bioRxiv Subject Collection: Neuroscience's Journal

Cross-region neuron co-firing mediated by ripple oscillations supports distributed working memory representations.
High-frequency (~90Hz) ripple oscillations may promote integrative processing in mammalian brains. Previous work has demonstrated that the co-occurrence of these ripple oscillations is associated with enhanced temporal binding of neural activity between human cortical neurons separated by up to 12mm. However, it remains unclear whether co-ripple facilitation of neuronal coupling supports cognitive processing, or if it occurs at greater distances. Here, we analyze intracranial recordings from patients implanted with microwire electrodes in the hippocampus, amygdala, ventromedial prefrontal cortex, anterior cingulate cortex, and pre-supplementary motor area, bilaterally, during a working memory task. We demonstrate that ripple oscillations significantly increase in all recorded regions, during encoding, maintenance and retrieval. Furthermore, co-occurrence of ripples increases between brain regions, associated with ~30% increases in cross-region co-firing, both without decrement over distances up to 220 mm. These increases in cross-regional co-rippling and associated co-firing scale with memory load during maintenance and retrieval. Critically, during retrieval, co-ripples promote the reinstatement of the stimulus-specific long-distance co-firing patterns observed during encoding, especially during rapid recognition. Our findings reveal that ripple oscillations orchestrate long-range neural communication that supports distributed neural representations during human cognition.

TBK1 activity regulates the directionality of axonal transport of signalling endosomes
The polarised and complex morphology of neurons pose massive challenges for efficient cargo delivery between the axon and soma, a process termed axonal transport. We have previously shown that the retrograde axonal transport of pro-survival, neurotrophic signalling endosomes relies on Rab7 in motor neurons, and that their trafficking is impaired in the early stages of amyotrophic lateral sclerosis (ALS) pathogenesis. Here, we report the effect of Rab7 phosphorylation on the transport of these signalling endosomes. We show that the ALS-linked kinase TBK1 phosphorylates Rab7 at S72 in neurons, altering its binding to cytoplasmic dynein adaptors. Accordingly, both TBK1 knockdown and the expression of a loss-of-function Rab7 mutant (S72E) induce aberrant bidirectional movement of signalling endosomes without modifying neuronal polarity or endosomal sorting. This alteration is specific for signalling endosomes, as axonal transport of lysosomes and mitochondria remain unaffected. We have therefore discovered a new TBK1 function that ensures the unidirectional transport of signalling endosomes, suggesting that reduced TBK1 activity determines retrograde transport dysfunctions and long-range signalling impairments.

The human cerebellum encodes temporally sensitive reinforcement learning signals
In addition to supervised motor learning, the cerebellum also supports nonmotor forms of learning, including reinforcement learning (RL). Recent studies in animal models have identified core RL signals related to reward processing, reward prediction, and prediction errors in specific regions in cerebellar cortex. However, the computational constraints on these signals remain poorly understood, particularly in humans. Here, we investigated cerebellar RL signals in a computationally-driven fMRI study. Human participants performed an RL task with no low-level sensorimotor contingencies. We observed robust RL signals related to reward processing and reward prediction errors in cognitive regions of the cerebellum (Crus I and II). These signals were not explained by oculomotor or physiological confounds. By manipulating the delay between choices and reward outcomes, we discovered that cerebellar RL signals are temporally sensitive, becoming undetectable at supra-second feedback delays. Similar delay effects were not found in other areas implicated in reward processing, including the ventral striatum and hippocampus. Connectivity analyses revealed that during RL feedback, cognitive areas of the cerebellum coactivate with a network that includes the medial and lateral prefrontal cortex and caudate nucleus. Together, these results highlight a temporally constrained contribution of the human cerebellum to a cognitive learning task.

Neural computations of visual, semantic, and memorability features in the human brain
Object recognition requires integrated processing that extends beyond the visual cortex, incorporating semantic and memory-related processes. However, it remains unclear how different attributes, such as visual, semantic, and memorability features, are encoded and interact during perception. Here, we recorded intracranial electroencephalography from 5143 channels while participants viewed natural object images. We systematically characterized the spatiotemporal patterns of neural encoding for visual, semantic, and memorability attributes and showed that memorability was encoded in a distributed manner, which can be dissociated from visual and semantic coding. While the ventral temporal cortex (VTC) was engaged in encoding all three attributes, the representations were dissociable. Interestingly, memorability representations in the prefrontal cortex appeared to arise from integrated visual and semantic signals from the VTC; and memorability influenced early stages of visual and semantic processing. Our results were corroborated by high-resolution 7T fMRI, which revealed continuous encoding across the brain, and further validated using a separate dataset featuring within-category object variability. Lastly, single-neuron recordings confirmed semantic and memorability coding in the medial temporal lobe. Together, these findings provide a comprehensive view of how visual, semantic, and memorability attributes are dynamically encoded across the brain, highlighting the complex interplay between these attributes that collectively shape object recognition and memory formation.

