Biology subjects

Hill, C.

Publications and source records attributed to Hill, C..

At least 19 recordsLinked to original sources

bioRxiv2026

The molecular structural basis of the braking action of muscle

Skeletal muscles generate force and shortening through a structural powerstroke in actin-bound myosin motors linked to ATP hydrolysis, but they also act as brakes to resist stretching by an external load. The molecular structural basis of muscle braking has remained obscure. Here we show that muscle braking is driven both by structural changes in myosin motors and by distortion of the lattice of thick and thin filaments including their M-band and Z-disk connections. A strained motor conformation is established early during stretch, but force continues to increase as thin filaments are displaced from their symmetrical lattice positions, the lateral shear between thick filaments increases, and the lattice volume decreases. Moreover, whilst the motor conformation rapidly recovers after the stretch, the distortion of the filament lattice is largely maintained, storing the work done on the muscle during stretch and accounting for the residual force enhancement after stretch.

biophysics↗

bioRxiv2026

A Chairside Multimodal Platform for Temporomandibular Joint Biomechanical Assessment: Technical Evaluation and Illustrative Application

BackgroundTemporomandibular joint (TMJ) biomechanics can be characterized by mandibular motion, masticatory muscle activity, and bite force generation. When acquired synchronously, these functional variables can serve as model-ready inputs for subject-specific computational analyses of internal joint mechanics. However, existing tools typically measure these signals using separate hardware and software platforms, limiting synchronized acquisition within a clinically practical chairside workflow. MethodsWe developed and technically evaluated a compact multimodal platform for chairside acquisition of TMJ functional data and demonstrated its analytical utility in an illustrative orthognathic surgery application. The platform integrates motion, bite force, muscle activity, acoustic, and event-timing measurements with software for real-time preview, protocol guidance, and synchronized export. We assessed technical performance and chairside feasibility and analyzed representative pre- and postoperative data from an orthognathic surgery patient using kinematic, force-control, and computational modeling workflows. FindingsMotion capture demonstrated submillimeter accuracy, with static and dynamic errors of approximately 0.04 mm and 0.12 mm. Bite force sensors showed excellent linearity (R{superscript 2} = 0.998). Chairside deployment required approximately 15 minutes each for setup and data collection. The illustrative case demonstrated that synchronized chairside data can support preoperative and postoperative kinematic analysis, bite force control capacity assessment, and estimation of TMJ disc stress. InterpretationThe proposed platform enables time-efficient chairside acquisition of synchronized, model-ready multimodal datasets for quantitative TMJ biomechanical assessment. This platform and workflow could support subject-specific biomechanical analysis and future clinical studies of temporomandibular joint function.

bioengineering↗

bioRxiv2026

Females Adapt to Dietary Protein Restriction on Enhanced Gut-Brain Axis during Aging

Growing evidence supports a critical role for the gut-brain axis in regulating metabolic health, inflammation, and cognitive function during aging. Age-associated gut dysbiosis has been linked to metabolic dysfunction and cognitive decline, with females exhibiting increased susceptibility to these age-related impairments. Diet is a major determinant of gut microbiome composition and function. Previous studies from our laboratory demonstrated that dietary protein restriction (DPR) induces fibroblast growth factor 21 (FGF21), improves metabolic health, and extends lifespan in male mice. However, the effects of DPR on the gut microbiome and associated health outcomes in aged female mice remain poorly understood. Female mice were assigned at 16 months of age to either a normal-protein (NP) or low-protein (LP) diet for 26 weeks. Metabolic assessments included food intake, fasting glucose concentrations, and glucose tolerance testing. Senescence-associated markers in mesenteric white adipose tissue (mWAT), fecal microbiome composition, and behavioral outcomes were evaluated to determine relationships among dietary protein intake, microbial communities, metabolic health, and cognitive function. Low-protein diet significantly improved metabolic health in aged female mice, as evidenced by improved glucose regulation. Microbiome analyses revealed increased abundance of Akkermansia at 17 months and Faecalibaculum in LP-fed animals at 22 months of age. More so, functional profiling and gene set enrichment analyses indicated enrichment of microbial pathways associated with membrane integrity and metal ion binding. Lastly, LP-fed female mice displayed improved memory performance at 22 months of age compared with age-matched NP-fed controls. Collectively, these findings demonstrate that DPR remodels the gut microbiome and improves metabolic and cognitive health in aged female mice. The observed microbial adaptations may contribute to the beneficial effects of DPR on aging-related physiology, highlighting the gut microbiome as a potential mediator of dietary interventions that promote healthy aging.

