bioRxiv Subject Collection: Neuroscience's Journal

Mechanistic Variability in Corneal Nerve Recovery Linked to Injury Type
The cornea, a transparent tissue covering the eye, is essential for clear vision and represents the most densely innervated tissue in the body. Its extensive sensory innervation provides both sensory perception and crucial trophic support, maintaining corneal health and integrity. Disruption of corneal innervation leads to neurotrophic keratitis (NK), a pathological condition caused by ocular injury, surgical procedures, or underlying diseases. The limited understanding of NK pathophysiological mechanisms has hindered the development of innovative therapeutic approaches. In this study, we comparatively analyzed corneal innervation morphogenesis and regeneration across two clinically relevant injury models, highlighting both commonalities and differences among these contexts. Our results demonstrate that corneal nerve morphogenesis and maturation span approximately 13 weeks, from embryonic day 12 (E12) through three months of age. Additionally, we observed that the specification of distinct nerve fiber types coincided temporally with a significant enhancement in corneal sensitivity. Furthermore, we found that the type of innervation loss, either via axotomy or abrasion, differentially affected corneal sensitivity and epithelial cell homeostasis. Importantly, the regeneration mechanisms following injury were also distinctly dependent on the type of nerve damage sustained. Collectively, these findings underscore both the shared characteristics and unique aspects inherent in each NK model, highlighting the necessity for tailored therapeutic strategies specific to individual patterns of corneal innervation disruption.

Replicating the facilitatory effects of transcranial random noise stimulation on motion processing: A registered report
Non-invasive brain stimulation (NIBS) techniques have the potential to demonstrate the causal impact of targeted brain regions on specific behaviors, and to regulate or facilitate behavior in clinical applications. Transcranial random noise stimulation (tRNS) is one form of transcranial electric stimulation (tES) in which an alternating current is passed between electrodes at random frequencies. High-frequency tRNS (hf-tRNS) is thought to enhance excitability and has been reported to have facilitatory effects on behavior in healthy and clinical populations. Due to the potential application of tRNS, clear demonstrations of the efficacy and replicability of stimulation are critical. Here, we focused on replicating the facilitatory effect of hf-tRNS over the human middle temporal complex (hMT+) on contralateral motion processing, initially demonstrated by Ghin et al. (2018). In this prior study, the improvement in performance was specific to global motion processing in the visual field contralateral to stimulation. The motivation to replicate this effect was reinforced by the well-supported hypothesis that hMT+ is critical for contralateral global motion processing. However, our results indicated that hf-tRNS does not improve motion discrimination. Specifically, we were unable to replicate a contralateral global motion processing facilitation following hf-tRNS to hMT+. In our within-subject controls, we also found no difference between hf-tRNS to hMT+ on contralateral global motion processing in comparison to sham stimulation, or in comparison to hf-tRNS to the forehead. While it remains possible that our lack of replication could be due to minor changes in the protocol from the original Ghin et al., study, for hf-tRNS to become a widely applied method, the modulatory effect of hf-tRNS should be robust to slight adjustments to the procedure.

Neural signatures of recollection are sensitive to memory quality and specific event features
Episodic memories reflect a bound representation of multimodal features that can be recollected with varying levels of precision. Recent fMRI investigations have demonstrated that the precision and content of information retrieved from memory engage a network of posterior medial temporal and parietal regions co-activated with the hippocampus. Yet, comparatively little is known about how memory content and precision affect common neural signatures of memory captured by electroencephalography (EEG), where recollection has been associated with changes in event-related potential (ERP) and oscillatory measures of neural activity. Here, we used a multi-feature paradigm previously reported in Cooper & Ritchey (2019) with continuous measures of memory, in conjunction with scalp EEG, to characterize the content and quality of information that drives ERP and oscillatory markers of episodic memory. A common signature of memory retrieval in left posterior regions, called the late positive component (LPC), was sensitive to overall memory quality and also to precision of recollection for spatial features. Analysis of oscillatory markers during recollection revealed that alpha/beta desynchronization was modulated by overall memory quality and also by individual features in memory. Importantly, we found evidence of a relationship between these two neural markers of memory retrieval, suggesting that they may represent complementary aspects of the recollection experience. These findings demonstrate how time-sensitive and dynamic processes identified with EEG correspond to overall episodic recollection, and also to the retrieval of precise features in memory.

Mitofusin 2 controls mitochondrial and synaptic dynamics of suprachiasmatic VIP neurons and related circadian rhythms including sleep
Sustaining the strong rhythmic interactions between cellular adaptations and environmental cues has been posited as essential for preserving the physiological and behavioral alignment of an organism to the proper phase of the daily light/dark cycle. Here, we show that mitochondria and synaptic input organization of suprachiasmatic (SCN) vasoactive intestinal peptide (VIP)-expressing neurons show circadian rhythmicity. Perturbed mitochondrial dynamics achieved by conditional ablation of the fusogenic protein mitofusin 2 (Mfn2) in VIP neurons cause disrupted circadian oscillation in mitochondria and synapses in SCN VIP neurons leading to desynchronization of entrainment to the light/dark cycle in Mfn2 deficient mice that resulted in advanced phase angle of their locomotor activity onset, alterations in core body temperature and sleep-wake amount and architecture. Our data provide direct evidence of circadian SCN clock machinery dependence on high-performance Mfn2-regulated mitochondrial dynamics in VIP neurons for maintaining the coherence in daily biological rhythms of the mammalian organism.

Axonal distribution of Layers IV-VI Connections in Human Visual Cortex Revealed by in-vitro extracellular injections of Biocytin.
The organization of interlaminar axonal connections was studied in slices of adult human visual association cortex. Cortical tissue was obtained during surgical glioma treatment in three subjects. Slices were injected with the highly sensitive anterograde tracer biocytin. In some cases, tract tracing was combined with laminar recordings of electrically evoked potentials, employing current-source density analysis. This study is based on 23 injections placed at various cortical layers and analyzed in detail. The results reveal a striking columnar architecture of local axons organized in vertically oriented bundles as well as vertical columns of neuronal cells bodies. A high degree of specificity was observed in the connectional patterns of various layers. This specificity was reflected in the tendency of lateral connections to spread into different cortical layers, and in the large variations in the axonal bouton densities in different layers. More specifically. Layer IV injections resulted in a more circular projection pattern including massive projections into layer II-III. Layer V injections produced extensive intra-laminar lateral connections while layer VI injections resulted in a vertical projection leading up to layer IV and a lateral intra-laminar projection. Thus, our results reveal highly specific connectional patterns in the different cortical layers in adult human association visual cortex.

Widely used CaMKII regulatory segment mutations cause tight actinin binding and dendritic spine enlargement in unstimulated neurons
Ca2+/calmodulin-dependent protein kinase II (CaMKII) is essential for long-term potentiation (LTP) of excitatory synapses that underlies learning. CaMKII responds to Ca2+ influx into postsynaptic spines by phosphorylating proteins and forming new protein interactions. The relative importance of these enzymatic and structural functions is debated. LTP induction triggers CaMKII docking to NMDA receptors, and recent evidence suggests that LTP can proceed without kinase activity after this event. Furthermore, CaMKII interaction with -actinin-2 is required for dendritic spine enlargement following LTP induction. CaMKII can auto-phosphorylate at T286, which enables autonomous activity after Ca2+/CaM dissociation. CaMKII also bears threonine at positions 305 and 306 in its regulatory segment. Experiments with CaMKII variants including a T305A/T306A (AA) double substitution have led to a model whereby T305/T306 phosphorylation by autonomously active CaMKII prevents further Ca2+/CaM activation. However, this mechanism is not compatible with some existing data including CaMKII phospho-proteomics and measurements with reporters of CaMKII conformation. Furthermore, autonomous CaMKII activity is now thought to only endure for seconds after LTP induction. In this study, we show that the AA substitution has an unintended gain-of-function property - it enables tight binding to -actinin-2 in unstimulated neurons. In situ labelling shows that the AA substitution elevates CaMKII-actinin interactions in neurons to a level only normally observed after induction of LTP. The AA CaMKII variant also elevates the proportion of enlarged spines in unstimulated neurons without altering synaptic currents. Calorimetric measurements with purified protein confirm that -actinin-2 binds tightly to the AA variant of CaMKII with no requirement for kinase activation. Using x-ray crystallography, we show that the AA substitution enables -actinin-2 to adopt a different tighter binding mode. Our findings reinforce the notion that CaMKII primarily fulfils a structural role in LTP.

Slow scission of single synaptic vesicles by Dynamin at physiological temperature
Compensatory endocytosis is crucial for the presynapse to maintain a functional pool of fusion-competent vesicles. Slow Clathrin-mediated endocytosis has been widely regarded as the primary mechanism of retrieval but this has been challenged by competing Clathrin-independent endocytic models, most notably sub-second ultra-fast endocytosis, reported to be predominant at physiological temperature. Here, we sought to resolve the salience of the respective endocytic modes by using a purely presynaptic preparation, the Xenapse, amenable to total internal reflection fluorescence microscopy (TIRFM). While the role of Clathrin is in dispute, Dynamin is widely acknowledged to figure as the scission protein at the invaginated vesicle neck. Hence, we labelled the endogenous Dynamin I with EGFP by CRISPR-Cas9 techniques and visualized single synaptic Dynamin-mediated scission events at very high temporal resolution. This revealed only a single slow mode of Dynamin-dependent retrieval with a half time of ~ 9 seconds at physiological temperature. Cross-correlational analysis with fluorescently labelled Clathrin confirmed these Dynamin events to be Clathrin-dependent. We thereby affirm Clathrin-mediated endocytosis as the primary mode of compensatory retrieval.