Language laterality and cognitive skills: does anatomy matter?
Brain anatomy, particularly white matter microstructure, is thought to play a critical role in the relationships between cognitive function and language lateralisation. This study investigates whether white matter microstructural parameters of the arcuate fasciculus and corpus callosum is associated with cognitive performance across distinct language lateralisation groups. Neuroimaging and cognitive data from 279 healthy adults were sourced from the BIL&GIN database. Participants completed a sentence production fMRI task, diffusion MRI, and cognitive tasks assessing verbal, visuospatial, and arithmetic skills. Significant positive associations were observed in strongly atypical individuals between fractional anisotropy in the splenium and working memory (R2 = 0.96, pFDR = 0.042) and between FA in the genu and visuospatial attention (R2 = 0.92, pFDR = 0.042). ANCOVA revealed that cross-dominant individuals had significantly lower visuospatial attention scores compared to consistently lateralised individuals (pFDR = 0.02). These findings challenge the notion that atypical lateralisation is inherently maladaptive and suggest that white matter pathways may serve as an alternative mechanism for supporting cognitive function in individuals with rightward language dominance. Furthermore, the results highlight the cognitive disadvantages of crossed dominance, implicating disrupted interhemispheric communication as a potential underlying mechanism.

Brain structure-function coupling - relationship with language lateralisation
Language is one of the most extensively studied lateralised cognitive functions in the human brain, predominantly relying on the left hemisphere in most individuals. However, the mechanisms by which a stable white matter architecture underpins individual language functions remain unclear. Previous studies have employed structural connectivity (SC) and functional connectivity (FC) coupling for individual fingerprinting and task decoding, suggesting that variability in brain entropy may serve as a distinguishing characteristic for language lateralisation. We examined a large cohort of healthy adults (n = 285) to investigate SC-FC coupling and identify markers distinguishing different language laterality groups. Functional connectivity was measured using resting-state fMRI (rsfMRI) time-series data, while structural connectivity was determined via probabilistic fibre tractography. SC-FC coupling was investigated using the SENSAAS language atlas and defined as the Pearson correlation between non-zero elements of regional structural and functional connectivity profiles. Group differences were assessed using the PALM toolbox in FSL. Our findings revealed that increased SC-FC coupling in the left precentral sulcus was associated with typical language lateralisation, while increased coupling in the right middle temporal gyrus and left anterior insula was observed in individuals with atypical language lateralisation (pFDR < 0.05). Non-lateralised individuals exhibited increased coupling in the left anterior insula compared to lateralised (pFDR<0.05). SC-FC coupling offers a promising framework to uncover functional and anatomical differences among individuals with varying language lateralisation. This regional specificity indicates that typical, atypical, and non-lateralised profiles rely on different structural-functional alignments, likely reflecting the recruitment of alternative pathways for language processing.

Critical point drying of brain tissue for X-ray phase contrast imaging
X-ray phase contrast tomography is emerging as a powerful method for imaging large volumes of brain tissue at sub-cellular resolution. However, current sample preparation methods are largely inherited from visible light or electron microscopy workflows and hence are not optimised to exploit the full potential of X-ray contrast mechanisms. Here we propose to replace interstitial material by air to enhance X-ray phase contrast of the ultrastructural features. We used critical point drying (CPD) of heavy metal-stained mouse brain tissue to produce mechanically stable samples with preserved ultrastructure and enhanced refractive index boundaries, a nanofoam-like material that remains compatible with follow-up conventional resin embedding. Using two complementary synchrotron-based setups, a high-throughput microtomography beamline (P14, DESY) and a nanoscale holographic tomography beamline (ID16A, ESRF), we found that CPD samples consistently showed 2-4x stronger phase-shift signal than conventional resin-embedded tissue. The contrast gain remained consistent across samples, imaging conditions, and beamlines. Our results suggest that CPD offers a generalisable route for preparing tissue for high-resolution X-ray imaging. It retains structural detail while improving signal, and is compatible with other processing procedures like femtosecond laser milling or electron microscopy, paving the path for large-scale biological tissue imaging.

Non-synaptic Mechanism of Ocular Dominance Plasticity
Classic experiments showing that monocular visual disruption alters synaptic connections to binocular neurons in the brain established the fundamental concept of synaptic plasticity through coincident spike time arrival. However, if the speed of impulse transmission from the eye is altered by visual deprivation, spike time arrival at binocular neurons would be affected, thereby inducing synaptic plasticity. This possibility is tested here in adult mice by monocular eyelid suture and action potential inhibition in retinal axons. The results show that spike time arrival in visual cortex is altered by monocular visual disruption in association with morphological changes in myelin (nodes of Ranvier) on axons in optic nerve and optic tract. This non-synaptic mechanism of ocular dominance plasticity, mediated by myelin-forming cells, supplements and may drive synaptic plasticity.