physiology↗

bioRxiv2026

Putative Herbicolin A, an antifungal lipopeptide produced by Pantoea agglomerans APC 4211 is a promising biocontrol agent against food spoilage fungi

Fungal contamination of food with yeast and moulds is associated with major economic losses due to spoilage and also poses health risks in the form of mycotoxin production. The strain Pantoea agglomerans APC 4211 isolated from leaves of Ilex aquifolium (holly tree) has broad spectrum antifungal activity against a variety of food spoilage fungi. Genomic analysis of the strain confirmed the presence of biosynthetic gene clusters potentially encoding for the enzymatic machinery required for the production of the antifungal lipopeptide herbicolin A. Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) analysis of the cell-free supernatant (CFS) confirmed the presence of molecular masses corresponding to herbicolin A (1300.8 Da), and herbicolin B (1138 Da). Purified herbicolin A has desirable properties for biotechnological applications, including potent antifungal activity against a range of spoilage fungi, thermal stability and resistance to proteases. Herbicolin A has low cytotoxicity against epithelial cell lines and has minimum inhibitory concentrations (MICs) lower than those of some commercial antifungal drugs (0.2 - 2.5 {micro}g/ml). In a model dairy system (10% skim milk), herbicolin A demonstrated excellent solubility and stability, effectively eliminating Aspergillus niger and Penicillium notatum at a concentration of 5 {micro}g/mL. In conclusion, herbicolin A is a potent, naturally occurring antifungal agent with the potential to be applied as a biopreservative in food systems, providing a safe, clean-label, and efficient compound for synthetic preservatives replacement. HighlightsO_LIHerbicolin A has a strong potential as a natural preservative for food protection C_LIO_LIHerbicolin A shows lower MICs than several antifungal agents C_LIO_LIHerbicolin A is stable under heat and resistant to proteolytic degradation C_LIO_LIHerbicolin A has strong solubility and stability in a model dairy system C_LIO_LIHerbicolin A indicates low cytotoxicity against epithelial cell lines C_LI Data summaryThe authors confirm all supporting data, code and protocols have been provided within the article or through supplementary data files.

microbiology↗

bioRxiv2026

Eggerthella lenta evades bacteriophage through reversible megabase-scale inversions of capsular polysaccharide gene clusters

Bacteriophages are a promising tool for microbiome editing, yet their development has been constrained by limited insights into bacteriophage-host interactions within their shared mammalian body habitat. We isolated a lytic phage {Phi}KL11 that efficiently targets a disease-associated member of the human gut microbiota, Eggerthella lenta, during in vitro growth. However, {Phi}KL11 selects for a pre-existing and reversible bacteriophage-resistant sub-population in mice. Long-read sequencing revealed a massive genomic inversion event, representing >50% of the E. lenta genome, enriched in response to bacteriophage infection. Transcriptomics linked this inversion to the altered expression of three capsular polysaccharide synthesis (CPS) gene clusters and transmission electron microscopy confirmed differential capsule production. Finally, we show that {Phi}KL11 has a broad host range attributable to CPS and other strain-variable genes. These findings suggest a previously unrecognized strategy for phage evasion in the gut, involving megabase-scale genomic inversions and reversible capsule variation driving phage resistance.