Lower GABA levels in the posterior cingulate are linked with poorer episodic memory in healthy older adults
Age-related deficits in episodic memory and mnemonic discrimination are associated with an increased risk of neurodegenerative diseases, such as Alzheimer's disease (AD) (Stark et al., 2013). While much research has focused on hippocampal contributions to these age-related changes (Stark et al., 2019), less is known about the role of posterior cingulate cortex (PCC) especially reduced inhibition in episodic memory deficit. PCC has connections to the medial temporal lobe and is linked to memory declines (Greicius et al., 2004). It is also one of the most vulnerable regions to amyloid deposition in AD (Yokoi et al., 2018). This study hypothesized and found that age-related declines in GABAergic function (brain's major inhibitory neurotransmitter) within the PCC contributes to individual differences in memory performance in healthy older adults. Using Magnetic Resonance Spectroscopy, we measured GABA levels in the PCC in 22 healthy younger and 30 older adults. We assessed episodic memory using Rey Delayed Auditory Verbal Learning Test (RAVLT) and Mnemonic Similarity Task (MST). We found that both raw GABA levels and episodic memory performance are lower in older adults compared to young. This reduction in GABA levels is subserved by age-related changes in tissue-composition as evidenced by no age-group differences in corrected GABA levels. More importantly, lower GABA levels (independent of tissue-correction) were associated with poorer episodic performance including delayed recall and mnemonic discrimination. This research suggests that therapeutically targeting posterior cingulate GABA levels might help slow or alleviate memory decline.

Electrophysiological classification of CACNA1G gene variants associated with neurodevelopmental and neurological disorders
This study highlights the complementarity of automated patch-clamp (APC) and manual patch-clamp (MPC) approaches to describe the electrophysiological properties of eighteen Cav3.1 calcium channel variants associated with various neurological conditions. Current density was measured efficiently for all variants in APC experiments, with four variants (p.V184G, p.N1200S, p.S1263A and p.D2242N) showing high current densities, compared to wild-type Cav3.1 channel, while six variants (p.M197R, p.V392M, p.F956del, p.I962N, p.I1412T, and p.G1534D) displayed low current densities, and were therefore preferentially studied using MPC. The electrophysiological properties were well conserved in APC (e.g. inactivation and deactivation kinetics, steady-state properties), with only the APC-MPC correlation for the activation kinetics being less robust. In addition, neuronal modeling, using a deep cerebellar neuron (DCN) environment, revealed that most of the variants localized in the intracellular gate (S5 and S6 segments) could increase DCN spike frequencies. This DCN firing was critically dependent on the current density and further pointed to the gain-of-function (GOF) properties of p.A961T and p.M1531V, the recurrent variants associated with Spinocerebellar Ataxia type-42 with Neurodevelopmental Deficit (SCA42ND). Action-potential (AP) clamp experiments performed using cerebellar and thalamic neuron activities further established the GOF properties of p.A961T and p.M1531V variants. Overall, this study demonstrates that APC is well-suited to high-throughput analysis of Cav3.1 channel variants, and that MPC complements APC for characterizing low-expression variants. Furthermore, in silico modeling and AP clamp experiments establish that the gain- or loss-of-function properties of the variants are determined by how the Cav3.1 channel decodes the electrophysiological context of a neuron.

Proteomic Analysis Links Truncated Tau to Lysosome Motility, Autophagy and Endo-lysosomal Dysfunction
Tauopathies are characterized by the progressive accumulation of abnormal tau species, which disrupt the autophagy-lysosomal pathway (ALP), a critical system for degrading intracellular macromolecules and aggregated proteins, causing toxicity and cell death. This study investigates the impact of the N-terminally truncated Tau35 protein overexpression on proteolytic pathways, including effects on autophagy and endo-lysosomal processes. Using a Tau35 mouse model and SH-SY5Y cell lines stably expressing either the Tau35 fragment or full-length tau, we employed western blotting, proteomic analysis of lysosome-enriched brain fractions, proteolysis/endocytosis assays, and live-cell imaging with the lysotracker reporter to assess protein degradation and lysosomal function. Our findings identify early pathological changes in endo-lysosomal processes, including increased endocytosis, proteolytic dysfunction and lysosomal motility abnormalities, associated with Tau35-induced toxicity. This work extends previous research by providing new insights into the mechanisms of Tau35-induced neurotoxicity, offering a foundation for developing targeted therapeutic strategies to address tauopathies.

Optimal Neuromuscular Performance Requires Motor Neuron Phosphagen Kinases
Phosphagen systems are crucial for muscle bioenergetics - rapidly regenerating ATP to support the high metabolic demands of intense musculoskeletal activity. However, their roles in motor neurons that drive muscle contraction have received little attention. Here, we knocked down expression of the primary phosphagen kinase [Arginine Kinase 1; ArgK1] in Drosophila larval motor neurons and assessed the impact on presynaptic energy metabolism and neurotransmission in situ. Fluorescent metabolic probes showed a deficit in presynaptic energy metabolism and some glycolytic compensation. Glycolytic compensation was revealed through a faster elevation in lactate at high firing frequencies, and the accumulation of pyruvate subsequent to firing. Our performance assays included two tests of endurance: enforced cycles of presynaptic calcium pumping, and, separately, enforced body-wall contractions for extended periods. Neither test of endurance revealed deficits when ArgK1 was knocked down. The only performance deficits were detected at firing frequencies that approached, or exceeded, twice the firing frequencies recorded during fictive locomotion, where both electrophysiology and SynaptopHluorin imaging showed an inability to sustain neurotransmitter release. Our computational modeling of presynaptic bioenergetics indicates that the phosphagen systems contribution to motor neuron performance is likely through the removal of ADP in microdomains close to sites of ATP hydrolysis, rather than the provision of a deeper reservoir of ATP. Taken together, these data demonstrate that, as in muscle fibers, motor neurons rely on phosphagen systems during activity that imposes intense energetic demands.

How the Heart Shapes the Mind: The Role of Cardiac Interoception in the Interaction between Autonomic Nervous Activity and Self-related Thoughts
Our thoughts often drift away from the tasks at hand. Various factors influence this phenomenon, including changes in the external environment, individual cognitive characteristics, and fluctuations in bodily responses. This study investigated the relationship between autonomic nervous fluctuations and thought state transitions, focusing on individual differences such as cardiac interoception. First, the heartbeat counting task was conducted, and the difference between the reported and actual number of heartbeats was used as an index of interoceptive accuracy. The participants then completed an auditory attention task while their cardiac activities were monitored. During the task, thought probes were randomly presented, and participants selected their thought content from eight categories and rated aspects such as task concentration and arousal. We estimated trial-by-trial thought states in a data-driven manner and examined how the current thought state, autonomic nervous activity, and individual cardiac interoceptive accuracy influenced the thought state in the next trial. The results demonstrated a strong association between higher cardiac interoceptive accuracy and the maintenance of similar states in subsequent trials when accelerated heart rates occurred during self-related thought states. Furthermore, the participants with higher depressive tendencies and interoceptive accuracy exhibited an increased likelihood of transitioning to self-related states when experiencing decreased heart rate during task-concentrated states. These results suggest that accurately detecting heart rate changes associated with specific thought states facilitates updates in first-person conscious experience, thereby biasing the transition patterns of subsequent thought states. This study provides new insights into the cognitive and physiological mechanisms underlying the dynamics of spontaneous thought.

Intrinsic fluctuations in global connectivity reflect transitions between states of high and low prediction error
While numerous researchers claim that the minimization of prediction error (PE) is a general force underlying most brain functions, others argue instead that PE minimization drives low-level, sensory-related neuronal computations but not high-order, abstract cognitive operations. We investigated this issue using behavioral, fMRI, and EEG data. Studies 1A/1B examined semantic- and reward-processing PE using task-fMRI, yielding converging evidence of PE's global effects on large-scale connectivity: high-PE states broadly upregulated ventral-dorsal connectivity, and low-PE states upregulated posterior-anterior connectivity. Investigating whether these global patterns characterize cognition generally, Studies 2A/2B used resting-state fMRI and showed that individuals continuously fluctuate between ventral-dorsal (high-PE) and posterior-anterior (low-PE) dynamic connectivity states. Additionally, individual differences in PE task responses track differences in resting-state fluctuations, further endorsing that these fluctuations represent PE minimization at rest. Finally, Study 3 used combined fMRI-EEG and showed that these resting-state ventral-dorsal and posterior-anterior PE networks specifically oscillate at 3-6 Hz. This whole-brain layout and timeline together speak to high/low-PE fluctuations playing a role in integrative and general sub-second cognitive operations.

Functional and structural profiling of circulation via genetically encoded modular fluorescent probes.
Sustained labeling of fluids is crucial for their investigation in animal models. Here, we introduce a mouse line (Alb-mSc-ST), where blood and interstitial fluid are labeled with the red fluorescent protein mScarlet and SpyTag. The SpyTag-SpyCatcher technology is exploited to monitor circulating fluid properties by biosensors or detect blood-brain barrier disruption. This approach represents a valuable tool for studying vascular structure, permeability and microenvironment in body organs in vivo.

