Adaptive behavior under threat requires deciding when to act and when to withhold action to avoid harm, often under conditions where movement, arousal, and task demand covary. Medial prefrontal cortex (mPFC) activity is widely associated with such control, yet it remains unclear whether this activity reflects causal action generation or broader evaluative processes shaped by behavioral state. Here, we combined fiber photometry, single-cell calcium imaging, mixed-effects modeling, and optogenetic inhibition to examine how GABAergic neurons in mouse mPFC represent cues, actions, and outcomes during a series of learned avoidance tasks of increasing complexity that promote cautious responding. By explicitly controlling for baseline activity and movement, we show that much apparent task-related activity in mPFC reflects movement and cue-evoked signals that are also present in a control cortical region, the visual cortex. mPFC GABAergic neurons showed little encoding of simple avoidance contingencies but broadly encoded punished outcomes. A small subset of neurons with strong movement sensitivity encoded more demanding avoidance contingencies requiring selection between action generation and deferment. For equivalent avoidance actions, distinct neuronal populations preferentially encoded either cue onset or the action. Despite this encoding, optogenetic inhibition of mPFC had minimal effects on the learning or performance of the different contingencies. These findings reveal a dissociation between neural encoding and causal necessity, indicating that mPFC GABAergic activity primarily reflects evaluative and contextual aspects of cautious avoidance behavior rather than direct control of action execution.
Science Journals
During neuroinflammation, CD11c<sup>+</sup>CD11b<sup>+</sup> myeloid cells accumulate at the cribriform plate, a key cerebrospinal fluid and antigen outflow site in mice. At this site, podoplanin-expressing cells, including lymphatic vessels and meningeal layers, expand to create a distinct drainage microenvironment. In this study, we sought to characterize myeloid cells, which populate this region, using a mouse model of neuroinflammation, experimental autoimmune encephalomyelitis. Utilizing a combination of immunohistochemistry, flow cytometry, and scRNAseq, we report that macrophages and dendritic cells from this region display unique expressional signatures related to tolerance, cell death, and reduced inflammatory profile. Together, this data supports that myeloid retention at the cribriform plate and olfactory bulb meninges promotes a local immunosuppressive environment.
Tissue microenvironments shape lymphocyte differentiation to align immune function with local physiological demands. Uterine natural killer (NK) cells are critical for reproductive success, yet the molecular cues in the uterus that instruct their specialized identities remain incompletely understood. Here, we identify a TGF-β-dependent differentiation pathway by which circulating conventional NK cells convert into uterine tissue-resident NK cells during murine pregnancy. Loss of TGF-β receptor II expression in <i>Ncr1</i>-expressing cells disrupted this conversion, markedly reducing tissue-resident NK cells in the gravid uterus. Impaired TGF-β-driven uterine tissue-resident NK cell differentiation during murine pregnancy led to abnormal spiral artery remodeling and increased fetal resorption rates at mid-gestation, ultimately reducing litter sizes at birth. Collectively, these findings define TGF-β as a pivotal driver of tissue-resident NK cell differentiation in the gravid uterus and establish a mechanistic framework through which the uterine microenvironment programs NK cell identity to meet the physiological demands of gestation.
The tricarboxylic acid (TCA) cycle enzymes malate dehydrogenase (MDH1) and citrate synthase (CIT1) form a multienzyme complex, referred to as a metabolon, that channels intermediate oxaloacetate between their reaction centers. Given that the MDH1–CIT1 metabolon enhances pathway reactions in vitro, its dynamic assembly is hypothesized to contribute to TCA cycle regulation in response to cellular metabolic demands. Here, we demonstrated that yeast mitochondrial MDH1 and CIT1 dissociated when aerobic respiration was suppressed by the Crabtree effect and associated when the respiratory activity was enhanced by acetate. Pharmacological TCA cycle inhibition dissociated the complex, whereas electron transport chain inhibition enhanced the interaction. The multienzyme complex assembly was related to the mitochondrial matrix acidification and oxidation, as well as cellular levels of malate, fumarate, and citrate. These factors significantly affected the MDH1–CIT1 complex affinity in vitro. Especially, variations in buffer pH within the physiological pH range between 6.0 and 7.0 in the mitochondrial matrix significantly impacted the MDH1–CIT1 affinity. These results demonstrate the dynamic association and dissociation of the MDH1–CIT1 metabolon and its relationship with respiratory activity, supporting metabolon dynamics as an integral factor in metabolic regulation governed by multiple factors such as mitochondrial pH and metabolite levels.