microbiology↗

bioRxiv2026

Salmonella Genomic Markers for Risk to Food Safety

Foodborne non-typhoidal Salmonella remains a major public health concern, yet many isolates recovered through food surveillance are not associated with human illness. To investigate whether genomic factors influence infection risk, we analysed whole-genome sequencing data from over 900 food and environmental isolates collected through UK Health Security Agency surveillance. Hierarchical clustering and comparative genomics identified distinct lineages associated with clinical cases, which were further contextualised using the global EnteroBase database. By combining pangenome and genome-wide association analyses, we identified distinct lineages within several serovars that differed in their association with human cases. In Salmonella Agona, all clinical isolates belonged to a single lineage carrying a highly conserved 7 kb marker that was absent from low-risk strains and demonstrated strong sensitivity and specificity across global datasets. This marker was located within a prophage closely related to the well-characterised Fels-2 phage and encodes a DNA invertase previously implicated in phase variation, a mechanism that promotes bacterial adaptability. Our findings indicate that infection risk can be structured at the lineage level and associated with mobile genomic elements, particularly prophages, that may contribute to environmental persistence and host adaptation. This work advances genomic surveillance from retrospective linkage towards mechanistic and predictive risk assessment, with direct relevance for supporting risk-based decision-making during outbreak investigations. IMPORTANCENot all strains found in food pose the same risk to human health, yet current surveillance systems generally treat them as equivalent hazards. We analysed over 900 genomes from food and environmental sources and found that human infection risk can be concentrated within specific genetic lineages rather than distributed across an entire serovar. In Salmonella Agona, we identified a highly conserved prophage-associated marker with strong sensitivity and specificity for infection-associated lineages. This marker was located within a prophage related to the well-characterised Fels-2 phage and encoded a DNA invertase previously associated to bacterial adaptation. These findings show how genome sequencing can move beyond outbreak detection to identify lineages with elevated public health relevance. Incorporating such information into surveillance programmes could improve risk-based decision-making, helping public health agencies prioritise investigations and interventions more effectively.

genomics↗

bioRxiv2026

Prevalent Gut Phages Encode Modular Adhesins Mediating Epithelial Binding and Endoplasmic Reticulum Trafficking

Bacteriophages are crucial components of the human microbiome and hold promise as therapeutic agents. Yet, their physical interactions with mammalian cells remain poorly understood. Here, we developed a high-throughput platform to identify phages that adhere to epithelial layers and the proteins that mediate this interaction. The identified phages encode immunoglobulin (Ig)-like domain-containing proteins that, when displayed on a non-adherent phage, confer epithelial binding and internalization in vitro, and increased phage retention in the mouse gut in vivo. Phages encoding these adhesins are among the most abundant and prevalent human gut phages, including crAss-like phages and a distinct cluster of uncharacterized myophages. Domain sequence variation alters epithelial interaction profiles, and internalized phages traffic to the endoplasmic reticulum through the Golgi apparatus, suggesting access to non-degradative internalization pathways. These findings reveal widespread phage-human interactions in the human virome, with potential impacts on health and implications for next-generation phage therapeutics.

microbiology↗

bioRxiv2026

Ratatoskr: A tool for automated retrieval of taxonomic type strain sequences and metadata

Bacterial taxonomic type strains anchor species names to physical and genomic reference material, making them essential for reproducible and comparable prokaryotic research. While reference strains are often well-characterised through curated metadata, nomenclature histories, and sequence records, no single database holds up-to-date information on all these aspects, resulting in fragmented information. Gathering the complete set of information for a type strain is further complicated by inconsistencies in nomenclature between sources due to the often-numerous synonyms that can describe a single strain. As a result, collecting type strain data for taxonomic proposals and emendations can be an onerous task requiring extensive manual curation. To address this issue, we introduce Ratatoskr, a Python-based tool that automates the retrieval of sequences and metadata for bacterial taxonomic type strains. Ratatoskr facilitates this by collecting the latest type strain information of the List of Prokaryotic names with Standing in Nomenclature (LPSN) and using this information to query the BacDive and NCBI databases. By applying known taxonomic synonym information Ratatoskr is able to resolve cross-database inconsistencies and streamline the retrieval process. We show that through its use, Ratatoskr can obtain metadata and sequence data for type strains of bacteria within minutes to seconds, depending on the number of members within the requested taxon. By automating this retrieval, Ratatoskr provides fast, accurate, and readily shareable starting points for studies involving the use of taxonomic type strains and data, such as new taxonomic proposals or emendations. Data summaryRatatoskr was developed using Python 3 and is freely available at https://github.com/Fabian-Bastiaanssen/Ratatoskr under a GPL-3.0 licence.

