bioRxiv Subject Collection: Neuroscience's Journal

Non-canonical internalization mechanisms of mGlu receptors
Cell surface density of G protein-coupled receptors (GPCRs) is tightly regulated through constitutive and agonist-induced internalization. Whereas the mechanisms of constitutive internalization remain elusive, agonist-induced internalization is accepted to involve receptor phosphorylation by GPCR kinases (GRKs), {beta}-arrestin binding and AP2 recruitment, targeting receptors to clathrin-coated pits. Dimeric class C metabotropic glutamate (mGlu1 to 8) receptors regulate synaptic transmission but their internalization process is ambiguous. Here, we used diffusion-enhanced energy transfer (DERET) to decipher their internalization kinetics. We showed that all mGlu receptors are constitutively internalized. However, only mGlu1, 5 and 3 homodimers are agonist-induced internalized, that require neither GRKs, nor {beta}-arrestins. In contrast, the constitutive internalization involves only {beta}-arrestins. This systematic study further illustrates how different class C receptors are relative to most other GPCRs, revealing non-canonical internalization mechanisms. These insights in mGlu receptor dynamics will help promoting the therapeutic action of drugs targeting mGlu receptors.

Exposure to bullying engages social distress circuits in the adolescent and adult brain
Despite advances in understanding the psychological and social consequences of peer victimization, the immediate effects of bullying on the central nervous system remain elusive. Here we mapped the neural responses to simulated bullying in adolescents and adults and tested whether these responses are associated with real-life victimization experiences. Fifty-one adolescents aged 11-14 years, and 47 adults underwent a functional MRI (fMRI) while watching first-person videos of bullying (victimization) in the school environment, as well as neutral and positive social interactions in a similar setting. Exposure to bullying versus positive social interaction engaged the socio-emotional and threat response systems, as well as regions related to social cognition, sensory and interoceptive processing, and motor control. These responses were consistent across adolescents and adults and dependent on the current and past victimization experiences of the participants. This large-scale activation of neural systems subserving socioemotional, somatosensory, and interoceptive processing highlights how peer victimization evokes a severe stress and alarm state in the central nervous system.

Elusive scents: neurocomputational mechanisms of verbal omissions in free odor naming
Odor naming is considered a particularly challenging cognitive test, but the underlying cause of this difficulty is unknown. People often fail to report any source label to identify common odors, resulting in omissions (i.e., a lack of response). Here, with the support of a computational model, we offer a hypothesis about the neural network mechanisms underlying odor naming omissions. Based on an evaluation of behavioral data from almost 40,000 odor naming attempts, we suggest that high omission rates are driven by odors that are referred to by multiple linguistic labels. To explain this observation at the systems level, where olfactory perception and language (semantic) processing are produced by interacting cortical systems, we developed a computational model consisting of two associatively coupled attractor memory networks (odor and language networks), and investigated the effect of Hebbian-like learning on the simulated task performance. We used distributed network representations for the odor percepts and word label mental objects, and accounted for their statistical inter-relationships (correlations) extracted from collected data on odor perceptual similarity, and from a large Swedish odor language corpus, respectively. We evaluated a novel hypothesis, that Bayesian-Hebbian synaptic plasticity mechanisms can explain behavioral omissions in odor naming tasks, casting new light on the underlying mechanisms of this frequently observed memory phenomenon. Due to the nature of Bayesian-Hebbian associative learning connecting the two networks, there was a progressively weaker coupling for odors paired with multiple different labels in the encoding process (one-to-many mapping). Thus, when the model was cued with perceptual odor stimuli that established multiple word label associations (one-to-many mapping), the olfactory language network often produced subthreshold network responses, resulting in elevated omissions (opposite to one-to-few mapping scenario that led to improved performance scores). Our results are of theoretical interest, as they suggest a biologically plausible mechanism to explain a common, but poorly understood, behavioral phenomenon.