<i>Legionella pneumophila</i> (<i>L.p</i>.), an intracellular bacterial pathogen, hijacks the ubiquitin signaling network of its eukaryotic host cells to establish infection. Two families of <i>L.p</i>. secreted ubiquitin ligases are instrumental in the maturation of the <i>Legionella</i>-containing vacuole (LCV): the SidC/SdcA family, which catalyzes canonical ubiquitination, and the SidE family, which bypasses the E1-E2-E3 enzymatic cascade and directly conjugates ubiquitin to a target through a phosphoribosyl (PR) linkage. Here, we demonstrate that the coordinated activities of these two effector families generate a hyperstable, ubiquitin-rich structure surrounding the LCV. We propose a model in which an initial wave of SidC/SdcA-mediated canonical ubiquitination around the LCV is further modified by SidE family-driven PR-ubiquitination, resulting in a detergent-resistant ‘cloud’. The ‘cloud’ is transient, breaking down as infection progresses, suggesting that <i>L.p</i>. reshapes the properties of the proteinaceous shell surrounding the vacuole to meet changing needs throughout its intracellular lifecycle. This unusual structure likely stabilizes and protects the LCV, shielding it from host defense mechanisms during early infection. Our findings reveal cellular consequences of effector interplay during infection and provide a foundation for future studies into the structure and function of the proteinaceous ‘cloud’ surrounding the LCV.
The central apparatus of motile cilia, consisting of central microtubules and various protein projections, is essential for dictating the ciliary movement. Although three proteins (FAP65, FAP147, and FAP70) have been localized to the C2a projection in <i>Chlamydomonas reinhardtii</i>, the full protein composition and functional roles of the vertebrate C2a remain inadequately defined. Here, we use three knockout mouse models corresponding to their respective homologs (<i>Ccdc108</i>, <i>Mycbpap</i>, and <i>Cfap70</i>) to systematically investigate their functions in vertebrates. Notably, all three knockout strains exhibit distinct phenotypes related to primary ciliary dyskinesia (PCD), including hydrocephalus and sinusitis. The ciliary incorporation of CCDC108, MYCBPAP, and CFAP70 is essential for one another’s stability, with the loss of any single component triggering C2a collapse, which destabilizes the central pair microtubules, and ultimately alters the ciliary movement pattern. Furthermore, we significantly expand the vertebrate C2a proteome by identifying ARMC3 and MYCBP as additional C2a components. Collectively, our findings illuminate the proteomic composition and strict physiological requirements of the vertebrate C2a projection, providing new insights into the molecular pathogenesis of PCD.
Analysis of multimodal and multidimensional data capturing dynamic interactions between diverse cell populations is a current challenge in bioimaging, especially in the context of immunology and immunotherapy research. Here, we introduce Celldetective, an open-source Python-based software tool designed for high-performance end-to-end analysis of image-based in vitro immune and immunotherapy assays. Celldetective is purpose-built for multicondition, 2D multi-channel time-lapse microscopy of mixed cell populations. Although it is optimised for the needs of immunology assays, it is nevertheless broadly applicable to any biological system involving interacting cell populations. The software seamlessly integrates AI-based segmentation, tracking, and automated single-cell event detection, all within an intuitive graphical interface that supports interactive visualisation, annotation, and training options. We showcase its capabilities with original datasets of single immune effector cell interactions with an activating surface mediated by bispecific antibodies and pairwise interactions in antibody-dependent cell cytotoxicity events.
Autosomal monoallelic gene expression and asynchronous replication between alleles are established features of imprinted genes and genes regulated by allelic exclusion. Inactivation/Stability Centers (I/SCs) are recently described autosomal loci that exhibit epigenetic regulation of allelic expression and replication timing, with differences that can be comparable to those observed between the active and inactive X chromosomes . Here, we characterize >100 autosomal loci with allele-specific epigenetic regulation of replication timing and gene expression, defining them as I/SCs. I/SCs are approximately 1 Mbb in size and can contain both protein-coding and noncoding genes. In different single-cell derived clones, these genes may be expressed from a single allele, the opposite allele, both alleles, or not expressed at all. This stochastic, yet mitotically stable, pattern indicates that the choice of which allele is expressed is independent of parent of origin and independent of the expression status of the other allele. Similarly, alleles within I/SCs show varying replication timing, either earlier or later, that is also independent of the other allele. Additionally, we identify syntenic loci in the mouse genome that display epigenetic regulation of allelic replication timing, highlighting the genomic organization and conservation of I/SC-associated regulation between human and mouse genomes. The allele-restricted regulation described here creates extensive cellular mosaicism through a stable epigenetic mechanism. This mosaicism impacts numerous dosage-sensitive genes associated with human diseases such as Alzheimer, Parkinson, epilepsy, deafness, and impaired intellectual development.
