in the HP1 homodimer, the functional form of the protein. It has been shown that some HP1 binding partners can enhance its phase separation ability while others disrupt the process. Here, we focus on the interactions between HP1 and three binding partners, namely the p150 subunit of the chromatin assembly factor 1 (CAF-1), the N-terminal domain of the lamin B receptor (LBR), and the mitotic protein Shugoshin 1 (Sgo1). Using phase separation assays, we show that CAF-1 prevents HP1 phase separation while LBR and Sgo1 enhance it. Binding assays, mutational studies, NMR spectroscopy and computational analysis allow us to dissect the contributions of the HAC motifs, the charge patterns of the binding partner sequences and the role of N-terminal phosphorylation on HP1 in condensate formation. Our results demonstrate that each binding partner uniquely balances these contributions to modulate the properties of HP1, while electrostatic interactions dominate the regulation of phosphorylated HP1. These results suggest that HP1 binding partners play an important role in the modulation of its properties and the regulation of its functions in distinct biological contexts.

Temporal niche partitioning is a strategy for reducing interspecies competition and strengthening reproductive isolation. It relies on animals confining their daily activity to distinct diurnal, crepuscular, or nocturnal windows. However, a hardwired temporal niche is only advantageous under stable, predictable ecological regimes; surviving dynamic environments demands behavioral flexibility. Yet, it remains unclear how animals override rigid biological constraints to rapidly exploit transiently available fitness-critical time windows. To address this, we leveraged the twilight-active, species-rich Drosophila genus and monitored their daily activity under naturalistic conditions. Here, we show that intense sociosexual interactions rapidly drive a species-specific reformatting of their canonical crepuscular niche. The dominant sensory modality used for sexual communication predicts niche shift direction: reliance on chemosensation for courtship redirects behavioral activity into the night, while visual reliance shifts it into the day. This temporal plasticity bypasses the circadian clock, instead operating via a conserved dopaminergic pathway. Dopamine operates a dual-output brain circuit that simultaneously inhibits sleep and sustains sexual motivation. Our results reveal how mating imperatives decouple behavioral timing from circadian command, enabling conditional colonization of otherwise restricted temporal windows. Ultimately, by driving the divergence of previously overlapping niches, sociosexually induced temporal plasticity provides a powerful mechanism for sympatric coexistence in crowded environments.

Single-nucleus transcriptomic atlases offer an unprecedented opportunity to connect cellular molecular states with Alzheimer's disease (AD) neuropathology, but whether these profiles encode reproducible, predictive information about pathological burden remains unclear. We present the SEA-AD DREAM Challenge, an open, international, model-to-data competition built on the Seattle Alzheimer's Disease Brain Cell Atlas to predict Alzheimer's disease neuropathological severity from single-nucleus RNA-sequencing data. Participants developed containerized models to predict categorical neuropathological staging, including overall Alzheimer's disease neuropathologic change, Braak stage, Thal phase, and CERAD score, as well as quantitative amyloid-{beta} and phospho-tau burden measured by 6E10 and AT8 immunohistochemistry. Across 17 eligible teams from 15 countries, the crowdsourcing framework enabled systematic comparison of diverse computational approaches and surfaced a broad landscape of modeling strategies and candidate predictive features. Top-performing methods achieved near-perfect prediction of categorical staging, with the best submission reaching a quadratic weighted kappa of 1.0 for the Overall AD Neuropathological Change score (ADNC), and competitive prediction of quantitative pathological burden in held-out data, with a best concordance correlation coefficient of 0.48. Post hoc perturbation analyses revealed that top categorical-stage predictions relied heavily on donor-level metadata-driven signals rather than transcriptomic features, whereas quantitative pathology prediction was more robust and supported by transcriptomic and cell-type-associated features with potential biological relevance to AD progression. The challenge also introduced the first AI Agent Track in a DREAM Challenge, providing an early benchmark for autonomous and human-guided agentic model development in single-cell neuroscience. This work demonstrates that single-nucleus transcriptomes encode substantial information about Alzheimer's disease pathology, establishes a reproducible benchmark for molecular neuropathology prediction, and highlights critical principles for designing privacy-preserving, leakage-aware community challenges using deeply phenotyped human brain data.