Genetic targeting of myelinated primary afferent neurons using a new NefhCreERT2 knock-in mouse
Primary afferent neurons that convey somatosensory modalities comprise two large, heterogeneous populations: small-diameter neurons that give rise to slowly conducting unmyelinated axonal C fibers and medium-to-large diameter neurons with fast myelinated A fibers. Despite these two major groupings, tools to differentiate between unmyelinated and myelinated primary afferent fibers by genetic targeting have not been available; in particular, whereas numerous mouse driver lines exist to target different C fiber populations, genetic tools that target myelinated primary afferent populations are scarce. Here we describe a knock-in mouse line expressing tamoxifen-dependent CreERT2 under control of the Nefh gene, which encodes neurofilament heavy chain (NFH or NF200), a protein that is highly enriched in myelinated fibers. This mouse enables highly selective and efficient recombination of Cre-dependent reporters for functional and anatomical interrogation of myelinated fibers while excluding unmyelinated C fibers. In combination with other recombinase-expressing mouse lines, this genetic tool will be valuable for intersectional targeting of subpopulations of myelinated primary afferent fibers.

Medial temporal lobe encodes cognitive maps of real-world social networks
People routinely navigate their complex social networks[1]: From gossiping strategically with others[2-4] to brokering connections between siloed groups[5,6], our ability to make adaptive social choices hinges on whether we can construct useful mental representations of the social ties within our communities[7]. While decades of neuroscience research have shown that the medial temporal lobe encodes cognitive maps of physical[8-10] or conceptual space[11], how the brain represents our social networks in the wild to solve social problems remains unknown. By combining computational models with functional neuroimaging and longitudinal measurement of an evolving and densely interconnected real-world human network (N=187), we show that the entorhinal cortex encodes a cognitive map of the long-range connectivity between pairs of network members. This social map reflects the particular demands of social navigation and is specifically formatted to encode the simultaneous connectivity between network members, which critically enables tracking how information diffuses across the network. Moreover, the strength of its encoding in the entorhinal cortex aids in brokering connections that improve cohesion within people's social communities. Our results illuminate how a domain-general neural mechanism[12,13] is tailored to prioritize the natural dynamics of social phenomena in order to support adaptive navigation through these highly complex environments.

A corticostriatal learning mechanism linking excess striatal dopamine and auditory hallucinations
Auditory hallucinations are linked to elevated striatal dopamine, but their underlying computational mechanisms have been obscured by regional heterogeneity in striatal dopamine signaling. To address this, we developed a normative circuit model in which corticostriatal plasticity in the ventral striatum is modulated by reward prediction errors to drive reinforcement learning while that in the sensory-dorsal striatum is modulated by sensory prediction errors derived from internal belief to drive self-supervised learning. We then validate the key predictions of this model using dopamine recordings across striatal regions in mice, as well as human behavior in a hybrid learning task. Finally, we find that changes in learning resulting from optogenetic stimulation of the sensory striatum in mice and individual variability in hallucination proneness in humans are best explained by selectively enhancing dopamine levels in the model sensory striatum. These findings identify plasticity mechanisms underlying biased learning of sensory expectations as a biologically plausible link between excess dopamine and hallucinations.

Overexpression of alpha synuclein in Midbrain Dopamine Neurons Reduces Dopamine Release Without Cell Loss and Drives Mild Motor Deficits in Mice
It has proven challenging to faithfully recapitulate the key pathological, physiological, and behavioral features of Parkinson Disease (PD) in animals. Here we used adeno-associated virus (AAV) vectors to achieve cell type-specific overexpression of wild-type human -synuclein (syn) and a fluorophore (mCherry) in midbrain dopamine neurons to model PD in mice. We found that AAVs drove selective expression of both syn and mCherry in midbrain dopamine neurons. In conjunction with approximately 2-fold overexpression of syn, we found several histopathological markers of PD-like pathology, including progressive accumulation of phosphorylated and aggregated syn, ubiquitin, and a reduction in the expression of tyrosine hydroxylase, without overt cell loss. In parallel, syn overexpression drove a profound loss of evoked dopamine release, without a substantive change in the intrinsic properties of dopamine neurons, nor in striatal dopamine content. Finally, syn overexpression led to mild locomotor deficits. Together, these findings suggest that moderate syn overexpression can mimic some aspects of premotor and early symptomatic phases of PD, including markers of Lewy Body-like pathology and functional loss of evoked dopamine release. This model may be useful for investigating cellular and circuit mechanisms related to PD pathogenesis and progression.

Catecholamine Precursor Modulation of Human Exploration:Evidence From a Large Gender-Balanced Sample
The catecholamine precursor tyrosine has been linked to improved cognitive performance, but investigations into decision-making and reinforcement learning processes known to be under catecholamine control are sparse. We examined the impact of a single dose of Tyrosine (2g) on reinforcement learning and exploration in a large (n=63) gender-balanced sample in a within-subjects preregistered study. Reinforcement learning performance was improved under Tyrosine, and computational modeling revealed that this performance increase was due to a stabilization of choice behavior reflected in increased value-driven exploitation. Further non-preregistered modeling analyses confirmed that accounting for higher-order perseveration substantially improved model fit, and substantiated the observation of increased value-driven exploitation under Tyrosine. Furthermore, it revealed a more fine-grained computational impact of Tyrosine, showing attenuated effects of directed exploration and value-independent perseveration. Supplementation with Tyrosine therefore improved reinforcement learning performance by stabilizing choice patterns in the service of optimizing reward accumulation. Results confirm that Tyrosine supplementation modulates specific computational mechanisms thought to be under catecholamine control.

KCa3.1 Contributes to Neuroinflammation and Nigral Dopaminergic Neurodegeneration in Experimental models of Parkinson's Disease
Chronic neuroinflammation and misfolded -synuclein (Syn) have been identified as key pathological correlates driving Parkinsons disease (PD) pathogenesis; however, the contribution of ion channels to microglia activation in the context of -synucleinopathy remains elusive. Herein, we show that KCa3.1, a calcium-activated potassium channel, is robustly upregulated within microglia in multiple preclinical models of PD and, most importantly, in human PD and dementia with Lewy bodies (DLB) brains. Pharmacological inhibition of KCa3.1 via senicapoc or TRAM-34 inhibits KCa3.1 channel activity and the associated reactive microglial phenotype in response to aggregated Syn, as well as ameliorates of PD like pathology in diverse PD mouse models. Additionally, proteomic and transcriptomic profiling of microglia revealed that senicapoc ameliorates aggregated Syn-induced, inflammation-associated pathways and dysregulated metabolism in primary microglial cells. Mechanistically, FYN kinase in a STAT1 dependent manner regulates KCa3.1 mediated the microglial reactive activation phenotype after -synucleinopathy. Moreover, reduced neuroinflammation and subsequent PD-like neuropathology were observed in SYN AAV inoculated KCa3.1 knockout mice. Together, these findings suggest that KCa3.1 inhibition represents a novel therapeutic strategy for treating patients with PD and related -synucleinopathies.

T1-weighted fMRI in mouse visual cortex using an Iron Oxide Nanoparticle contrast agent and Ultrashort Echo Time (UTE) imaging at 9.4 T
Purpose: This study aims to investigate the feasibility of using T1-weighted fMRI with an iron oxide nanoparticle contrast agent and Ultrashort Echo Time (UTE) imaging at 9.4T to measure functional hyperaemia in the mouse visual cortex. The goal is to capture positive signal changes in both the parenchyma and pial surface, and to test whether surface vessels respond during neuronal activation. Methods: The study involved scanning of nine mice after administration of iron oxide-based superparamagnetic contrast agent (Molday ION) via the tail vein. Two functional imaging experiments were conducted: one to investigate the effect of echo time on the functional response, and another to characterize the impact of higher resolution on UTE functional contrast. Regions of interest (ROIs) were defined in the parenchyma and pial surface of the visual cortex. Results: The administration of the contrast agent produced a bright-blood signal in the vasculature in structural MRI when using a UTE acquisition. Positive signal changes were observed at the shortest echo time (0.164 ms) in both the parenchyma (0.2% +/- 0.08) and pial surface (0.2% +/- 0.1 %), providing evidence that UTE fMRI experiments can detect changes in both pial and parenchymal vessels. Measurements using longer echo times ([≥]1 ms) showed negative signal changes. Higher spatial resolution resulted in increased percent signal change at the pial surface, suggesting less partial volume effects and better delineation of surface vessels. Conclusion: The findings demonstrate that T1-weighted fMRI with UTE imaging and iron oxide nanoparticles captures positive signal changes across all vascular compartments, providing additional insights into the involvement of surface vessels during functional hyperemia.