Preserved synaptic architecture but impaired ketamine-induced synaptic plasticity of layer 5 pyramidal neurons in the aged frontal cortex
Healthy aging is accompanied by a gradual decline in higher-order cognitive functions, including working memory, attention, and cognitive flexibility, processes that critically rely on intact frontal cortical circuits. While neuronal loss is minimal during aging, whether there are changes in functional plasticity in this region remains unexplored. In this regard, dendritic spines, the primary postsynaptic structures of excitatory synapses, act as key hubs for experience-dependent synaptic remodeling. Using longitudinal in vivo two-photon imaging in Thy1-eGFP-M mice, we examined age-related changes in dendritic spine density and dynamics in layer 5 pyramidal neurons of the secondary motor area (MOs), a frontal cortical region essential for strategy switching and cognitive flexibility, and that was assessed using an operant conditioning paradigm. We found that aged mice (18 to 22 months) exhibited significant impairments in cognitive flexibility relative to young mice (3 to 5 months) in the four-odor choice discrimination and reversal task. Analysis of dendritic spine plasticity revealed that baseline spine density, turnover, and morphology were largely preserved in aged mice. Sex differences were evident, with females displaying higher spine density and a greater fraction of stable spines, a feature maintained across aging. Importantly, despite preserved baseline architecture, aged mice showed impaired ketamine-induced spinogenesis and reduced stabilization of newly formed spines, in contrast to the robust structural plasticity observed in young mice. These results indicate that healthy aging selectively impairs activity-dependent synaptic remodeling without affecting steady-state spine architecture in frontal cortical circuits. By linking deficits in induced synaptic plasticity to age-related impairments in cognitive flexibility, our study highlights the critical need to target plasticity mechanisms as a therapeutic strategy to restore executive function and cognitive adaptability in the aging brain.

Conserved Axonal Transcriptome Dynamics Underlie PGE2-Induced Sensitisation and Identify Tnfrsf12a/Fn14 as a Regulator of Neuronal Excitability in DRG Neurons
Chronic pain arises when dorsal root ganglion (DRG) neurons become sensitised to noxious inputs, a process driven by inflammatory mediators such as prostaglandin E2 (PGE2). Local translation of axonal mRNAs is a key regulator of nociceptor plasticity, yet how axonal transcriptome dynamics contribute to inflammatory sensitisation remains unclear. Using compartmentalised culture systems and RNA-sequencing, we defined axonal and somatic transcriptomes in embryonic (E16.5) and adult (W8) DRG neurons and assessed their remodelling after PGE2 exposure. We identify a conserved core axonal transcriptome spanning embryonic to adult stages, prominently enriched for ribosomal and mitochondrial functions, consistent with sustained translational and metabolic demands. PGE2 elicited compartment-specific reprogramming: pathways related to sensory processing and pain were upregulated in axons but downregulated in somata. Functionally, prolonged axonal PGE2 exposure enhanced capsaicin-evoked calcium responses and drove retrograde sensitisation of neuronal somata. Integrating transcriptomics with functional assays, we pinpointed Tnfrsf12a (Fn14), a cytokine receptor linked to regeneration and neuropathic pain, as a PGE2-induced axonal mRNA. Crucially, local axonal knockdown of Tnfrsf12a significantly reduced neuronal excitability, providing proof-of-concept that axonally enriched transcripts can be targeted to modulate sensitisation. These findings position conserved axonal transcriptome programmes as drivers of peripheral sensitisation and establish Tnfrsf12a/Fn14 as a therapeutic candidate for inflammatory pain.

Benchmarking Orientation Distribution Function Estimation Methods for Tractometry in Single-Shell Diffusion Magnetic Resonance Imaging - An Evaluation of Test-Retest Reliability and Predictive Capability
Deriving white matter (WM) bundles in-vivo has thus far mainly been applied in research settings, leveraging high angular resolution, multi-shell diffusion MRI (dMRI) acquisitions that enable advanced reconstruction methods. However, these advanced acquisitions are both time-consuming and costly to acquire. The ability to reconstruct WM bundles in the massive amounts of existing single-shelled, lower angular resolution data from legacy research studies and healthcare systems would offer much broader clinical applications and population-level generalizability. While legacy scans may offer a valuable, large-scale complement to contemporary research datasets, the reliability of white matter bundles derived from these scans remains unclear. Here, we leverage a large research dataset where each 64-direction dMRI scan was acquired as two independent 32-direction runs per subject. To investigate how recently developed bundle segmentation methods generalize to this data, we evaluated the test-retest reliability of the two 32-direction scans, of WM bundle extraction across three orientation distribution function (ODF) reconstruction methods: generalized q-sampling imaging (GQI), constrained spherical deconvolution (CSD), and single-shell three-tissue CSD (SS3T). We found that the majority of WM bundles could be reliably extracted from dMRI scans that were acquired using the 32-direction, single-shell acquisition scheme. The mean dice coefficient of reconstructed WM bundles was consistently higher within-subject than between-subject for all WM bundles and ODF reconstruction methods, illustrating preservation of person-specific anatomy. Further, when using features of the bundles to predict complex reasoning assessed using a computerized cognitive battery, we observed stable prediction accuracies (r: 0.15-0.36) across the test-retest data. Among the three ODF reconstruction methods, SS3T had a good balance between sensitivity and specificity in external validation, a high intra-class correlation of extracted features, more plausible bundles, and strong predictive performance. More broadly, these results demonstrate that bundle segmentation can achieve robust performance even on lower angular resolution, single-shell dMRI, with particular advantages for ODF methods optimized for single-shell data. This highlights the considerable potential for dMRI collected in healthcare settings and legacy research datasets to accelerate and expand the scope of WM research.