bioinformatics↗

bioRxiv2025

Bacterial resistance and co-existence with temperate bacteriophages in the human gut

Our understanding of the eco-evolutionary dynamics and consequences of bacteria-phage interactions in nature is limited. By combining longitudinal sampling and analysis of Enterobacterales and associated phages isolated from the human gut, with experimental evolution, our work identifies mechanisms that underpin observed patterns of temperate phage-bacteria co-existence and bacterial resistance in this clinically relevant microbiome. Specifically, we show that limited phage infectivity observed against sympatric and allopatric hosts can be explained by the prevalence of spontaneous prophage induction (SPI) bacterial phenotypes and ecological shifts and turnover in focal host populations at multiple taxonomic levels. Despite the ubiquity of temperate phages in the human gut, these findings challenge the assumption that they are major drivers of within-host bacterial evolution and diversification over short-time scales in this natural ecosystem.

microbiology↗

bioRxiv2025

ACSS2 mediates prenatal alcohol exposure-related morphological and behavioral phenotypes

The metabolic enzyme Acetyl-CoA Synthetase 2 (ACSS2) recently emerged as an unexpected regulator of molecular and behavioral changes associated with alcohol use. Its role during prenatal exposure, however, remains unknown. Here, we use a combination of proteomic, genomic and behavioral approaches to establish ACSS2 as a key mediator of prenatal alcohol exposure-related phenotypes. We define the developmental window during which ACSS2 translocates to nuclei in the mouse brain, and show that alcohol-derived acetate is incorporated into fetal brain histone acetylation in utero. Using genetically engineered mice not expressing ACSS2, we demonstrate that loss of this enzyme attenuates chronic prenatal alcohol exposure-induced craniofacial abnormalities, motor function deficits, cognitive impairments as well as associated chromatin and gene expression changes in the dorsal hippocampus and the cerebellar vermis. Our results outline a previously unknown mechanism underlying prenatal alcohol exposure-related phenotypes regulated by ACSS2, which will inform the development of future therapeutic interventions. HIGHLIGHTSO_LIACSS2 translocates to nuclei during in utero brain development C_LIO_LIAlcohol-derived acetate is incorporated into fetal brain histone acetylation C_LIO_LIPrenatal alcohol exposure results in long-lasting and ACSS2-dependent chromatin and gene expression changes in the hippocampus and cerebellar vermis C_LIO_LILoss of ACSS2 attenuates molecular changes, craniofacial abnormalities and cognitive impairments linked to prenatal alcohol exposure C_LI

neuroscience↗

bioRxiv2025

Distinct distributions of myosin motor conformations during contraction of slow and fast skeletal muscle

Slow skeletal muscles maintain posture and produce graded movement at low metabolic cost. Force development and ATP utilisation during fixed-end contractions are typically five times slower in slow than fast muscles from the same species. Mechanical measurements previously suggested that more myosins are attached to thin filaments during contraction of slow muscle, which seems incompatible with its high efficiency. We therefore used small-angle X-ray diffraction to provide a structural estimate of the fraction of myosins attached to thin filaments in slow muscle. X-ray signals associated with myosin binding to actin indicate that only about 10% of myosin motors are actin-bound during fixed-end tetani of rat soleus slow muscles, compared with about 25% in mouse EDL fast muscle. Moreover, X-ray signals associated with the helical organisation of OFF myosin motors in the thick filaments show that about 70% of myosin motors remain in the OFF conformation during tetanic contraction of slow muscle, compared with only 30% in fast muscle. The much slower force development in soleus muscle also allowed clear separation of early structural changes in thick filaments on activation, some of which are distinct from those reported previously in fast muscles. Moreover, the early structural changes in soleus muscle have about the same amplitude in a twitch and a tetanus, suggesting that they are triggered by thin filament activation rather than thick filament stress, and implying a fast signalling pathway between thin and thick filaments. Key PointsO_LIThe interaction between myosin motors and actin filaments in slow skeletal muscles maintain posture and produce graded movement at low metabolic cost. C_LIO_LIMechanical studies have suggested that more myosins are attached to actin filaments in slow than in fast muscle, but this seems incompatible with its high efficiency. C_LIO_LIWe used X-ray diffraction to show that there are fewer myosin motors attached to actin in slow muscle than in fast muscle because more motors are sequestered on the myosin filament. C_LIO_LIThe slower force development in slow muscle also allowed us to isolate and characterise fast changes in myosin motor conformation associated with activation of the actin filaments. C_LIO_LIThe results reveal a distinct pathway of inter-filament signalling in slow muscle that could help the development of novel therapies for muscle weakness. C_LI

biophysics↗

bioRxiv2025

The metallophore staphylopine is essential for survival of Staphylococcus epidermidis in human synovial fluid