Interpersonal Neural Synchrony Across Levels of Interpersonal Closeness and Social Interactivity
Interpersonal neural synchrony is a key marker of social interactions, offering insights into the neural mechanisms underlying human connection and developmental outcomes. So far, hyperscanning studies have examined synchrony across diverse dyads and tasks, leading to inconsistencies and limiting cross-study comparability. This variability challenges the establishment of a unified theoretical framework for neural synchrony. This study investigated the effects of interpersonal closeness and social interactivity on neural synchrony using functional near-infrared spectroscopy hyperscanning. We recorded brain activity from 142 dyads (70 close-friend, 39 romantic-partner, and 33 mother-child dyads) across three interaction conditions: video co-watching (passive), a cooperative game (structured active), and free interaction (unstructured active). Neural synchrony was computed between participants' bilateral inferior frontal gyrus (IFG) and temporoparietal junction (TPJ) using wavelet transform coherence. Results showed that true dyads exhibited significantly higher synchrony than non-interacting surrogate dyads (q < .001), particularly in combinations involving the right IFG, left IFG, and left TPJ. Mother-child dyads displayed lower synchrony than adult-adult dyads at the global (p < .001) and local level of analysis. At the global level, synchrony was highest during video co-watching, followed by the cooperative game and free interaction (p < .001). However, left IFG-left IFG and left IFG-right TPJ synchrony peaked during the cooperative game. These findings highlight the role of brain maturation and social task structure in shaping neural synchrony. By providing an experimental framework, this study lays the groundwork for future hypothesis-driven hyperscanning research, advancing our understanding of the neural mechanisms underlying human social interactions.

Beyond Pairwise Connections in Complex Systems: Insights into the Human Multiscale Psychotic Brain
Complex biological systems, like the brain, exhibit intricate multiway and multiscale interactions that drive emergent behaviors. In psychiatry, neural processes extend beyond pairwise connectivity, involving higher-order interactions critical for understanding mental disorders. Conventional brain network studies focus on pairwise links, offering insights into basic connectivity but failing to capture the complexity of neural dysfunction in psychiatric conditions. This study aims to bridge this gap by applying a matrix-based entropy functional to estimate total correlation, a mathematical framework that incorporates multivariate information measures extending beyond pairwise interactions. We apply this framework to fMRI-ICA-derived multiscale brain networks, enabling the investigation of interactions beyond pairwise relationships in the human multiscale brain. Additionally, this approach holds promise for psychiatric studies, providing a new lens through which to explore beyond pairwise brain network interactions. By examining both triple interactions and the latent factors underlying the triadic relationships among intrinsic brain connectivity networks through tensor decomposition, our study presents a novel approach to understanding higher-order brain dynamics. This framework not only enhances our understanding of complex brain functions but also offers new opportunities for investigating pathophysiology, potentially informing more targeted diagnostic and therapeutic strategies. Moreover, the methodology of analyzing multiway interactions beyond pairwise connections can be applied to any signal analysis. In this study, we specifically explore its application to neural signals, demonstrating its power in uncovering complex multiway interaction patterns of brain activity, which provide a window to explore connectivity beyond pairwise interactions in the multiscale functionality of the brain.

Utility of the Recombinase Driver CX3CR1::Cre Rat Strain to Evaluate Microglial Contributions to Nicotine Addiction
Smoking remains a leading preventable cause of death, and nicotine is the primary substance responsible for maintaining use of tobacco products. Preclinical rodent models have shown that neuroimmune signaling is dysregulated by nicotine self-administration (SA) within the nucleus accumbens core (NAcore), which is a key region within the mesolimbic brain reward pathway. Microglia are the resident brain immune cell and prior studies have shown that they play an important role in nicotine-related behaviors. However, while there are transgenic mouse lines that allow for specific evaluations of microglia to neurobiology and behavior, there are fewer tools available for rats as a model species thus limiting our ability to evaluate specific contributions of microglia to nicotine SA. Using transgenic rats expressing Cre under the control of the CX3CR1 promoter bred on a Long Evans (LE) background, we show that NAcore microglia can be specifically transduced with Designer Receptors Exclusively Activated by Designer Drugs (DREADDs). We further show that CX3CR1::Cre rats readily self-administer nicotine, and display a characteristic extinction curve that is not different from outbred LE or Cre-negative littermates. Together, these validation studies lay the foundation for future use of this transgenic rat line to evaluate the specific contributions of microglia in the brain to neurobehavioral underpinnings of nicotine addiction.

Atrophin-1 Antisense Oligonucleotide Provides Robust Protection from Pathology in a Fully Humanized DRPLA Model
Dentatorubral-pallidoluysian atrophy (DRPLA) is a fatal neurodegenerative disease arising from a CAG repeat expansion in the atrophin-1 (ATN1) gene. Because DRPLA, like many repeat expansion disorders (REDs), arises predominantly from toxic gain-of-function mechanisms, we hypothesized that ATN1 knockdown would have therapeutic potential. To test this, we established the first fully-humanized mouse model of a RED, in which one allele of mouse Atn1 is completely replaced by human ATN1, including 112 pure CAG repeats. This novel approach to exploring RED biology provides significant advantages, notably the ability to test sequence-specific therapeutics targeting human sequences, even in introns and untranslated regions of pre-mRNA. We found that our model, the Atn1Q112/+ mouse, recapitulates key features of human DRPLA, including behavioral alterations, reduced brain size and aggregate accumulation. We treated Atn1Q112/+ mice with antisense oligonucleotides (ASOs) targeting mouse Atn1 (to probe for loss of function concerns), human ATN1, or a combination. Treatment with human, but not mouse, ATN1-targeting ASOs provides remarkable protection from a range of disease-related behavioral phenotypes, including aggregation of mutant ATN1 (mATN1), and marked rescue of transcriptional dysregulation in the cerebellum. These results have helped motivate an ongoing human clinical study of ASOs targeting ATN1 for DRPLA.

Associative coding of conditioned fear in the thalamic nucleus reuniens in rodents and humans
The nucleus reuniens (RE) is a midline thalamic structure interconnecting the medial prefrontal cortex (mPFC) and the hippocampus (HPC). Recent work in both rodents and humans implicates the RE in the adaptive regulation of emotional memories, including the suppression of learned fear. However, the neural correlates of aversive learning in the RE of rodents and humans remains unclear. To address this, we recorded RE activity in humans (BOLD fMRI) and rats (fiber photometry) during Pavlovian fear conditioning and extinction. In both rats and humans, we found that conditioned stimulus (CS)-evoked activity in RE reflects the associative value of the CS. In rats, we additionally found that spontaneous neural activity in RE tracks defensive freezing and shows anticipatory increases in calcium activity that precede the termination of freezing behavior. Single-unit recordings in rats confirmed that individual RE neurons index both the associative value of the CS and defensive behavior transitions. Moreover, distinct neuronal ensembles in the RE encode fear versus extinction memories. These findings suggest a conserved role of the RE across species in modulating defensive states and emotional memory processes, providing a foundation for future translational re-search on fear-related disorders.

Sex-specific and age-related progression of auditory neurophysiological deficits in the Cln3 mouse model of Batten disease
CLN3 disease is a prevalent form of Neuronal Ceroid Lipofuscinosis (NCL) caused by inherited mutations in the CLN3 gene, with symptoms such as vision loss, language impairment, and cognitive decline. The early onset of visual deficits complicates neurological assessment of brain pathophysiology underlying cognitive decline, while the small number of CLN3 mutation cases in humans hinders the study of sex differences. Building on our recent progress in assessing auditory neurophysiological changes in CLN3 patients, we developed a parallel approach using electroencephalography arrays in Cln3 knockout (Cln3-/-) mice to investigate the longitudinal progression of auditory processing deficits in both sexes. We employed a duration mismatch negativity (MMN) paradigm, similar to that used in our CLN3 patient studies, to assess the automatic detection of pattern changes in a sequence of stimuli. Wild-type mice of both sexes showed robust duration MMN responses when assessed longitudinally in the same subjects from 3 to 9 months of age. In contrast, female Cln3-/- mice developed consistent MMN deficits throughout this age range, while male Cln3-/- mice exhibited MMN deficits at younger ages that were mitigated at older ages. Analyses of auditory brainstem responses indicate that MMN abnormalities in Cln3-/- mice are not due to peripheral hearing loss. Instead, these deficits originate centrally from sex-specific and age-related changes in auditory evoked potentials elicited by standard and deviant stimuli. Our findings reveal a sex-specific progression of central auditory processing deficits in Cln3-/- mice, supporting auditory duration MMN as a translational neurophysiological biomarker for mechanistic studies and therapeutic development.

Visualization of functional and effective connectivity underlying auditory descriptive naming
Objective: We visualized functional and effective connectivity within specific white matter networks in response to auditory descriptive questions. Methods: We investigated 40 Japanese-speaking patients with focal epilepsy and estimated connectivity measures using cortical high-gamma dynamics and MRI tractography. Results: Hearing a wh-interrogative at question onset enhanced inter-hemispheric functional connectivity, with left-to-right callosal facilitatory flows between the superior-temporal gyri, contrasted by functional connectivity diminution with right-to-left callosal suppressive flows between dorsolateral prefrontal regions. Processing verbs associated with concrete objects or adverbs increased left intra-hemispheric connectivity, with bidirectional facilitatory flows through extensive white matter pathways. Questions beginning with what, compared to where, induced greater neural engagement in the left posterior inferior-frontal gyrus at question offset, linked to enhanced functional connectivity and bidirectional facilitatory flows to the temporal lobe neocortex via the arcuate fasciculus. During overt responses, inter-hemispheric functional connectivity was enhanced, with bidirectional callosal flows between Rolandic areas, and individuals with higher IQ scores exhibited less prolonged neural engagement in the left posterior middle frontal gyrus. Conclusions: Visualization of directional neural interactions within white matter networks during overt naming is feasible. Significance: Phrase order may influence network dynamics in listeners, even when presented with auditory descriptive questions conveying similar meanings.