Redefined Strategies to generate Conditional miR-141/200c miRNA cluster Knockout mice to eliminate confounding neo cassettes
MicroRNAs (miRNAs) of the miR-200 family specifically miR-141 and miR0-200c regulate neurogenesis, differentiation, and epithelial mesenchymal transitions in development and several diseases including cancer and stroke. The STOCK Mirc13tm1Mtm /Mmjax mouse line, which targets the miR-141/200c cluster, was originally described by Park et al. (2012) as a conditional knockout-first allele requiring a two-step breeding strategy: Flp recombination to excise lacZ/neo cassettes followed by Cre recombination to delete the floxed miRNA cluster. However, subsequent studies frequently bypassed this step and reported knockouts based on direct crosses with Cre mouse lines, leaving residual lacZ/neo sequences that may silence upstream elements or introduce transcriptional artifacts. Here, we present a detailed and refined strategy to generate both global and tissue-specific miR-141/200c knockouts that adheres to the original design intent and eliminates confounding cassettes. Our approach confirmed robust baseline expression of miR-141 and miR-200c in various organs such olfactory bulbs and lungs where these miRNAs are robustly expressed, with near-complete loss of expression in knockout animals as validated by qPCR and in situ hybridization. By restoring a clean floxed allele prior to Cre deletion, we establish a reliable and interpretable mouse model for dissecting the roles of the miR-141/200c cluster miRNA in various disease models in these mice.

Obesogenic Diet Impairs Social Memory Through Alterations of Hippocampal CA2 Excitability and Oxytocin Signaling
While obesity induces cardio-metabolic disorders and cognitive deficits, the underlying neural mechanisms remain unexplored. In mice, exposure to an obesogenic high-fat and sugar diet (HFD) resulted in social recognition memory deficits, a process that is dependent upon hippocampal area CA2 and oxytocin signaling. HFD-fed mice had stronger inputs onto CA2 pyramidal neurons that led to increased action potential firing, without altering intrinsic properties or inhibitory transmission. Chemogenetic CA2 inhibition rescued HFD-induced social memory deficits, confirming the role of CA2 hyperexcitability in these effects. In CA2, oxytocin receptor activation resulted in membrane depolarization, spontaneous action-potential firing and permitted endocannabinoid-mediated plasticity in control diet-fed mice, but not HFD-fed littermates. In a concentration-dependent manner, oxytocin restored potentiation of excitatory responses and allowed for endocannabinoid plasticity at CA2 inhibitory synapses as well as social memory deficits in HFD-fed mice. By investigating the influence of diet on hippocampal area CA2, this study uncovers novel mechanisms linking neuromodulation and plasticity in social memory encoding.

The Proximity Prediction Hypothesis: How predictive coding of CT-touch explains Autonomous Sensory Meridian Response and its therapeutic applications.
Autonomous Sensory Meridian Response (ASMR) is a pleasant tingling sensation felt across the scalp and neck, widely reported to reduce anxiety and improve sleep. The Proximity Prediction Hypothesis (PPH) is the first comprehensive predictive coding model explaining ASMR's underlying neural mechanism. PPH posits that near-field acoustic cues from common ASMR triggers (e.g., brushing sounds, whispered speech) engage the audio-tactile Peripersonal Space Network, generating a top-down prediction of gentle C-tactile (CT) touch on CT fibre-rich skin of the scalp and neck. This prediction suppresses locus coeruleus (LC) arousal and increases vagal output, offering a mechanistic explanation for the phenomenon's therapeutic benefits. In a subjective-experience survey (N = 64), ASMR-labelled trials were rated significantly more pleasant but only slightly more arousing than controls. Pleasantness predicted both the presence and intensity of tingles, supporting PPH's core claim that hedonic value, rather than sympathetic activation, drives the graded somatosensory response. PPH situates ASMR within the Neurovisceral Integration framework, predicting measurable Central Nervous System-Autonomic Nervous System (CNS-ANS) markers (beta-band desynchronisation in the posterior insula and proportional increases in high-frequency heart rate variability with tingle intensity). It further predicts reduced LC activity during ASMR, stronger effects in individuals with high interoceptive prediction error (e.g., anxiety, autism), and attenuation of tingles when spatial proximity cues are removed. By integrating auditory proximity, CT-touch anticipation, and autonomic regulation into a single predictive-coding account, PPH provides a unified, testable framework for explaining ASMR, offering a blueprint for translating this sensory phenomenon into targeted, evidence-based interventions for anxiety and sleep disorders.