Circuit-Based Understanding of Fine Spatial Scale Clustering of Orientation Tuning in Mouse Visual Cortex
In sensory cortex of brain it is often the case that neurons are spatially organized by their functional properties. A hallmark of primary visual cortex (V1) in higher mammals is a columnar functional map, where neurons tuned to different stimuli features are regularly organized in space. However, rodent visual cortex is at odds with this rule and lacks any spatially ordered functional architecture, and rather neuron feature preference is haphazardly organized in patterns termed "salt-and-pepper". This sharp contrast in feature organization between the visual cortices of rodents and higher mammals has been a persistent mystery, fueled in part by abundant evidence of conserved cortical physiology between species. In this work, we applied a novel GCaMP indicator that are localized in the nucleus of neurons during two photon imaging in mouse V1, which enabled us to overcome most spurious spatially correlated activity due to fluorescence contamination, and to ensure a faithful observation of functional organization over space. We found that the orientation tuning properties of distant neuron pairs (> 20 um) are irregularly and randomly organized, while neuron pairs that are extremely close (< 20 um) have strongly correlated orientation tuning, indicating a narrow yet strong spatially clustered organization of orientation preference, which we term "micro-clustered" organization. Exploring a circuit based model of recurrently coupled mouse V1 we derived two key predictions for the micro cluster: spatially localized recurrent connections over a comparable narrow spatial scale, and common relative spatial spreads of balanced excitation and inhibition in the network over broad spatial scales. These predictions are validated by both anatomical and optogenetic-based physiological circuit mapping experiments. Altogether, our work takes an important step in building a circuit-based theory of visual processing in mouse V1 over spatial scales that are often ignored, yet contain powerful synaptic interactions.

Dissociating the incubation of appetitive and consummatory behavior in a model of oral cocaine self-administration
Cocaine use disorder remains a persistent public health dilemma that currently lacks effective treatment strategies. One key impediment to successful treatment outcomes is increased drug craving that occurs over the course of abstinence and subsequent relapse to drug use. This phenomenon, known as the incubation of drug craving, has been modeled extensively in rodent models of intravenous drug self-administration. As commonly implemented, the design of intravenous self-administration preclinical studies precludes disentangling appetitive and consummatory behaviors as drug seeking (appetitive) and taking (consummatory) is simultaneous. Here, we employed a model of oral cocaine self-administration to interrogate the incubation of drug vs nondrug craving, where the route of administration is identical between reinforcers and appetitive and consummatory behaviors are dissociable. Oral self-administration of cocaine produced detectable levels of cocaine and its metabolite, benzoylecgonine, within the blood and brain, and blood and brain levels of both substrates correlated with cocaine consumption. When tested for seeking- (lever pressing) and taking-related (magazine head entries) behavior after 1 or 21 days of forced abstinence from cocaine or saccharin, we observed incubation of lever pressing among cocaine-administering mice and incubation of magazine entries among saccharin-administering mice. These behavioral changes were accompanied by reduced expression of the glial glutamate transporter GLT-1 within the nucleus accumbens (NAc) of cocaine self-administering mice, regardless of abstinence. Altogether, these results underscore the utility of this model of cocaine self-administration, highlight the conserved nature of incubated cocaine seeking across routes of administration, and demonstrate the dissociable neurobehavioral sequelae of the incubation of reward seeking across reinforcer types.

The Thermal Grill Elicits Central Sensitization
The thermal grill, in which innocuous warm and cool stimuli are interlaced, can produce a paradoxical burning pain sensation--the thermal grill illusion (TGI). While the mechanisms underlying TGI remain unclear, prominent theories point to spinal dorsal horn integration of innocuous thermal inputs to elicit pain. It remains unknown whether the TGI activates peripheral nociceptors, or solely thermosensitive afferents and is integrated within the spinal cord. Different types of sensitization have established mechanisms and can inform TGI mechanisms: if the TGI elicits (1) primary hyperalgesia, peripheral nociceptors are activated; (2) secondary hyperalgesia in the absence of primary hyperalgesia, spinal integration is required; and (3) brush allodynia, wide-dynamic range neurons are involved in mediating the TGI. Here, we determine whether the TGI elicits primary hyperalgesia, secondary hyperalgesia or brush allodynia. Fifty-two participants underwent individually calibrated phasic thermal grill stimulation. We found that the TGI elicited primary hyperalgesia, but only in participants with component temperatures in the noxious range (<19 {degrees}C and >41 {degrees}C). The TGI also elicited secondary hyperalgesia, even in participants with strictly innocuous thermal inputs. No participants developed brush allodynia. We observed sex differences in primary hyperalgesia: only males exhibited thermal grill-induced primary hyperalgesia. These findings suggest that the TGI is integrated in the spinal dorsal horn, likely mediated by heat-pinch-cold (HPC) neurons, and, to some degree, by primary nociceptive afferents in males. This study shows that the TGI may have sex-dependent mechanisms and determines that HPC cells are involved in the illusory sensation of pain from innocuous thermal inputs.