Due to extended life expectancies, prosthetic joint infections are an increasing burden on healthcare institutions worldwide. The most commonly isolated causative agents are staphylococci, though the mechanisms underpinning survival and proliferation in synovial fluid are still not fully understood. In this study, we aimed to identify genes important for survival in synovial fluid using a transposon mutant library and RNAseq. We produced a transposon mutant library, containing approximately 57,000 unique insertion mutants, in Staphylococcus epidermidis strain 846. This library was grown in Muller Hinton broth or processed human synovial fluid samples, and Transposon-directed Insertion Sequencing (TraDIS) was used to identify genes involved in survival in synovial fluid. This identified importance of the his, pur and cnt operons. These genes were also upregulated in both Staphylococcus epidermidis 846 and the model Staphylococcus epidermidis strain RP62A when exposed to human synovial fluid. All these key pathways contribute to production of the metallophore staphylopine. To confirm staphyopine production is essential for survival in synovial fluid, a defined transposon insertion in the gene encoding for staphylopine export (cntE) mutant was used. This demonstrated impaired survival in synovial fluid compared to the wild type. RT-qPCR also showed that cntE was more highly expressed after exposure to infected synovial fluid (where metals will be depleted) than non-infected fluid. In conclusion, TraDIS and RNASeq both identified the importance of staphylopine for survival in human synovial fluid. This suggests an opportunity for exploitation for therapeutic or diagnostic use. Author SummaryStaphylococcus epidermidis is a common cause of prosthetic joint infection, however accurate diagnosis remains difficult. In this work we explored the genetic basis of Staphylococcus epidermidis survival in human synovial fluid using a large transposon mutant library, and identified which genes were differentially expressed upon exposure to the fluid. We found crossover between the datasets, which pointed to the importance of the metal acquisition compound staphylopine. The expression levels of the gene required for staphylopine export were shown to be dependent on the infection status of the individual samples were obtained from. We also found the gene cluster to be conserved in a range of staphylococcal species isolated from cases of prosthetic joint infection. Our work provides a valuable resource in the from of a large transposon mutant library, and provides a greater understanding of the requirements for staphylococcal survival in human synovial fluid, providing potential biomarkers for future diagnostic development.

microbiology↗

bioRxiv2025

A novel approach to Caudoviricetes taxonomy utilising whole proteome structure-structure comparison

Viral proteins often evolve so rapidly that sequence similarity is lost even among conserved functional genes, complicating phylogenetic analysis and classification. This challenge is acute for Caudoviricetes, a highly diverse class of dsDNA tailed bacteriophages recently redefined by the International Committee on Taxonomy of Viruses (ICTV). To address the limitations of sequence-based taxonomy, we present a structure-based classification framework using whole-proteome structural clustering. From 4,082 exemplar genomes, we predicted 445,098 protein structures with ESMFold and clustered them using Foldseek. Genomes were encoded as binary profiles of structural fold presence, and phylogenetic distances were inferred and normalized using Relative Evolutionary Divergence (RED). This yielded a revised structural taxonomy comprising 159 orders, 267 families, 502 subfamilies, and 1,189 genera. We also introduce PhagePleats, a Python tool for classifying novel phage genomes based on structural similarity. Our approach highlights the utility of protein structure for resolving distant viral relationships.

microbiology↗

bioRxiv2025

Stable coexistence of Citrobacter rodentium with a lytic bacteriophage during in vivo murine infection