Enhancing experience-dependent plasticity accelerates vision loss in a murine model of retinitis pigmentosa
Modulating neural plasticity is pursued as a therapeutic approach for several neurologic conditions. Here we evaluated if enhancing experience-dependent plasticity prolongs vision in a murine model of retinitis pigmentosa. First, we quantified the loss of visual acuity under both scotopic and photopic conditions for mice heterozygous for the P23H mutation in the Rhodopsin gene (Rho P23H/+). Acuity progressively declined under scotopic conditions followed by photopic conditions. Acuity deficits only broadly correlated with the retinal response measured by the electroretinogram. In contrast, acuity deficits were consistent with the percent of cortical excitatory layer 2/3 neurons responsive to higher spatial frequency visual stimuli. Then, we tested if enhancing plasticity in adult visual circuity by deleting the nogo-66 receptor gene (Ngr1) would preserve vision in Rho P23H/+ mice. However, loss of vision was accelerated in Ngr1 -/-; Rho P23H/+ mice. Thus, enhancing plasticity can be maladaptive in the context of neural degeneration.

Enduring Autism-like Phenotypes and Deregulated Hypothalamic Prosocial Peptides After Early-Life Exposure to Indoor Flame Retardants in Male C57BL/6 Mice
Background: Polybrominated diphenyl ethers (PBDEs) are neuroendocrine disrupting chemicals that produce adverse neurodevelopmental effects. PBDEs have been implicated as risk factors for autism spectrum disorder (ASD), which is characterized by abnormal psychosocial functioning and is commonly accompanied by co-morbidities such as cognitive and attentional deficits. Here, we used a mouse model with translationally relevant exposure to establish direct causal evidence that maternal transfer of a commercial mixture of PBDEs, DE-71, produces ASD-relevant behavioral and neurochemical deficits in male offspring. Methods: C57Bl6/N mouse dams were exposed to a commercial PBDE mixture, DE-71, via oral administration of 0 (vehicle control, VEH/CON), 0.1 (L-DE-71), or 0.4 (H-DE-71) mg/kg bw/d for 10 weeks, spanning three weeks prior to gestation through the end of lactation at postnatal day (PND) 21. Results: Mass spectrometric analysis indicated dose-dependent transfer of PBDEs (in ppb) to brains of F1 male offspring at PND 30, with reduction in levels by PND 110. Adult F1 male offspring displayed ASD-relevant neurobehavioral phenotypes, including impaired short- and long-term social recognition memory (SRM), despite intact general sociability, and exaggerated repetitive behavior. Exposed mice also displayed altered olfactory discrimination of social odors, impaired novel object recognition memory, and reduced open field habituation. However, no changes were observed in anxiety-like, sensorimotor, or depressive-like behaviors relative to VEH/CON. At the molecular level, DE-71 exposed males displayed deregulated gene markers of prosocial neuropeptides. Oxt was upregulated in the paraventricular nucleus (PVN); Avp was upregulated in the PVN and bed nucleus of the stria terminalis (BNST) but downregulated in the lateral septum (LS); Avp1ar and Adcyap1 were upregulated in the BNST; and Adcyap1r1 was upregulated in the PVN, supraoptic nucleus (SON), and BNST. Conclusions: These findings demonstrate that developmental PBDE exposure produces enduring behavioral and neurochemical phenotypes that resemble core domains of ASD, which may result from early neurodevelopmental reprogramming within central social and memory networks.

Neuroinflammation links the neurogenic and neurodegenerative phenotypes of Nrmt1-/- mice
It is widely thought that age-related damage is the single biggest contributing factor to neurodegenerative diseases. However, recent studies are beginning to indicate that many of these diseases may have developmental origins that become unmasked overtime. It has been difficult to prove these developmental origins, as there are still few known links between defective embryonic neurogenesis and progressive neurodegeneration. We have created a constitutive knockout mouse for the N-terminal methyltransferase NRMT1 (Nrmt1-/- mice). Nrmt1-/- mice display phenotypes associated with premature aging. Specifically in the brain, they exhibit age-related striatal and hippocampal degeneration, which is accompanied by impaired short and long-term memory. These phenotypes are preceded by depletion of the postnatal neural stem cell (NSC) pools, which appears to be driven by their premature differentiation and migration. However, this differentiation is often incomplete, as many resulting neurons cannot permanently exit the cell cycle and ultimately undergo apoptosis. Here, we show that the onset of apoptosis corresponds to increased cleavage of p35 into the CDK5 activator p25, which can promote neuroinflammation. Accordingly, Nrmt1-/- brains exhibit an increase in pro-inflammatory cytokine signaling, astrogliosis, complement activation, microgliosis, and markers of a compromised blood brain barrier, all of which indicate an activated neuroimmune response. We also find Nrmt1-/- mice do not activate a corresponding anti-inflammatory response. These data indicate that abnormal neurogenesis can trigger neuroinflammation, which in the absence of compensatory anti-inflammatory signaling, could lead to neuronal apoptosis and progressive neurodegeneration.