Second-Scale Neural Dynamics Shape Hormonal Outputs in Hypothalamic CRH Neurons
The elevation of glucocorticoids is a hallmark of stress, arising from the integration of rapid neuronal signals into sustained hormonal outputs. A key interface for this neuroendocrine signal translation is corticotropin releasing hormone (CRH) neurons in the paraventricular nucleus of the hypothalamus (PVN), which release CRH at the median eminence (ME). We recently discovered that CRHPVN neurons exhibit a characteristic shift in firing patterns from rhythmic short-bursts (low-activity state) to tonic firing (high-activity state) in response to stress. This raised a critical question: how are these distinct firing patterns are integrated into slower, sustained hormonal outputs at neuroendocrine terminals? Here, we implemented optical approaches to detect CRH release ex vivo and in vivo. Using newly developed sniffer cells for CRH, we measured CRH release at ME ex vivo, triggered by the distinct, in vivo-like firing patterns. Our results demonstrated that the primary determinant of neuroendocrine CRH release was the firing rate sustained over the timescale of seconds, with little contribution from specific firing patterns. These results collaborated with second-scale increase in firing rate triggered by stress stimuli. Additionally, we recorded the dynamics of CRH at the ME in the freely moving mice using genetically-encoded GPCR-activation based (GRAB) sensors for CRH. Foot shock stress triggered transient, time-locked increases in CRH release on the timescale of seconds. Importantly, these second-scale CRH pulses, when elicited during repeated foot shocks, were integrated over minutes to scale downstream hormone releases. Together, our data revealed critical roles of second-scale dynamics in CRHPVN neuron activity for the neuroendocrine translation of stress signals.

Hitting the right pitch: Cortical tracking of fundamental frequency changes across speech rates in auditory and sensorimotor regions
Neuronal entrainment to speech properties is essential for language processing, particularly through oscillatory tracking of slower rhythms corresponding to the syllabic rate. However, it remains less explored whether brain rhythms also synchronize with higher-frequency speech components, particularly the fundamental frequency (F0) or pitch. We used magnetoencephalography (MEG) to investigate cortical tracking of F0 while participants listened to sentences produced at natural normal and fast rates, but also to time-compressed speech. We examined how pitch changes accompanying natural increases in speech rate modulate brain-to-speech coupling and compared this with artificially accelerated speech, where F0 is unchanged. We also explored whether this coupling is confined to the auditory cortex or involves a broader cortical network. We computed whole-brain cortico-acoustic coupling between the MEG source time-series and the speech signal, alongside spectral power modulations in frequency bands centered on the mean F0 of the speech material. We found significant brain-to-F0 coupling in the right auditory, inferior parietal, insular, and pre- and postcentral regions across all speech rate conditions. Importantly, the peak neuro-acoustic coupling frequency adjusted to reflect the F0 increase due to natural rate acceleration. Interestingly, we found significant brain-speech coupling around F0 not only in the primary auditory cortex but also in a postcentral somatosensory region likely corresponding to the ventral larynx area. These findings provide new insights into frequency-specific cortical tracking of F0 during the perception of naturally-produced speech at varying rates and suggest the involvement of an auditory-somato-motor network that may facilitate the recognition of articulatory features during speech perception.

Neural Correlates of Social Withdrawal and Preference for Solitude in Adolescence
Social isolation during development, especially in adolescence, has detrimental but incompletely understood effects on the brain. This study investigated the neural correlates of preference for solitude and social withdrawal in a sample of 2809 youth (median (IQR) age = 12.0 (1.1) years, 1440 (51.26%) females) from the Adolescent Brain Cognitive Development study. Older youth whose parents had mental health issues more frequently preferred solitude and/or were socially withdrawn (beta = 0.04 - 0.14, CI=[0.002,0.19], p<0.05), both of which were associated with internalizing and externalizing behaviors, depression, and anxiety (beta = 0.25 - 0.45, CI=[0.20,0.49], p<0.05). Youth who preferred solitude and/or were socially withdrawn had lower cortical thickness in regions supporting social function (cuneus, insula, anterior cingulate and superior temporal gyri) and/or mental health (beta = -0.09 to -0.02, CI=[-0.14,-0.003], p<0.05), and higher amygdala, entorhinal cortex, parahippocampal gyrus, and basal ganglia volume (beta = 2.62 - 668.10, CI=[0.13,668.10], p<0.05). Youth preferring solitude had more topologically segregated dorsal attention, temporoparietal, and social networks (beta = 0.07 - 0.10, CI=[0.02,0.14], p<=0.03). Socially withdrawn youth had a less topologically robustness and efficient (beta = -0.05 to -0.80, CI=[-1.34,-0.01], p<0.03), and more fragile cerebellum (beta = 0.04, CI=[0.01,0.07], p<0.05). These findings suggest that social isolation in adolescence may be a risk factor for widespread alterations in brain regions supporting social function and mental health.

Maternal and child immune profiles are associated with neurometabolite measures of early-life neuroinflammation in children who are HIV-exposed and uninfected: a South African birth cohort
Children who are HIV-exposed and uninfected (HEU) are at risk of neurodevelopmental delays, which may be partially due to maternal immune dysregulation during pregnancy. This study investigates associations between maternal and child immune profiles and early neurometabolite profiles in HEU and HIV-unexposed (HU) children from a South African birth cohort. A subgroup of 156 children (66 HEU, 90 HU) from the Drakenstein Child Health Study underwent magnetic resonance spectroscopy at age 2-3 years, and maternal and child serum markers were measured at multiple timepoints via immunoassays. In HEU children, serum concentrations of maternal pro-inflammatory cytokines IL-5 ({beta}=0.79, p=0.005) and IL-8 ({beta}=0.64, p=0.02) were associated with myo-inositol ratios in parietal grey and white matter regions, respectively, while child serum MMP-9 at two years was associated with myo-inositol ratios in the midline parietal grey matter ({beta}=1.30, p=0.03). The association of maternal anti-inflammatory cytokine IL-13 with glutamate ratios in the midline parietal grey matter was negative in HEU ({beta}=-0.41, p=0.038) and positive in HU children ({beta}=0.42, p<0.0001). These findings suggest maternal immune activation may affect neurometabolite profiles in HEU children.

Soleus H-reflex size versus stimulation rate in the presence of background muscle activity: A methodological study
Introduction: Hoffmann reflex (HR) operant conditioning has emerged as an important intervention in neurorehabilitation. During conditioning, the HR is elicited at low rates (~0.2 Hz) to avoid the initial reduction in HR size that can occur over repeated stimulation, i.e., rate-dependent depression (RDD), thereby maintaining reflex size. This study investigated the impact of higher stimulation rates on HR size, where a stable, low-level, background electromyographic (EMG) signal is maintained over 225 conditioning trials in each of 30 sessions. A higher rate could shorten session length and/or number. Methods: Fifteen healthy participants maintained low background soleus EMG (5-18 microV, ~1-3% of the maximum stimulation evoked direct muscle (M-wave) EMG response (Mmax) while standing. Soleus HR and M-wave recruitment curves were obtained at rates of 0.2, 1, and 2 Hz, from which Mmax and Hmax were calculated. Seventy-five HR trials (HRT) were collected for each stimulation rate at a target M-wave size (~10-20% of Mmax). Results: There was no evidence of RDD at higher stimulation rates. In addition, the mean HR over trials was reliable across participants and rates. The Intraclass Correlation Coefficient (ICC) was 0.965 (95%CI:0.915, 0.987). Discussion: This study shows that H-reflex conditioning might be performed at rates up to 2 Hz with no RDD and with consistent HR values. A faster rate could increase the number of conditioning trials per session, reduce session duration, and/or reduce the number of sessions. It could thereby accelerate the conditioning process and make the process less demanding for participants.

The Role of Syllabic Rhythm in Speech Perception Across Languages
The insertion of silences at regular intervals restores the intelligibility of English utterances that have been accelerated beyond comprehension, as long as the duration of the resulting speech-silence chunks falls within the theta rhythm of natural speech, i.e. the temporal modulation associated to the syllabic rate. We test whether such a rhythmic strategy works in languages rhythmically different from English, an stress-timed language. Thus, we assess whether comprehension of time-compressed Semantically Unpredictable Sentences (SUS) is restored in the syllable-timed language French and the mora-timed language Japanese, when silences re-establishing theta rhythm are inserted. Restoring the theta rhythm also improved intelligibility in French, but not in Japanese, in which best performance was instead achieved at faster rhythms, which suggests that modulation at the rate of a language's basic rhythmic unit plays a key role in understanding speech. In a second experiment, French speakers listened to SUS with speech-silence chunks adapted to the range of the temporal modulations of the delta, gamma, and high gamma rhythms, which correspond to the rate of prosodic phrases, phonemes, and subsegmental features, respectively. Unlike the theta rhythm, we found no restorative effects, providing further evidence for the special status of the theta rhythm in speech comprehension.