How does internal representation held in visual working memory (VWM), known as the attentional template, guide attention in humans? A longstanding debate concerns whether only one (Single-Item-Template theory) or multiple (Multiple-Item-Template theory) items serve as attentional templates simultaneously. Here, we propose a Rhythmic-Item-Template hypothesis, successfully reconciling these seemingly contradictory theories. Using the classical VWM-guided attention task with human participants, we found that two VWM items alternately dominate behavioral guidance in theta-rhythmic (4–8 Hz), with anti-correlated activation states in time, and more importantly, this rhythmic oscillation was not driven by the retro-cue processing. Neural recordings revealed that occipital alpha oscillation (8–14 Hz) governed item-specific prioritization, and its amplitude closely tracked subjects’ behavioral guidance, while frontal theta-oscillations phase-led and coupled with occipital alpha oscillations during the item transition. Our Rhythmic-Item-Template results not only resolve previous Single-Item-Template versus Multiple-Item-Template debate but also advance our understanding of how distributed brain rhythms coordinate flexible resource allocation in multi-item memory systems.
Cephalopod chromatophores are skin pigment organs enabling rapid, neurally controlled camouflage, yet the organization of their motor control remains poorly understood. Previously, we developed CHROMAS, a computer-vision pipeline for high-resolution analysis of chromatophore dynamics (Ukrow et al., 2025). Here, we apply it to investigate motor control and innervation in <i>Euprymna berryi</i> and <i>Sepia officinalis</i>. By segmenting chromatophores into radial slices and analyzing anisotropic deformations, we used dimensionality reduction and source separation to estimate the number and spatial influence of motor neurons controlling individual chromatophores and groups thereof. On average, four independent components were detected per chromatophore, each forming contiguous petal-shaped domains. Clustering thousands of components revealed motor units spanning multiple chromatophores, most involving fewer than 14, with diverse geometries ranging from compact local groups to elongated or fragmented structures; chromatophore pairs were co-innervated more often than expected by chance. Expansion was consistently faster and more stereotyped than relaxation, consistent with active contraction and passive recoil. These results show that chromatophores are not uniform pixels but contrast elements fractionable into sub-territories coordinated across neighbors. This geometry of neural control enables the generation of ‘virtual chromatophores’, that is, functional groupings of adjacent chromatophore territories that act as single units, as well as that of noise in the distribution of pixel shapes.
<p>by Kristian Händler, Varun K. A. Sreenivasan, Violetta Pilorz, Celia Martinez-Perez, Iratxe Elorduy, Tomas J. Casas, Marianne Lehmann, Jon Olano Bringas, Laura Escobar Castañondo, Nora Bengoa-Vergniory, Federico N. Soria, Henrik Oster, Malte Spielmann, Mariana Astiz</p>
The circadian system synchronizes physiology, improving the adaptation to daily environmental changes. In mammals, the central pacemaker, in the suprachiasmatic nuclei (SCN) of the hypothalamus, coordinates “wake” functions by inducing the circadian release of glucocorticoids (GCs). GCs entrain the clocks of a wide variety of tissues through GC receptor (GR) activation, however, the influence of GCs on the SCN is unclear and seems to depend on the maturity of the circuit. During the perinatal period, the mouse SCN express GR and respond directly to GCs while the adult SCN express low GR and have been traditionally considered resistant to GCs. To understand the change of sensitivity to GCs we followed the developmental trajectory of the mouse SCN, and found that while GR is expressed in all SCN cells early in life, it remains expressed mainly in astrocytes in the adult. Using a model of prenatal exposure to GCs, we found that offspring from treated mothers, adapt slower to shifted light–dark cycle and shows reduced expression of GR in SCN astrocytes. The adult SCN astrocytes can indeed sense and respond to GCs with rapid astrocytic Ca<sup>2+</sup> events that propagate across neighboring cells, an effect that is prevented by the specific inhibition of astrocyte–astrocyte communication. Our findings provide a conceptual advance on how the mouse clock develops and on the influence that GCs have on the SCN. This might be relevant to understand how circadian synchrony is restored in conditions of temporal misalignment, such as jet lag.
This cost-effectiveness analysis assesses the lifetime health benefits and costs of invitation to 1-time Helicobacter pylori stool antigen testing added to biennial fecal immunochemical test (FIT) screening compared with FIT alone.