Ischemic stroke increases levels of the folate receptor and one-carbon enzymes in male and female brain tissue
Stroke is the second most common cause of death worldwide and predominantly affects individuals over 65 years old. Its prevalence is projected to increase in parallel with the aging global population. Nutrition is a modifiable risk factor for ischemic stroke. Folates, B-vitamins and choline play a central role in one-carbon metabolism (1C), which is a key metabolic network that integrates nutritional signals with biosynthesis, redox homeostasis, epigenetics, regulation of cell proliferation, and stress resistance. Our research group has previously shown that deficiencies in 1C lead to worsened stroke outcomes using preclinical models. However, the impact of ischemic stroke on 1C enzymes in affected brain tissue remains unknown. The objective of this study is to investigate whether ischemic stroke contributes to a change in the levels of 1C enzymes after ischemic stroke in male and female patients. Cortical brain tissue sections from ischemic stroke patients and controls were stained for enzymes involved in 1C. All tissue was co-stained with neuronal nuclei (NeuN) and DAPI (4',6-diamidino-2-phenylindole). The colocalization of all three markers was evaluated by two individuals who were blinded to the experimental groups. Ischemic stroke increased neuronal levels of the folate receptor and 1C enzymes, methylenetetrahydrofolate reductase (MTHFR), thymidylate synthase (TS) and serine hydroxy methyltransferase (SHMT). In male stroke brain tissue was observed to have increased levels of MTHFR, TS, and SHMT. Female brain tissue had increases in the folate receptor and TS. The results suggest that ischemic stroke leads to increased demand of 1C and that there are some differences between males and females.

Erucamide regulates retinal neurovascular crosstalk
Neurovasculoglial crosstalk is critical in establishing and maintaining a functional neurovascular unit. Breakdown in the unit is central to many neurodegenerative disorders of the CNS of which the retina is a component. A growing literature indicated that primary fatty acid amides (PFAMs) can regulate this crosstalk between vasculature and neuronal tissues. In this study we describe a central role for erucamide, a 22:1 mono-unsaturated omega-9 fatty acid amide, in degenerating retinal tissues. Using high-resolution global mass spectrometry-based metabolomics, we cataloged metabolites in murine models of retinal degeneration and show that while PFAMs, in general, are highly dysregulated, erucamide is the one most significantly diminished during photoreceptor atrophy. Using rodent models of retinal degeneration and novel organosilane-modified porous silicon nanoparticles (pSiNPs) for the in vivo delivery of erucamide, we demonstrate that erucamide activates CD11b+ myeloid cells, leading to the upregulation of angiogenic and neurotrophic cytokines that stabilize retinal degeneration. We identified TMEM19 as a novel binding protein for erucamide that is crucial for human iPSC-derived macrophage precursor cells activation and subsequent neurotrophic and angiogenic factor production. These findings reveal a previously unknown PFAM pathway that is modulated during retinal degenerative diseases, demonstrating that erucamide or functional analogues and their action through TMEM19 may be useful as a therapeutic alternative to neuroprotective and stem cell-based approaches for the treatment of retinal degenerative diseases.

Heterochronic myeloid cell replacement reveals the local brain environment as key driver of microglia aging
Aging, the key risk factor for cognitive decline, impacts the brain in a region-specific manner, with microglia among the most affected cell types. However, it remains unclear whether this is intrinsically mediated or driven by age-related changes in neighboring cells. Here, we describe a scalable, genetically modifiable system for in vivo heterochronic myeloid cell replacement. We find reconstituted myeloid cells adopt region-specific transcriptional, morphological and tiling profiles characteristic of resident microglia. Young donor cells in aged brains rapidly acquired aging phenotypes, particularly in the cerebellum, while old cells in young brains adopted youthful profiles. We identified STAT1-mediated signaling as one axis controlling microglia aging, as STAT1-loss prevented aging trajectories in reconstituted cells. Spatial transcriptomics combined with cell ablation models identified rare natural killer cells as necessary drivers of interferon signaling in aged microglia. These findings establish the local environment, rather than cell-autonomous programming, as a primary driver of microglia aging phenotypes.

Hypoimmunogenic human motor neurons induced from iPSCs in vivo substantially ameliorate ALS disease in large animal models
Stem cell-based therapy holds great potential for substituting degenerated motor neurons (MNs) in amyotrophic lateral sclerosis (ALS). Missing protocols for advanced differentiation of transplanted cells into MNs, immune rejection, and the lack of suitable ALS models for preclinical trials have slowed the development of effective therapies. Here, we employed multiplex genetic-editing to generate a novel human pluripotent stem cell line containing doxycycline (Dox)-inducible MNs-specific transcription factors and comprehensively modified immunomodulatory genes. We transplanted these cells into the spinal cord of ALS large animal models (SOD1G93A pigs and TIA1P362L rabbits), which faithfully recapitulate pathologies and symptoms observed in ALS patients. The transplanted cells could efficiently differentiate into functional MNs upon Dox treatment in vivo, distribute throughout the spinal cord and motor cortex via extensive migration, survive long-term without the need for immunosuppression. Notably, these MNs integrated into host neural circuits, as evidenced by their long projection of peripheral axons to target muscle and reformation of neuromuscular junctions. As result, pathologies and motor deficits were substantially ameliorated in both animal models.