Bacteriophages are ubiquitously present in bacterial communities, yet phage-bacteria interactions in complex environments like the gut remain poorly understood. While antibiotic resistance is driving a renewed interest in phage therapy, most studies have been conducted in in vitro systems, offering limited insight into the complexity of such dynamics in physiological contexts. Here, we use Citrobacter rodentium (CR), a natural mouse-restricted enteric pathogen and well-established model for human enteropathogenic and enterohaemorrhagic Escherichia coli (EPEC and EHEC) infections, to investigate phage-pathogen interactions in vivo. We isolate and characterise Eifel2, a novel lytic phage infecting CR, and generate anti-phage specific antibodies that enable the visualisation of phage infections in vitro. In a murine model of CR infection, oral administration of Eifel2 led to robust phage replication in the gut without reducing the bacterial burden or infection-associated inflammation, confirming the establishment of a stable coexistence in the gut. Despite the emergence of a sub-population of phage-resistant CR mutants in vivo, they did not undergo clonal expansion, indicating that additional selective pressures impaired their widespread dissemination in the gut. Together, our findings demonstrate that imaging approaches can capture key infection stages in vitro, while in vivo models are essential for capturing the complexity of phage-bacteria interactions. This work highlights the importance of studying phage therapy in host-pathogen contexts that include a normal microbiota and a suitable host environment, where dynamic co-existence rather than eradication may define therapeutic outcomes. ImportanceBacteriophages, or phages, are viruses which can either kill or persist inside bacteria. Current interests in phage biology are in part ignited by the fact that they could be used to treat infections caused by antibiotic-resistant bacteria. However, most of our understanding of phage-bacterial interactions comes from in vitro models and/or in vivo gut models relying on altering the endogenous microbiota. Here we report the finding of a novel phage, Eifel2, which specifically targets Citrobacter rodentium (CR), the mouse equivalent of human diarrhoeagenic E. coli pathogens. Despite effectively killing CR in vitro, CR and Eifel2 develop a coexistence relationship in mice with an intact microbiota. While CR phage-resistant mutants emerge, host and microbial factors constrain their expansion. This work highlights the importance of studying phage therapy in host-pathogen contexts that include the complete microbiota, where therapeutic outcomes may rely on dynamic co-existence and containment rather than eradication.

microbiology↗

bioRxiv2025

Evolutionary implications of stipule occurrence in Permo-Carboniferous wetland forests dominated by Psaroniaceae

Stipules are specialized appendages borne at the leaf base with roles as diverse as sheltering of delicate growing tissues from environmental exposure, vegetative propagation and dispersal, and modification into climbing hooks or protective spines. Stipules are widely present in extant vascular plants, but their origins in geological history remain obscure and must have involved convergent evolution more than once, perhaps as early as the late Paleozoic emergence of forested terrestrial ecosystems. Based on extraordinary collections from the early Permian Wuda Tuff Flora, the earliest known stipule structure in the plant kingdom has been identified in marattialean tree ferns Psaroniaceae, the dominant element of Permo-Carboniferous wetland forests. Psaroniaceous stipules are consistent with protection of juvenile fronds and the stem apex as well as retention of a functional role in mature and withered fronds. Furthermore, the continued and perhaps indeterminate growth and the fully laminated stipules invite speculation of a potential role in vegetative propagation following detachment from the parent frond. However, no direct fossil evidence of stipules acting as vegetative propagules is currently available.

paleontology↗

bioRxiv2025

Plasticity in a bacterial global regulatory switch that drives a shift in antibiotic resistance and virulence

Antibiotic resistance and expression of virulence factors can impact the outcome of infection by Pseudomonas aeruginosa. Pathogenesis is often modelled using the P. aeruginosa PAO1 reference strain but laboratory lineages vary in the sequence and activity of MexT, a global regulator impacting virulence, biofilm formation and ciprofloxacin resistance. We defined the impact of active versus inactive MexT in PAO1 and observed global transcriptomic changes affecting the expression of 900 genes. Phenotyping revealed altered metabolism, antibiotic resistance and virulence - resulting in striking variation across a single model organism. We propose that antibiotic-resistance, introduced during work with the original PAO1 strain, has caused plasticity in mexT that accounts for variation across lineages. To test this, we introduced antibiotic resistance into clinical P. aeruginosa isolates and observed downstream mutations in mexT when selective pressure was removed, supporting the proposed evolutionary pathway. Overall, we have demonstrated the transcriptomic basis of MexT as a phenotypic switch in PAO1 and implicated antibiotic resistance as a cause of downstream changes in mexT in P. aeruginosa. Furthermore, MexS/MexT-regulated efflux is implicated in the antibiotic stress response and virulence, helping identify the mechanisms for rapid phenotypic switching through mexT and confirming that PAO1 is a very different organism compared to most isolates. Improved understanding of the regulatory changes linked to antibiotic resistance is particularly relevant to P. aeruginosa where cycles of antibiotic treatment are common.