Cross-Frequency Coupling as a Neural Substrate for Prediction Error Evaluation: A Laminar Neural Mass Modeling Approach
Predictive coding frameworks suggest that neural computations rely on hierarchical error minimization, where sensory signals are evaluated against internal model predictions. However, the neural implementation of this inference process remains unclear. We propose that cross-frequency coupling (CFC) furnishes a fundamental mechanism for this form of inference. We first demonstrate that our previously described laminar neural mass model (LaNMM) supports two key forms of CFC: (i) Signal-Envelope Coupling (SEC), where low-frequency rhythms modulate the amplitude envelope of higher-frequency oscillations, and (ii) Envelope-Envelope Coupling (EEC), where the envelopes of slower oscillations modulate the envelopes of higher-frequency rhythms. Then, we propose that - by encoding information in signals and their envelopes - these processes instantiate a hierarchical ``Comparator'' mechanism at the columnar level. Specifically, SEC generates fast prediction-error signals by subtracting top-down predictions from bottom-up oscillatory envelopes, while EEC operates at slower timescales to instantiate gating - a critical computational mechanism for precision-weighting and selective information routing. To establish the face validity - and clinical implications of - this proposal, we model perturbations of these CFC mechanisms to investigate their roles in pathophysiological and altered neuronal function. We illustrate how, in disorders such as Alzheimer's disease, disruptions in gamma oscillations following dysfunction in fast-spiking inhibitory interneurons impact Comparator function with an aberrant amplification of prediction errors in the early stages and a drastic attenuation in late phases of the disease. In contrast, by increasing excitatory gain, serotonergic psychedelics diminish the modulatory effect of predictions, resulting in a failure to attenuate prediction error signals (c.f., a failure of sensory attenuation). Together, these results establish cross-frequency coupling - across temporal scales - as candidate computational processes underlying hierarchical predictive coding in health and disease.

Flexibility and Neural Correlates of Action-Sound Predictions
Abstract To interact efficiently with our environment, our brain predicts the sensory effects of our actions and compares them with the actual outcomes. This allows us to adapt our actions when predictions and sensory outcomes mismatch. While this process is generally well understood for action-sound predictions, it is an open question how flexibly these predictions can adapt in frequently changing environments, as they occur in real life. To investigate the flexibility of top-down predictions, we asked participants (N = 41) to press one of two buttons, a left-hand and a right-hand button, and switch hands autonomously. One button frequently produced a sound (80%) and rarely no sound. The other button frequently generated no sound (80%) and rarely produced a sound. In a third, separate condition, each button produced a sound in 50% of the trials. Unexpected sounds and unexpected sound omissions elicited a series of error-related brain responses in the electroencephalogram (EEG) at different levels of auditory processing, including a mismatch negativity (MMN) and the P3 complex for unexpected sounds, and the oN1, oN2, and oP3 complex for unexpected omissions. Moreover, unexpected sounds elicited an equivalent MMN-regardless of whether silence was expected (80%) or no reliable expectation was possible (50%), while later P3 components showed different amplitudes. Our results demonstrate flexible action-sound predictions at sensory and higher cortical levels. Furthermore, they indicate that predicted silence does not have an explicit sensory representation at lower levels but emerges at later stages, when higher-level information has been integrated.

Non-invasive in vivo bidirectional magnetogenetic modulation of pain circuits.
Primary nociceptors in the dorsal root ganglion (DRG) receive sensory information from discrete parts of the body and are responsible for initiating signaling events that in supraspinal regions will be interpreted as physiological or pathological pain. Genetic, pharmacologic and electric neuromodulation of nociceptor activity in freely moving non-transgenic animals has been shown to be challenging due to many factors including the immunogenicity of non-mammalian proteins, procedure invasiveness and poor temporal precision. Here, we introduce a magnetogenetic strategy that enables remote bidirectional regulation of nociceptor activity. Magnetogenetics utilizes a source of direct magnetic field (DMF) to control neuronal activity in cells that express an anti-ferritin nanobody-TRPV1 receptor fusion protein (Nb-Ft-TRPV1). In our study, RetroAAV2-mediated delivery of an excitatory Nb-Ft-TRPV1 construct into the sciatic nerve of wild-type mice resulted in stable long-term transgene expression accompanied by significant reduction of mechanical withdrawal thresholds during DMF exposure, place aversion of the DMF zone and activity changes in the anterior cingulate (ACC) nucleus. Conversely, delivery of an inhibitory variant of the Nb-Ft-TRPV1 construct, engineered to gate chloride ions in response to DMF, led to reversed behavioral manifestations of mechanical allodynia and showed place preference for the DMF zone, suggestive of functional pain relief. Changes in DRG activity were confirmed by post-mortem levels, immediately following DMF exposure, of the activity-induced gene cfos, which increased with the excitatory construct in normal mice and decreased with the inhibitory construct in pain models Our study demonstrates that magnetogenetic channels can achieve long-term expression in the periphery without losing functionality, providing a stable gene therapy system for non-invasive, magnetic field regulation of pain-related neurons for research and potential clinical applications.

Uncoupling overeating and fat storage by modulation of different serotonergic receptors
Psychotropic drugs such as antipsychotics improve symptoms of psychiatric disorders. However, they are associated with severe metabolic side effects that remodel energy balance, resulting in weight gain and increased food intake (hyperphagia). Here, we compare how antipsychotics and exogenous serotonin induce hyperphagia by remodeling energy balance. We find that the ability of serotonin and antipsychotics to remodel energy balance strictly depends on the serotonergic receptors SER-7 and SER-5, respectively. While both molecules induce hyperphagia, serotonin does so by increasing energy expenditure and reducing fat stores. In contrast, antipsychotics block the inhibitory effect of fat storage on feeding, thereby inducing hyperphagia and increasing fat stores. Thus, it is possible to manipulate energy balance to induce hyperphagia while either increasing or decreasing fat storage. Inactivation of the germline remodels energy balance similar to antipsychotic treatment, promoting hyperphagia while increasing fat storage. Consistent with overlapping mechanisms, antipsychotics are no longer able to remodel energy balance in both C. elegans and mice lacking an intact germline. Thus, our results uncouple overeating from fat storage and show that overeating can be induced by mechanisms that reduce or increase fat stores.

Concentration of myelin debris-like myelin basic protein-immunoreactive particles in the distal (anterior)-most part of the myelinated region in the normal rat optic nerve
In the myelinated region of the normal rat optic nerve, neuronal fibers are encircled by myelin sheaths except at nodes of Ranvier. In addition to these myelinated fibers, concentration of small particles was observed in the distal (anterior)-most part of the myelinated region. These particles were visualized by fluorescent immunohistochemistry using mouse monoclonal anti-human myelin basic protein (MBPh) antibody (clone SMI-99). Fluorescent double immunohistochemistry by using both the rat monoclonal anti-cow myelin basic protein (MBPc) antibody (clone 12) and the anti-MBPh antibody demonstrated that myelin basic protein immunoreactive-particles detected by the anti-MBPc antibody were almost completely overlapped with those immunostained by the anti-MBPh antibody. Since these antibodies have different target sites, these particles contained the real myelin basic protein. We hypothesized that the MBPh-immunoreactive particles were myelin debris-like structures in the normal rat optic nerve. Quantitative morphological analyses indicated that only 2 out of 6 differences in sizes and shape descriptors between the particles and myelin debris observed in the damaged-optic nerve were statistically significant. Glial fibrillary acidic protein-immunoreactivity and glutamine synthetase-immunoreactivity were seen in the particles. Majority of the particles were isolated from ionized calcium binding adapter molecule 1-labeled microglia. These results demonstrate that the myelin debris-like MBPh-immunoreactive particles are concentrated on the distal-most part of the myelinated region. This evidence suggests that the distal-most part is under mildly pathological condition. Furthermore, the evidence may provide clues as to the pathophysiological background that induces localized vulnerability of the myelin sheaths.

Expression of Tacr1 and Gpr83 by spinal projection neurons
Anterolateral system (ALS) projection neurons underlie perception of pain, itch and skin temperature. These cells are heterogeneous, and there have therefore been many attempts to define functional populations. A recent study identified two classes of ALS neuron in mouse superficial dorsal horn (SDH) based on expression of the G protein-coupled receptors Tacr1 or Gpr83. It was reported that cells expressing these receptors formed largely non-overlapping populations, and that ~60% of ALS cells in SDH expressed Tacr1. An additional finding was that while Tacr1- and Gpr83-expressing ALS cells projected to several brain nuclei, their axons did not reach the ventral posterolateral (VPL) thalamic nucleus, which is reciprocally connected to the primary somatosensory cortex. These results were surprising, because we had reported that ~90% of SDH ALS neurons in the mouse possess the neurokinin 1 receptor (NK1r), which is encoded by Tacr1, and in addition the VPL is thought to receive input from lamina I ALS cells. Here we use retrograde and anterograde labelling in Tacr1CreERT2 and Gpr83CreERT2 mice to reinvestigate the expression of the receptors by ALS neurons and to reassess their projection patterns. We find that ~90% of ALS neurons in SDH express Tacr1, with 40-50% expressing Gpr83. We also show that axons of both Tacr1- and Gpr83-expressing ALS neurons reach the VPL. These results suggest that ALS neurons in the SDH that express these GPCRs show considerable overlap, and that they are likely to contribute to the sensory-discriminative dimension of pain through their projections to VPL.