Relative value learning in Drosophila melanogaster larvae
The ability to learn from past experiences to inform future decision-making is crucial for humans and animals alike. One question with important implications for adaptive decision-making is whether we learn about the absolute values of cues we encounter (how good or bad?), or about their relative values (how much better or worse than the alternative?). Humans have been shown to use relative value learning, even when it leads to suboptimal decisions. In this study, we ask whether insects use absolute or relative value learning. Using the larvae of the fruit fly Drosophila melanogaster, we designed associative odour-taste learning experiments to distinguish both kinds of learning and find that larvae learn about the relative rather than the absolute values of both rewards and punishments, irrespective of the number and sequence of training trials. This suggests that relative value learning is a facility shared across the animal kingdom from maggots to humans, and can be realized even by simple insect brains. Given the great potential of D. melanogaster as a model organism for in-depth neurobiological analyses, our study opens up the opportunity to reveal the mechanism underlying relative value learning in unprecedented detail.

Quantitative spectral Linear Unmixing and Ratiometric FRET for live-cell imaging of protein interactions
We present a biophysical imaging strategy based on linear unmixing Forster resonance energy transfer (lux-FRET) for investigating protein-protein interactions and receptor-mediated signaling in live cells. This method utilizes spectral unmixing of FRET signals acquired via confocal laser scanning microscopy (LSM), enabling high-resolution quantification of molecular interactions with both spatial and temporal precision. Applying lux-FRET, we examined receptor-receptor interactions and downstream signaling events, including agonist specificity for 5-HT receptors. Ratiometric FRET measurements with a genetically encoded cAMP biosensor allowed us to assess biosensor sensitivity to cyclic nucleotides and receptor efficacy. Additionally, we explored physiological interactions between CD44 and 5-HT receptors and characterized the oligomerization state of the 5-HT1A receptor through apparent FRET efficiency analysis. Our findings demonstrate the utility of lux-FRET combined with quantitative molecular microscopy as a powerful tool for dissecting dynamic signaling mechanisms in live cells. This approach offers broad applicability for researchers studying receptor pharmacology, cellular signaling, and protein interaction dynamics.

Using Tools as Cues for Motor Adaptation in Virtual Reality
Humans are adept at using multiple tools, often switching between them even when each involves distinct and potentially conflicting motor demands. This study investigated how different features of a tool influence the formation of distinct motor memories during dual adaptation to opposing visuomotor perturbations. Using an immersive virtual reality (VR) setup, participants performed an aiming task in which they launched a ball toward a target using virtual tools, each consistently associated with a different visuomotor rotation. We manipulated tool features across three groups: one in which tools differed only in colour (Colour Control), one in which they differed in shape but involved similar operational dynamics (Motor Congruent), and one in which they differed in both shape and mode of operation (Motor Incongruent). A control group that adapted to a single perturbation with a single tool at a time was also included. Only the Motor Incongruent group demonstrated robust dual adaptation and clear aftereffects, comparable to those observed during single-tool learning. These results suggest that distinct modes of tool operation play a critical role in supporting the formation and retention of separate internal models during sensorimotor adaptation.

Deciphering the mechanistic basis for the pathological effect of the Gαo E246K mutation in neurodevelopmental disorder
Mutations in the GNAO1 gene, which encodes for Go, a major neuronal G protein, are associated with neurodevelopmental disorders, epilepsy, and movement disorders. We identified and characterized a spontaneous heterozygous GNAO1 E246K mutation in an Israeli female infant with complex developmental delays and substantial motor difficulties. This mutation has been reported in other cases as a prevalent pathogenic mutation in patients with motor dysfunction and a broad range of neurological outcomes. To investigate the molecular and functional consequences of the Go E246K mutation, we employed structural modeling and analysis, biochemical assays, mass spectrometry-based proteomics, and cellular functional assays. We show that this mutation does not affect nucleotide binding, nor basal or RGS-accelerated GTP hydrolysis. Despite the E246 position located within a predicted effector binding region, proteomics analysis did not identify any new cellular partners. Instead, we demonstrate that the E246K mutation disrupts the Go regulatory GTPase cycle by directly impairing G{beta}{gamma} dissociation. This impairment overrides the presence of wild-type Go, explaining the dominant effect of the severe neurogenetic phenotype in the heterozygous background. These findings establish a new molecular mechanism for a GNAO1 mutation with dominant-negative effects on the GTPase regulatory cycle. The insights gained from studying this mechanism of action provide a basis for developing specific and personalized therapeutic strategies based on the outcome of a missense mutation in GNAO1.