Common Neural Choice Signals reflect Accumulated Evidence, not Confidence
While much research has investigated decision confidence, its neural underpinnings remain unclear. Although several studies have shown that centro-parietal EEG signals correlate with the reported level of confidence, according to recent computational work these signals reflect evidence which feeds into the computation of confidence, but they do not themselves track confidence. To test this prediction, we capitalized on our recent finding that a causal manipulation of prior beliefs selectively affects confidence, while leaving objective task performance unaffected. Using EEG recordings, we tested whether neural signals known to correlate with confidence (CPP and Pe) reflect evidence accumulation or confidence. Behaviorally, we replicated the finding that manipulating prior beliefs causally and selectively affected confidence without changing objective task performance. The EEG data showed a monotonic relation between the reported level of confidence and both CPP and Pe amplitudes. Importantly, this finding is compatible both with the theory that these signals track confidence as well as with the alternative theory that they track accumulated evidence. Critically, both neural signals were insensitive to the influence of prior beliefs on confidence, showing that they reflect the accumulated evidence that is used by the system to compute confidence, instead of directly reflecting confidence. Likewise, oscillatory activity in alpha and beta band was insensitive to the influence of prior beliefs on confidence. Decoding analyses revealed that the brain does hold shared representations for prior beliefs and confidence, and we identified a frontal signal that is sensitive to both confidence and prior beliefs. These findings are in line with recent views suggesting that the CPP and the Pe reflect evidence accumulation, which is combined with prior beliefs to form a confidence judgment.

Successful axonal regeneration is driven by evolutionarily conserved metabolic reprogramming
Unlike mammals, zebrafish can regrow axons after injury and restore circuit function in the central nervous system (CNS). Mitochondria have been identified as key players in this process, but how different metabolic pathways work together to sustain regeneration remains unclear. Using RNA sequencing of adult zebrafish retinal ganglion cells after optic nerve crush injury, we found that oxidative phosphorylation is downregulated during axonal regrowth. Simultaneously, the thioredoxin antioxidant system was upregulated, likely to limit oxidative damage. Additionally, we observed an integrated upregulation of glycolysis and the pentose phosphate pathway during the initial regrowth phases, possibly to provide energy and supplying NADPH for biosynthesis and antioxidant responses. We show that this metabolic reprogramming is evolutionarily conserved, as a comparable one occurs in the pro-regenerative mammalian Pten and Socs3 co-deletion model. Inhibiting glycolysis and thioredoxin in zebrafish impairs axonal regrowth, suggesting that targeting these pathways could enhance CNS regeneration in mammals.

A Novel Mouse Model of Parkinson's Disease for Investigating Progressive Pathology and Neuroprotection
Developing animal models that successfully recapitulate the features of progressive Parkinson's disease (PD) is crucial for understanding disease progression mechanisms and creating effective therapeutic interventions. In this study, we created a mouse model of PD by overexpressing -synuclein through a combined injection of AAV6--synuclein and preformed fibrils (PFFs) into the medial and lateral substantia nigra (SN). We also demonstrated that chronic administration of the c-Abl inhibitor PD180970 provides neuroprotection in this model. Mice injected with the AAV6--synuclein and PFF combination showed a progressive loss of dopaminergic (DA) neurons in the SN and their projections in the striatum over 24 weeks. This neuronal loss coincided with a time-dependent accumulation of phosphorylated -synuclein (p-syn) in the SN. The p-syn aggregates spread to synaptically connected DARPP-32-positive neurons in the striatum and further extended to the cortex. We also observed a contralateral spread of p-syn aggregates. Additionally, -synuclein overexpression led to a significant increase in activated microglia and astrocytes at all timepoints, with the strongest activation occurring early and gradually diminishing over time. Daily administration of PD180970 significantly reduced the loss of DA neurons caused by -synuclein injection and decreased the accumulation of p-syn in the SN. PD180970 treatment also reduced the neuroinflammation significantly. Overall, the combined injection of AAV6--synuclein and preformed fibrils into the mouse brain establishes a robust PD model, enabling detailed mechanistic studies of the disease. We further demonstrate the model's utility for chronic neuroprotection studies using the potential drug PD180970, highlighting its broad applicability.