A human electrophysiological biomarker of Fragile X Syndrome is shared in V1 of Fmr1 KO mice and caused by loss of FMRP in cortical excitatory neurons
Predicting clinical therapeutic outcomes from preclinical animal studies remains an obstacle to developing treatments for neuropsychiatric disorders. Electrophysiological biomarkers analyzed consistently across species could bridge this divide. In humans, alpha oscillations in the resting state electroencephalogram (rsEEG) are altered in many disorders, but these disruptions have not yet been characterized in animal models. Here, we employ a uniform analytical method to show in males with fragile X syndrome (FXS) that the slowed alpha oscillations observed in adults are also present in children and in visual cortex of adult and juvenile Fmr1 -/y mice. We find that alpha-like oscillations in mice reflect the differential activity of two classes of inhibitory interneurons, but the phenotype is caused by deletion of Fmr1 specifically in cortical excitatory neurons. These results provide a framework for studying alpha oscillation disruptions across species, advance understanding of a critical rsEEG signature in the human brain and inform the cellular basis for a putative biomarker of FXS.

Heterogeneous distribution of glial cell marker proteins and of cell nucleus marker in the myelinated region of the normal rat optic nerve
Glial cells are critically important for maintenance of neuronal activity in the optic nerve. The present study documents the distribution of glial structure proteins (GFAP: glial fibrillary acidic protein; MBP: myelin basic protein), a glial functional protein (GS: glutamine synthetase), and of a cell nucleus marker (bisBenzimide) in the various myelinated regions of the normal rat optic nerve. Fourteen optic nerves from 12 male Sprague-Dawley rats were employed. Immunohistochemistry and confocal microscopy were used to investigate the distribution of GFAP, MBP, GS, and of bisBenzimide along the longitudinal plane of the myelinated region. GFAP-immunoreactivity and GS-immunoreactivity were strong in the distal (anterior)-most part, but weak in the proximal (posterior) part with a significant decrease in strength along the longitudinal plane of the myelinated region. bisBenzimide labeling was also strong in the distal-most part, but weak in the proximal part with a significant difference in strength in the myelinated region. MBP-immunoreactive particles and cell nuclei were densely distributed in the distal-most part, however they were sparsely dispersed in the proximal part with a significant difference. The heterogeneous distribution of GFAP, GS, bisBenzimide, cell nuclei, and MBP-immunoreactive particles along the longitudinal plane may be an important functional adaptation that reflects the not uniform nature of the physiological and structural environment of the myelinated region. Notably, the concentrations of GFAP, GS, and of MBP-immunoreactive particles in the distal-most part of the myelinated region suggest that this part is under a mildly pathological condition in the normal rat optic nerve.

scFv intrabody targeting wildtype TDP-43 presents protective effects in a cellular model of TDP-43 proteinopathy
TDP-43 proteinopathies are a set of neurological disorders characterized by the abnormal accumulation and mislocalization of TDP-43 in the cytoplasm, leading to the disruption of the normal function of the protein. In most of the cases, it is the wildtype (wt) form of the protein that is involved. An untargeted high-throughput screen of a single-chain variable fragment (scFv) library was performed using phage display against human full-length wt TDP-43. Two scFvs (B1 and D7) were retained following cellular expression (then termed intrabodies) and colocalization with cytoplasmic TDP-43 in vitro. We generated a 3D structure of full length wt TDP-43 in silico, and used it for epitope mapping. In a cellular model of TDP-43 proteinopathy, D7 enhanced the proteasomal degradation of the insoluble 35-kDa C-terminal fragment TDP-43 and reversed some TDP-43-induced metabolomic alterations, particularly relating to the lipid metabolism. Our findings offer a new scFv intrabody that bind to human wtTDP-43 and modify cellular pathways associated with TDP-43 proteinopathies.

Parameter optimisation for mitigating somatosensory confounds during transcranial ultrasonic stimulation
Transcranial ultrasonic stimulation (TUS) redefines what is possible with non-invasive neuromodulation by offering unparalleled spatial precision and flexible targeting capabilities. However, peripheral confounds pose a significant challenge to reliably implementing this technology. While auditory confounds during TUS have been studied extensively, the somatosensory confound has been overlooked thus far. It will become increasingly vital to quantify and manage this confound as the field shifts towards higher doses, more compact stimulation devices, and more frequent stimulation through the temple where co-stimulation is more pronounced. Here, we provide a systematic characterisation of somatosensory co-stimulation during TUS. We also identify the conditions under which this confound can be mitigated most effectively by mapping the confound-parameter space. Specifically, we investigate dose-response effects, pulse shaping characteristics, and transducer-specific parameters. We demonstrate that somatosensory confounds can be mitigated by avoiding near-field intensity peaks in the scalp, spreading energy across a greater area of the scalp, ramping the pulse envelope, and delivering equivalent doses via longer, lower-intensity pulses rather than shorter, higher-intensity pulses. Additionally, higher pulse repetition frequencies and fundamental frequencies reduce somatosensory effects. Through our systematic mapping of the parameter space, we also find preliminary evidence that particle displacement (strain) may be a primary biophysical driving force behind peripheral somatosensory co-stimulation. This study provides actionable strategies to minimise somatosensory confounds, which will support the thorough experimental control required to unlock the full potential of TUS for scientific research and clinical interventions.

Hair cell population integrity necessary to preserve vestibular function
Regeneration of hair cells is a primary target of gene therapy aimed at restoring vestibular and cochlear functions. Indeed, vestibular dysfunction constitutes a major medical concern, as one of its manifestation, dizziness, affects 15-35% of the general population, with a prevalence rate of 85% for those over 80 of age. Age-related alterations of both vestibular function and the integrity of vestibular hair cells has been reported in humans. However, direct comparisons between structural pathology and vestibular dysfunctions quantifications are lacking in humans and rather limited in animal models, representing a significant gap in current knowledge. Thus far, therapeutic trials in animal models targeting vestibular loss associated with genetic diseases have yielded varied and partial results, and the functional identity and quantity of hair cells sufficient to restore minimal or normal vestibular function remain undefined. Here, we further develop an innovative methodology to bridge the gap between hair cells integrity and functional vestibular loss in individuals. Gradual vestibular deficits were induced through a dose-dependent ototoxic compound, quantified with canal or utricular-specific vestibulo-ocular reflex tests, and were then correlated in all individuals with the loss of type I and type II hair cells in different regions of ampulla and macula. Our findings reveal that the structure-function relationship is nonlinear, with lower bound of approximately 50% of hair cells necessary to retain minimal vestibular function, and threshold exceeding 80% to preserve normal function, thus shedding light on population-coding mechanisms for vestibular response. Our data further support the decisive role of type I, rather than type II, HC in the tested VOR functions.

Gamma oscillations in basal ganglia stem from the interplay between local inhibition and beta synchronization
Basal ganglia activity fluctuations have primarily been studied in the context of beta (12-30 Hz) oscillations, a well-established neural marker for Parkinson's Disease (PD). Recent studies have also identified gamma (30-100 Hz) oscillations within the basal ganglia, suggesting it could serve as an alternative marker, but the underlying circuit mechanisms remain poorly understood. Here, through a spiking network model of the basal ganglia, we identified two distinct gamma oscillations: a high-frequency gamma rhythm within the globus pallidus (GPe-TI) and a slower gamma rhythm within D2 medium spiny neurons (MSNs), both stemming from self-inhibition. When we simulated dopamine depletion to mimic the effects of PD, the intensity of gamma oscillations in the GPe-TI was not affected, but their peak frequency increased due to phase-amplitude coupling with pathological beta oscillations. This suggests that the GPe-TI loop, while robust to dopamine depletion, becomes more synchronized with beta activity in the context of PD, leading to faster gamma rhythms. In contrast, gamma oscillations in D2 MSNs were not present in simulated healthy condition and only emerged under dopamine-depleted pathological conditions. Moreover, both their intensity and peak frequency were strongly modulated by pathological beta activity. Together, these findings highlight the complementary roles of self-inhibition and beta oscillations in shaping gamma activity within basal ganglia circuits. The GPe-TI loop primarily sustains high-frequency gamma rhythms, while low frequency gamma rhythms in D2 MSNs are strongly dependent on dopamine-depletion-related beta modulation. These results underscore the importance of network-wide interactions in PD, where pathological beta oscillations influence gamma activity. This study offers insights into the mechanisms of gamma oscillations in PD and highlights the potential of gamma activity, in both the prototypical and striatal loops, as a marker for disease progression and monitoring pathological dysfunction in PD.

Predicting the Sound-Induced Flash Illusion: A Time-Window-of-Integration Approach
{The sound-induced flash illusion (SIFI) refers to the observation that pairing a single flash with 2 auditory beeps leads to the illusory perception of 2 visual stimuli (fission illusion). Susceptibility to the illusion depends on many factors like exact physical stimulus parameters, participants` expectations, attention, age, and clinical/non-clinical population membership. While there exist many experimental studies, very few models have been proposed to account for the phenomenon. Here we suggest a formal model (SIFI-TWIN) based on the notion of a temporal binding window that predicts the occurrence of illusory flashes as a function of the temporal and physical stimulus arrangement. The model`s performance is illustrated on a study investigating differences in SIFI performance between elderly hearing aid users and those with the same degree of mild hearing loss who were not using hearing aids. The results suggests that the higher incidence of reporting an illusory flash for hearing-aid users is due to both a larger temporal window of integration and a larger bias to report the illusion. The SIFI-TWIN model will help to better understand the diverse results from clinical and non-clinical studies as well as the cognitive foundations of the SIFI.