microbiology↗

bioRxiv2025

Inhibition of adenylyl cyclase 1 (AC1) and exchange protein directly activated by cAMP (EPAC) restores ATP-sensitive potassium (KATP) channel activity after chronic opioid exposure

Prolonged exposure to Gi/o receptor agonists such as opioids can lead to a sensitization of adenylyl cyclases (ACs), resulting in heterologous sensitization or cyclic AMP (cAMP) overshoot. The molecular consequences of cAMP overshoot are not well understood, but this adaptive response is suggested to play a critical role in the development of opioid tolerance and withdrawal. We found that genetic reduction of AC1 and simultaneous upregulation of ATP-sensitive potassium (KATP) channel subunits, SUR1 or Kir6.2, significantly attenuated morphine tolerance and reduced naloxone-precipitated withdrawal. In vitro models utilized an EPAC2-GFP-cAMP biosensor to investigate sensitization of adenylyl cyclase in SH-SY5Y neuroblastoma cells and HEK{Delta}AC3/6 knockout cells. Acute application of DAMGO significantly decreased the cAMP signal from the EPAC2-GFP-cAMP biosensor, while chronic DAMGO administration resulted in enhanced cAMP production following AC stimulation. Inhibition of cAMP overshoot was observed with naloxone (NAL), pertussis toxin (PTX), and the neddylation inhibitor, MLN4924 (Pevonedistat), as well as co-expression of {beta}-adrenergic receptor kinase C-terminus ({beta}ARK-CT). After establishment of the AC1-EPAC sensitization in the in vitro models, we found that inhibition of AC1 or EPAC enhanced potassium channel activity after chronic morphine treatment, using a thallium-based assay in SH-SY5Y cells. Similar data were obtained in mouse dorsal root ganglia (DRG) after chronic morphine treatment. This study presents evidence for investigating further AC1 signaling as a target for opioid tolerance and withdrawal, by increasing EPAC activity and affecting potassium channels downstream of opioid receptors. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=150 SRC="FIGDIR/small/636278v1_ufig1.gif" ALT="Figure 1"> View larger version (44K): org.highwire.dtl.DTLVardef@185bb82org.highwire.dtl.DTLVardef@d97ea4org.highwire.dtl.DTLVardef@1097addorg.highwire.dtl.DTLVardef@1616fcf_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOGraphical AbstractC_FLOATNO C_FIG

neuroscience↗

bioRxiv2024

Punishing temporal judgement boosts sense of agency and modulates its underlying neural correlates

Feeling in control of ones actions is fundamental to the formation of action-outcome relationships. Reinforcement and its valence also change the action-outcome relationship, either through behavior promotion or diminishment. In this study we evaluated how reward and punishment reinforcement modulate sense of agency, as measured by intentional binding. Moreover, using electroencephalography (EEG) we evaluated how reward and punishment reinforcement changes outcome event related potentials associated with the accuracy of participants judgement of the time interval between a key press and audio tone. We found that punishment reinforcement increased intentional binding between the action and outcome more than reward and control feedback. This was also reflected in the outcome event related potentials, where punishment elicited greater P300s and Late Positive Potentials compared to reward and control. We also found increased N100s and diminished P300s and Late Positive Potentials when the participants did not actively participate in evoking the tone. Taken together, our findings showcase that punishment reinforcement boosts sense of agency and modulates associated neural activity more than reward and no reinforcement, as a function of increasing attention and arousal. These findings illuminate the greater effect punishment reinforcement has on behavior and brain activity by its modification of sense of agency, which is important for the development of treatments in psychiatric and neurological diseases.

neuroscience↗