Neural dynamics of proactive and reactive cognitive control in medial and lateral prefrontal cortex
Goal-directed behavior requires adjusting cognitive control to both react to and prepare for conflict. Previous work indicates theta oscillations and population activity in dorsomedial prefrontal cortex (dmPFC) and dorsolateral prefrontal cortex (dlPFC) are critical for reactive control. However, the neural mechanisms supporting proactive control are less clear. Here, we investigated the neural basis of behavioral adaptations when control is prepared in anticipation of conflict using intracranial EEG (iEEG) in dmPFC and dlPFC during a Stroop task where conflict frequency was manipulated across blocks. We observed canonical conflict-driven increases in dmPFC theta and in dmPFC and dlPFC local population activity, as indexed by high frequency activity (HFA). Conflict also suppressed theta power in both regions after the response, accentuated a pre-response beta desynchronization selectively in dlPFC, and increased a post-response beta rebound in both regions. Importantly, we identified a pre-trial marker of proactive control where dmPFC theta power increased before trials when conflict was expected, and theta, beta, and HFA conflict signals in both regions were enhanced when conflict was rare and diminished when conflict was common. These findings reveal shared HFA but dissociable oscillatory dynamics in dmPFC and dlPFC during reactive conflict processing, highlight pre-trial dmPFC theta as a potential substrate for proactive control, and refine the roles of dmPFC and dlPFC in control adaptations.

Sex-specific effects of appetite suppressants and stereotypy in rats
This study investigated the sex-specific effects of commonly prescribed appetite suppressants on body weight and the manifestation of motor side effects, specifically stereotypy. Employing video recordings and DeepLabCut (DLC) for precise behavioral quantification, we analyzed stereotypy, defined as purposeless, repetitive motor behaviors, in male and female rats. Under control (saline) conditions, male rats exhibited a greater propensity for weight gain compared to females. However, in contrast, female rats demonstrated greater and more homogenous weight loss than males following the administration of diethylpropion and tesofensine. Phentermine and mazindol induced comparable weight loss in both sexes, whereas cathine elicited weight reduction exclusively in males. 5-HTP and d-amphetamine administration only prevented weight gain relative to controls. Analysis of motor side effects revealed that drugs primarily targeting dopamine pathways, specifically phentermine, mazindol, diethylpropion, cathine, and d-amphetamine, induced pronounced stereotypies, particularly head-weaving, in both sexes. Interestingly, tesofensine elicited head-weaving behavior exclusively in female subjects, albeit to a lesser extent than that observed with other dopaminergic agents; conversely, tesofensine was most frequently associated with orolingual dyskinesia. Moreover, one of the most potent forms of stereotypy backward locomotion, here referred to as "moonwalking" was sporadically observed only following the administration of phentermine, diethylpropion, cathine, and mazindol, with diethylpropion inducing it most frequently. Male subjects treated with these same drugs exhibited an unexpected effect: spontaneous ejaculations, potentially attributable to the combined effects on dopamine and serotonin signaling in brain regions regulating sexual function. Network analysis and Markov transition matrices revealed distinct behavioral profiles associated with head-weaving, which emerged as the dominant attractor state, suggesting potential mechanistic differences among these drugs. Collectively, this study provides a valuable database characterizing the behavioral side effects of appetite suppressants.

Social, cognitive and sensory dimensions of cortical network overconnectivity in young children with autism spectrum disorder
The heterogeneity in both the neurobiological mechanisms and the phenotypic presentation of autism spectrum disorder (ASD) poses a major challenge to clinical and translational research. Early inefficiencies in functional connectivity (FC) have been associated with ASD, yet it remains unclear whether and how abnormal brain network properties may account for individual differences across ASD-related symptomatology and behaviors. We applied source-level reconstruction to resting-state high-definition EEG data in a cohort of 113 young children (40 with ASD) to identify early global and local alterations of cortical network connectivity. We subsequently used regularized canonical correlation analysis (rCCA) to characterize specific FC patterns linked to variation in cognitive, social and sensory dimensions within the autism spectrum. We found increased low-frequency FC in frontotemporal cross-hemispheric networks and lateral-occipital regions of young ASD children. RCCA revealed three distinct FC patterns in recurrent ASD-related networks, each contributing to predict individual differences in cognitive, social and sensory features. These linked FC-behavior dimensions may shed light on atypical brain network topology conferring risk for specific phenotypic manifestations of ASD, which may implicate unique underlying neurobiological mechanisms.