In vivo genetic labeling of primary cilia in developing astrocytes.
Astrocyte cilia are largely understudied due to the lack of available tools. Astrocyte research advanced with the establishment of Aldh1l1-CreERT2, an inducible Cre line that specifically targets the astrocyte lineage. Here, we develop and compare genetic models that label astrocyte cilia in the developing prefrontal cortex (PFC) using Aldh1l1-CreERT2 and Cre-dependent cilia reporters. We evaluate these models by testing different tamoxifen-induction protocols and quantifying the percentage of astrocytes labeled with the cilia reporters. We show that tamoxifen dosage impacts the expression of cilia reporters in astrocytes. We validate the maximum cilia-labeling efficiency of tamoxifen using constitutively-expressed cilia reporters. The data reveal that only a subset of SOX9- positive astrocytes in the PFC possess cilia throughout development. Our work highlights the utility of Cre-Lox systems to target specific cell types and the importance of carefully validating genetic models.

Differential modulation of movement speed with state-dependent deep brain stimulation in Parkinson's disease
Subthalamic deep brain stimulation (STN-DBS) provides unprecedented spatiotemporal precision for the treatment of Parkinson's disease (PD), allowing for direct real-time state-specific adjustments. Inspired by findings from optogenetic stimulation in mice, we hypothesized that STN-DBS effects on movement speed depend on ongoing movement kinematics that patients exhibit during stimulation. To investigate this hypothesis, we implemented a motor state-dependent closed-loop neurostimulation algorithm, adapting DBS burst delivery to ongoing movement speed in 24 PD patients. We found a stronger anti-bradykinetic effect, raising movement speed to the level of healthy controls, when STN-DBS was applied during fast but not slow movements, while only stimulating 5% of overall movement time. To study underlying brain circuits and neurophysiological mechanisms, we investigated the behavioral effects with MRI connectomics and motor cortex electrocorticography. Finally, we demonstrate that machine learning-based brain signal decoding can be used to predict continuous movement speed for fully embedded state-dependent closed-loop algorithms. Our findings provide novel insights into the state-dependency of invasive neuromodulation, which could inspire advanced state-dependent neurostimulation algorithms for brain disorders.

Differential retinal ganglion cell resilience to optic nerve injury across vertebrate species
Optic neuropathies comprise a diverse group of disorders that ultimately lead to retinal ganglion cell (RGC) degeneration. Despite varying etiologies, these conditions share a conserved pathological progression: axonal damage in the optic nerve triggers progressive RGC degeneration. Understanding species-specific differences in neuronal resilience is critical for identifying key survival mechanisms and potential neuroprotective targets. In this study, we compare RGC densities and survival rates following optic nerve crush (ONC) in three vertebrate models-mice, zebrafish, and killifish-under standardized experimental conditions. Transcriptomic analysis confirmed that, similar to RBPMS in mice, Rbpms2 serves as a pan-RGC marker in zebrafish and killifish. Using these markers, we reveal significant species-specific differences in RGC density, with fish species exhibiting over a five-fold higher density than mice at equivalent life stage. Killifish also show an age-dependent decline in RGC density. Furthermore, we identify distinct injury responses across species: mice undergo rapid degeneration, losing ~80% of their RGCs by day 14 after ONC; zebrafish maintain full RGC retention for two weeks before experiencing a loss of ~12%; and killifish display a biphasic response to ONC, with young adults retaining two-thirds of their RGCs by day 21, while older fish exhibit a more pronounced second wave of RGC loss, ultimately preserving just over half of their RGCs by 21 days after injury. These findings highlight fundamental differences in neuroprotective capacity among species, providing a comparative framework to uncover molecular mechanisms governing RGC survival and to identify therapeutic strategies for treating optic neuropathies and neurodegeneration across diverse pathologies.

Beyond the Resection: Surgical White Matter Disruption alters Non-Resected Brain Anatomy
Resective neurosurgery is a cornerstone treatment for many neurological conditions. Although traditionally viewed as localised procedure, increasing evidence from advanced magnetic resonance imaging (MRI) shows that also non-resected anatomy can degenerate following surgery. The relationship between local tissue removal and these postoperative changes remains thus far speculative. Here, we investigate the hypothesis that degenerative changes to surgically preserved grey and white matter are mediated by transneuronal degeneration, a deterioration of intact neuronal populations due to lost axonal input. Using a robust structural and diffusion MRI framework, we first identify widespread postoperative atrophy: pronounced cortical thickness decreases near the resection, and extensive white matter impairments across the ipsilateral hemisphere. Importantly, we then link these alterations to surgical white matter disruption, revealing a sequential network atrophy following neurosurgery. Beyond degenerative effects, we also demonstrate often reported structural network reorganisations as an artefact of image processing, indicating limited capacity for macroscale plasticity post-resection.

Persistent Anhedonia After Intermittent Long-Access Nicotine Self-Administration in Rats
Tobacco use disorder is a chronic condition characterized by compulsive nicotine use and withdrawal symptoms after smoking cessation. Smoking is the leading preventable cause of morbidity and mortality worldwide. Smoking cessation leads to anhedonia, which is an inability to experience pleasure from previously enjoyed activities and is caused by dysregulation of the brain's reward and stress systems. It is also a key withdrawal symptom that contributes to relapse to smoking after a period of abstinence. To better understand the development of anhedonia, we investigated its onset and time course in rats that self-administered nicotine. Rats were implanted with intracranial self-stimulation (ICSS) electrodes to assess reward function and intravenous catheters for nicotine self-administration. Elevations in ICSS brain reward thresholds reflect decreased sensitivity to rewarding electrical stimuli, indicating anhedonia. The rats self-administered 0.06 mg/kg of nicotine intermittently, three days per week, for seven weeks. Brain reward thresholds were determined once a week 24 h after nicotine self-administration during weeks 1 to 3, and at 12, 24, and 48 h during weeks 4, 5, and 7. Elevations in brain reward thresholds were not observed during the first four weeks of nicotine self-administration. However, the brain reward thresholds were elevated in both weeks 5 and 7 at least 12 h after nicotine self-administration, indicating that anhedonia emerges gradually and then persists. As withdrawal severity gradually increases, smoking cessation may become more challenging. Therefore, behavioral or pharmacological interventions soon after smoking initiation are critical to prevent the development of a tobacco use disorder.

Uncovering Sleep's Hippocampal-Cortical Dialogue: The Role of Deltas' and Spindles' Cross-Area Synchronization and Ripple Subtypes
ippocampal ripples, critical for sleep-related memory consolidation, are heterogeneous events with various sources and functions. Here we applied principal component analysis to identify ripple sub-types and relate them to hippocampal-cortical interactions as well as their role in consolidating simple and complex semantic-like memories in rats. Three main ripple types were discovered: baseline, large-input, and small-input ripples. Small-input ripples, were associated with increased prefrontal cortex to hippocampus connectivity, followed hippocampal delta waves, and were sufficient for simple learning. In contrast, large-input ripples exhibited increased hippocampus to prefrontal cortex connectivity, occurred during hippocampal spindles together as a doublet with a small-input ripple, and were critical for complex memory consolidation. Finally, learning induced heightened coupling between hippocampal delta and spindle oscillations and their cortical counterparts, consequently leading to an increased synchronization of ripples with cortical oscillations.

Meditation induces shifts in neural oscillations, braincomplexity and critical dynamics: Novel insights fromMEG
While the beneficial impacts of meditation are increasingly acknowledged, its underlying neural mechanisms remain poorly understood. We examined the electrophysiological brain signals of expert Buddhist monks during two established meditation methods known as Samatha and Vipassana, which employ focused at- tention and open monitoring technique. By combining source-space magnetoencephalography (MEG) with advanced signal processing and machine learning tools, we provide an unprecedented assessment of the role of brain oscillations, complexity and criticality in meditation. In addition to power spectral density (PSD), we computed long-range temporal correlations (LRTC), deviation from criticality coefficient (DCC), Lempel-Ziv complexity (LZC), 1/f slope, Higuchi fractal dimension (HFD), and spectral entropy. Our findings indicate increased levels of neural signal complexity during both meditation practices compared to the resting state, alongside widespread reductions in gamma-band LRTC and 1/f slope. Importantly, the DCC analysis revealed a separation between Samatha and Vipassana, suggesting that their distinct phenomenological properties are mediated by specific computational characteristics of their dynamic states. Furthermore, in contrast to most previous reports, we observed a decrease in oscillatory gamma power during meditation, a divergence we attribute to the correction of the power spectrum by the 1/f slope. We discuss how these results advance our comprehension of the neural processes associated with focused attention and open monitoring meditation practices.

Characteristics of Tactile Orientation Perception: Oblique Effect, Active vs Passive Exploration, and Serial Dependence
Orientation perception is one of the most fundamental aspects of somatosensory perception, similar to its visual counterpart. In the current study, we examined several features of tactile orientation perception and their interactions, aiming to better understand the underlying mechanisms. We found that active exploration exhibits better orientation acuity compared to passive exploration. We also observed an exploration-independent and paradigm-independent robust anisotropy (i.e., tactile oblique effect) in tactile orientation acuity where the proximal-distal axis demonstrates superior orientation acuity compared to oblique and medial-lateral orientations. Finally, we demonstrated that, similar to its visual counterpart, tactile orientation perception showed a systematic attractive serial dependence effect, in which current orientation perception is biased towards the previous trial. However, this attractive effect is driven by post-perceptual processing/response rather than the stimulus per se.