This Viewpoint discusses the US Centers for Medicare & Medicaid Services’ Increasing Organ Transplant Access Model and outlines pathways to monitor the implementation of the model to ensure equitable access for all patients.
Humans across cultures not only share the ability to recognise music but also respond to it through movement. While the sensory encoding of music is well-studied, when and how infants naturally start moving to music is largely unexplored. This study simultaneously investigates infants’ neural (auditory) responses and spontaneous movements to music during the first postnatal year. Neural activity (EEG) and body kinematics (markerless pose estimation) were recorded from 79 infants (aged 3, 6, and 12 months) listening to refrains of children’s music, along with shuffled, high-pitched, and low-pitched versions of the same songs. Neural data revealed that, across all ages, infants exhibit enhanced auditory responses to music compared to shuffled music, indicating that auditory encoding of music emerges early in development. Movement data revealed a different outcome. While coarse auditory-motor coupling is present at all ages, more complex structured movement patterns emerge in response to music only by 12 months. Notably, no age group demonstrated evidence of coordinated movements to music. Additionally, enhanced auditory responses to high vs low pitch were only evident at 6 months, while infants’ movements were better predicted by high-pitched compared to low-pitched music at all ages. This study provides initial insights into how the developing brain gradually transforms music into spontaneous movements of increasing complexity.
Being a fluent language user involves recognizing words as they unfold in time. How does this skill develop over the course of early childhood? And how does facility in word recognition relate to the growth of vocabulary knowledge? We address these questions using data from Peekbank, an open database of experiments measuring children’s eye movements during early word recognition. In an observational study of 26 datasets from over 2500 children ages 6 months to 6 years, we show that word recognition becomes faster, more accurate, and less variable across development, consistent with a process of skill learning. Factor analysis reveals covariation of word recognition speed and accuracy with children’s vocabulary size in cross-sectional analysis. Further, across a range of longitudinal models, speed, accuracy, and vocabulary were coupled. Children with overall faster word recognition tended to show faster vocabulary growth, though developmental growth in word recognition skill was not specifically associated with growth in vocabulary. Together, these findings support the view that word recognition is a skill that develops gradually across early childhood and that this skill is deeply intertwined with early language learning.
Optimised genome editing for precise DNA insertion and substitution using prime editors in zebrafish
CRISPR/Cas9-mediated genome editing has rapidly become a popular tool for studying gene functions and generating genetically modified organisms. However, using this system, stochastic integration of random insertions and deletions restricts precise genome manipulation. Advanced CRISPR/Cas9 technologies using Prime Editors (PEs), Cas9 proteins fused with reverse transcriptase, enable programmed integration of short DNA modifications into the genome. However, its application in precise genome editing in animal models is challenging. Here, we utilise a nickase- and a nuclease-based PE to perform programmed short DNA substitutions and insertions at various loci in the zebrafish genome. Whereas nickase-based PE2 mediated a higher ratio of precise prime edits to the total edits, nuclease-based PEn was more efficient for short DNA modifications, achieving up to 27.3% precise insertion. To further evaluate our approach, we inserted a nuclear localisation signal into a reporter transgene to incorporate longer fragments by prime editing. These gene modifications were transmitted to the next generation. We show that PE-mediated prime editing can efficiently manipulate genome information in zebrafish without using exogenous donor DNA.
The human brain segments continuous experience into discrete events, with theoretical accounts proposing two distinct mechanisms: creating boundaries at points of high <i>prediction error</i> (mismatch between expected and observed information) and high <i>prediction uncertainty</i> (reduced precision in predictions). Using fMRI and computational modeling, we investigated the neural correlates of error-driven and uncertainty-driven boundaries. We developed computational models that generate boundaries based on prediction error or prediction uncertainty, and examined how both types of boundaries, and human-identified boundaries, related to fMRI pattern shifts and evoked responses. Multivariate analysis revealed a specific temporal sequence of neural pattern changes around human boundaries: early pattern shifts in anterior temporal regions (–11.9 s), followed by shifts in parietal areas (–4.5 s), and subsequent whole-brain pattern stabilization (+11.8 s). The core of this dynamic response was associated with both error-driven and uncertainty-driven boundaries. Critically, both error- and uncertainty-driven boundaries were associated with unique pattern shifts. Error-driven boundaries were associated with early pattern shifts in ventrolateral prefrontal areas, followed by pattern stabilization in prefrontal and temporal areas. Uncertainty-driven boundaries were linked to shifts in parietal regions within the dorsal attention network, with minimal subsequent stabilization. In addition, within the core regions responsive to both types of boundaries, the timing differed significantly. These findings provide evidence for two overlapping brain networks that maintain and update representations of the environment, controlled by two distinct prediction quality signals: prediction error and prediction uncertainty.