The effect of chronic stress and chronic alcohol intake on behavior, brain structure, and functional connectivity in a rat model
Pathological chronic stress is stress exceeding the organism's ability to cope physiologically, which may act as a risk factor in the onset and relapse of alcohol use disorder. Chronic- restraint stress (CRS) and ethanol intake are independently known to induce changes in brain structure and function, however, their combined effects on neurodevelopment over long periods of time remains largely unexplored. We conducted an in vivo longitudinal rat model with three main goals. 1) to determine if chronic stress increases ethanol intake; 2) to determine the effect of chronic- stress and ethanol intake in behavioral measures, brain structure, and function; and 3) to investigate the effect of sex. This observational study included Wistar rats assigned to four groups: 1) ethanol consumption (EtOH+/CRS-), 2) stress exposure (EtOH-/CRS+), 3) both ethanol and stress exposure (EtOH+/CRS+), and 4) control group (EtOH-/CRS-). Our results showed that chronic stress did not affect ethanol intake but led to reduced body weight gain, elevated corticosterone levels, and impaired recognition memory. Structural MRI revealed that both exposures produced additive brain volume changes in regions such as the olfactory bulb, orbitofrontal cortex, caudate-putamen, hippocampus, and cerebellum. Functional connectivity analysis using network-based statistics identified disrupted cortical-subcortical connections. Results found here were sex-dependent in terms of volumetric changes (higher effects on males) and functional connectivity (higher effects on females). Findings suggest sex-dependent mechanisms where both chronic- ethanol intake and stress affect brain plasticity during neurodevelopment. Understanding these region-specific vulnerabilities is crucial for addressing alcohol use disorders and stress-related neuropathology.

Variable processing shifts during perceptual acceleration: Evidence from temporal integration
The perception of a stimulus can be accelerated by another that precedes it. Research to date has focused on quantifying this acceleration, and localizing it in the chain of perceptual and cognitive processes that are involved. This is challenging, because these processes may interact unexpectedly, and because traditional (univariate) analyses of brain activity and behaviour may conflate processes with the representations they act on. By using multivariate pattern analysis of EEG data from a missing element task, designed to measure the visual temporal integration of two successive stimulus displays, we were able to track the representation associated with the integrated percept. We manipulated the delay between our displays, and observed commensurate acceleration of the resultant integrated representation. Furthermore, regardless of the delay, we found that although processing was already accelerated during the earliest processing stages at around 100ms after stimulus onset, intermediate stages, at around 200ms, were even more accelerated. In contrast, later processing stages, at around 400ms, again showed less acceleration. The results thus suggest that perceptual acceleration during temporal integration is nonlinear, and that some time that is gained at one moment in the process can be lost again at another.

Modeling cerebral development in vitro with L-MYC-immortalized human neural stem cell-derived organoids
A promising advance for ex vivo studies of human brain development and formulation of therapeutic strategies has been the adoption of brain organoids that, to a greater extent than monolayer or spheroid cultures, recapitulate to varying extents the patterns of tissue development and cell differentiation of human brain. Previously, such studies been hampered by limited access to relevant human tissue, inadequate human in vitro models, and the necessity of using rodent models that imperfectly reproduce human brain physiology. Here we present a novel organoid-based research platform utilizing L-MYC-immortalized human fetal neural stem cells (LMNSC01) grown in a physiological 4% oxygen environment. We visualized developmental processes in LMNSC01 brain organoids for over 120 days in vitro by immunofluorescence and NanoString gene expression profiling. Gene expression patterns revealed by NanoString profiling were quantitatively compared to those occurring during normal brain development (BrainSpan database) using the Singscore method. We observe similar developmental patterns in LMNSC01 organoids and developing cortex for genes characterizing neurons, astrocytes, and oligodendrocytes, and multiple pathways including those involved in apoptosis, neuronal cytoskeleton, neurotransmission, and metabolism. Notable properties of this LMNSC01 platform are its initiation with immortalized authentic human neural stem cells, growth in a physiological oxygen environment, the consistency of the organoids produced, and favorable comparison of their gene expression patterns with those reported for normal cortical development.