Mitochondrial dysfunction precedes neurodegeneration in DRPLA patient-derived neurons, and phenylbutyrate improves survival.
Dentatorubral-pallidoluysian atrophy (DRPLA) is a progressive autosomal-dominant neurodegenerative disorder caused by a CAG repeat expansion in the ATN1 gene, which encodes the transcriptional corepressor Atrophin-1. The temporal sequence of molecular mechanisms driving neuronal dysfunction and degeneration in DRPLA is poorly understood, limiting therapeutic development. We generated patient-derived induced pluripotent stem cells and differentiated them into cortical excitatory glutamatergic neurons to model early pathogenic processes. Early pathological features of DRPLA patient-derived neurons included mitochondrial dysfunction and oxidative stress. These alterations occurred before overt neuronal loss, highlighting bioenergetic stress as a key early driver of disease progression toward neurodegeneration. Pharmacological treatment with phenylbutyrate significantly improved neuronal survival and reduced mitochondrial reactive oxygen species production, demonstrating the therapeutic potential of targeting mitochondrial dysfunction and oxidative stress. These findings challenge the conventional aggregation-centric model of polyglutamine disease pathogenesis and position mitochondrial stress as a central and early promoter of neuronal degeneration in DRPLA. By providing mechanistic insight into early-stage disease processes, our study lays the foundation for therapeutic strategies targeting mitochondrial dysfunction in DRPLA and related polyglutamine disorders.

Activation of transposable elements is linked to a region- and cell-type-specific interferon response in Parkinson's disease
Parkinson's disease (PD) is a common age-related neurodegenerative disorder involving a neuroinflammatory response, the cause of which remains unclear. Transposable elements (TE) have been linked to inflammatory states, but their potential role in PD has not been explored. Using bulk- and single nuclei RNAseq of postmortem brain tissue from four brain regions, we studied TE transcriptional activation and its correlation with neuroinflammation in PD. Over a thousand TE loci, including LINE-1s and ERVs, were highly expressed in a cell-type and region-specific manner in the human brain. Increased TE expression was found in microglia and neurons in the substantia nigra and putamen in the PD brains, but not amygdala or prefrontal cortex, compared to age-matched control tissue. This TE activation correlated with innate immune transcriptional responses, characterized by the expression of interferon-related and viral response genes, in the same brain regions. The link between an interferon response and TE activation was mechanistically confirmed using human pluripotent stem cell-derived microglia and neurons. Our findings provide a unique insight into TE transcription in the PD brain and suggests that TEs play a role in chronic neuroinflammatory processes and the progression of this neurodegenerative disorder.

Poly(ADP-ribose) Polymerase 1 Deficiency Attenuates Amyloid Pathology, Neurodegeneration, and Cognitive Decline in a Familial Alzheimer Disease Model
Poly(ADP-ribose) (PAR) polymerase-1 (PARP1) has been implicated in DNA damage responses and neuroinflammation in Alzheimer disease (AD), yet its role in amyloid-{beta} (A{beta}) pathology remains unclear. Here, we show that PARP1 activation drives A{beta} pathology and neurodegeneration. Using a sensitive ELISA, we observed significantly elevated PAR levels in the cerebrospinal fluid (CSF) of patients with mild cognitive impairment (MCI) and AD compared to controls. In vitro, oligomeric A{beta} 1-42 activated PARP1 and induced DNA damage, while genetic or pharmacological inhibition of PARP1 conferred neuroprotection. In vivo, PARP1 knockout in the 5XFAD mouse model of amyloidosis led to reduced amyloid plaque burden, preserved synaptic and neuronal integrity, attenuated glial activation and neuroinflammation, and rescued cognitive deficits. Mechanistically, PARP1 deficiency decreased amyloid precursor protein (APP) and BACE1 levels, altered {gamma}-secretase complex composition, and enhanced A{beta} degradation via neprilysin. These findings position PARP1 as a critical mediator of A{beta} toxicity and neurodegeneration, suggesting its inhibition as a promising therapeutic strategy for AD.

Disrupted Lipid Homeostasis as a Pathogenic Mechanism in ABCA7-Associated Alzheimers Disease Risk
INTRODUCTION: ABCA7 (ATP-binding cassette sub-family A member 7) encodes a lipid transporter linked to Alzheimers disease (AD). While common variants confer modest risk in Europeans, a 44-base pair deletion (rs142076058; p.Arg578Alafs) is a strong risk factor in African Americans (AA). Despite this, the biological consequences of this ancestry-specific variant are not well understood. METHODS: We expressed the truncated ABCA7 protein in HEK and HepG2 cells to assess localization and lipid metabolism. Additionally, induced pluripotent stem cell (iPSC)-derived neurons carrying the deletion were compared with isogenic controls. RESULTS: The truncated ABCA7 localized to the plasma membrane similarly to wild type but induced significant lipid droplet accumulation in HepG2 cells and iPSC-derived neurons. DISCUSSION: These findings show that the AA-specific ABCA7 deletion disrupts lipid regulation despite normal localization, suggesting a mechanistic link between impaired lipid homeostasis and increased AD risk. This work underscores the importance of ancestry-specific studies in AD research.