Gorillas have a polygynous social system in which the highest-ranking male has almost exclusive access to females and sires most of the offspring in the troop. Such behavior results in a dramatic reduction of sperm competition, which is ultimately associated with numerous traits that cause low efficacy of gorilla spermatogenesis. However, the molecular basis behind the remarkable erosion of the gorilla male reproductive system remains unknown. Here, we explored the genetic implications of the polygynous social system in gorillas by testing for altered selection intensity across 13,310 orthologous protein-coding genes from 261 Eutherian mammals. We identified 578 genes with relaxed purifying selection in the gorilla lineage, compared with only 96 that were positively selected. Genes under relaxed purifying selection in gorillas have accumulated numerous deleterious amino acid substitutions; their expression is biased towards male germ cells, and they are enriched in functions related to meiosis and sperm biology. We tested the role of gorilla relaxed genes previously not implicated in male reproductive function using the <i>Drosophila</i> model system and identified 41 novel spermatogenesis genes required for normal fertility. Furthermore, by exploring exome/genome sequencing data of infertile men with severe spermatogenic impairment, we found that the human orthologs of the gorilla relaxed genes are enriched for loss-of-function variants in infertile men. These data provide compelling evidence that reduced sperm competition in gorillas is associated with relaxed purifying selection on genes related to male reproductive function. The accumulation of deleterious mutations in these genes likely provides the mechanistic basis behind the low efficacy of gorilla spermatogenesis and uncovers new candidate genes for human male infertility.
Genetic studies of human embryonic morphogenesis are constrained by ethical and practical challenges, restricting insights into developmental mechanisms and disorders. Human pluripotent stem cell (hPSC)-derived organoids provide a powerful alternative for the study of embryonic morphogenesis. However, screening for genetic drivers of morphogenesis in vitro has been infeasible due to organoid variability and the high costs of performing scaled tissue-wide single-gene perturbations. By overcoming both these limitations, we developed a platform that integrates reproducible organoid morphogenesis with uniform single-gene perturbations, enabling high-throughput arrayed CRISPR interference screening in hPSC-derived organoids. To demonstrate the power of this platform, we screened 77 transcription factors in an organoid model of anterior neurulation to identify <i>ZIC2</i>, <i>SOX11</i>, and <i>ZNF521</i> as essential regulators of neural tube closure. We discovered that <i>ZIC2</i> and <i>SOX11</i> are required for closure, while <i>ZNF521</i> prevents ectopic closure points. Single-cell transcriptomic analysis of perturbed organoids revealed co-regulated gene targets of <i>ZIC2</i> and <i>SOX11</i> and an opposing role for <i>ZNF521</i>, suggesting that these transcription factors jointly govern a gene regulatory program driving neural tube closure in the anterior forebrain region. Our single-gene perturbation platform enables high-throughput genetic screening of in vitro models of human embryonic morphogenesis.
Amateur soccer heading may be associated with acute increases in blood biomarkers of neural damage, findings published in JAMA Neurology suggest.
The Review titled “Chronic, Noninfectious Diarrhea: A Review,” published on March 2, 2026, was corrected to remove a sentence in the Secretory Diarrhea section that misstated the results of a cited reference. This article was corrected online.
This Viewpoint discusses slow life support, a care strategy used when clinicians believe patient recovery is impossible but cannot negotiate full withdrawal of therapies, and provides ethical analysis and practical recommendations for such practices.
This Review discusses the pathophysiology of cardiovascular complications in adults with cyanotic congenital heart disease after early management and examines late complications and management strategies.
Diabetes incidence is rising in the US, with more than 40 million US residents living with this condition. Diabetes morbidity and mortality are unequally distributed, and new diagnoses are higher among American Indian/Alaska Native, Black, and Hispanic adults, compared with White adults. Managing prescription medications is critical to reducing diabetes morbidity, as is controlling risk factors and associated comorbid conditions. However, the cost of pharmacologic therapy has increased substantially, driven in part by the growing use of newer glucose-lowering agents. The high cost of these medications comprises a substantial portion of diabetes-related health care expenditure for individuals with diabetes and may result in restrictive coverage by health plans insuring enrollees with diabetes, further limiting access to care. The out-of-pocket costs faced by patients are particularly burdensome, because patients also face the consequences of adverse downstream outcomes that result from reduced medication adherence due to costs.