Neurod2/6 transcription factors control basal progenitor differentiation and sequential production of neocortical cell subtypes
Neurod2 and Neurod6 (Neurod2/6) are key regulators to promote neuron differentiation and maturation during cerebral cortical development, but the cellular mechanism underlying their regulation is still unclear. As Neurod2/6 share similar expression and redundant regulatory elements, we took advantage of the mouse model deficient for both Neurod2/6 to investigate their roles in cortical neurogenesis. Here we reveal that the differentiation of Tbr2-positive (Tbr2+) basal progenitors (BPs), but not Pax6+ apical progenitors, was severely defected, resulting in ectopically expanded BPs in perinatal Neurod2/6 deficient brains. The sequential fate specification of cortical neurons was also impaired in the absence of Neurod2/6. Additionally, ectopic Tbr2+ BPs expressed multiple proliferation markers and were able to self-renew. Olig2+ glial precursors were consequently over-produced in Neurod2/6 deficient brains. Restoration of Neurod2/6 in the double deficient brains using in utero electroporation downregulated Tbr2 expression, and exhibited substantial rescue effects on defected laminar subtype specification and excessive gliogenesis by promoting BP differentiation. Our work indicates that Neurod2/6 regulate BP differentiation and sequential production of cortical cell subtypes via inhibiting Tbr2-dependent genetic program.

Neither exogenous, nor endogenous: evidence for a distinct role of negative emotion during attentional control
Negative or threatening stimuli capture attention. However, it remains unclear whether this phenomenon is best conceived as bottom-up (i.e. salience-driven) or top-down (i.e. goal-directed) instead. To address this question, we conducted two experiments using a previously validated dot-probe task (DPT) where physical salience (i.e. abrupt luminance change) and negative emotion (i.e. fearful face) competed with one another for attention selection (Experiment 1, n = 40) or negative (but also positive) emotion could be used as an endogenous cue by the participants to guide this process (Experiment 2, n = 39). Eye-tracking was used to ascertain that both cue and target were processed with peripheral vision. In Experiment 1, we found that negative emotion and physical salience both drove spatial attention in a bottom-up manner, yet their effects were under-additive, suggesting that they could mutually inhibit each other. Moreover, the results of Experiment 2 showed that fear, unlike happiness, could bias spatial attention in a top-down manner, yet only when participants were aware of the association created between the emotional cue and targets location at the block level. Combined together, these novel findings suggest that negative value does not influence the priority map independently from physical salience and goal but depending on the specific combination of cues available for attention selection in the environment, it acts either as an exogenous or endogenous cue, thereby revealing an enhanced flexibility for it.

Mixed Selectivity of Subthalamic Nucleus Neurons in Encoding Motor and Reward Behaviors
The subthalamic nucleus (STN) plays a critical role in modulating motor and cognitive functions within the basal ganglia, with its dysfunction being implicated in movement disorders such as Parkinson's disease. However, the behavioral representations of individual STN neurons remain incompletely understood. Using in vivo two-photon calcium imaging in behaving mice, we systematically mapped the activity of single STN neurons across diverse behavioral contexts, including locomotion, licking, and reward-driven actions. Our findings reveal that STN neurons exhibit mixed selectivity, encoding multiple behaviors with distinct temporal dynamics and excitatory or inhibitory response patterns. This mixed selectivity allows the STN to robustly encode motor parameters such as locomotion speed and licking intensity while integrating contextual information from different behavioral states. Comparisons with the adjacent zona incerta (ZI) revealed distinct encoding properties: while both regions represent locomotion, STN neurons more faithfully track motor states, whereas ZI neurons exhibit prolonged calcium events with weaker movement correlations. Population-level analysis showed STN activity in a low-dimensional neural manifold, with components linked to movement velocity and licking intensity. Notably, locomotion encoding in STN was context-dependent, diverging when movements were internally generated versus reward-modulated. Together, these findings highlight the specialized yet flexible role of the STN in integrating motor and reward-related signals, supporting a framework in which STN neurons contribute to motor control through multiplexed and context-dependent encoding. This work provides new insights into the functional organization of basal ganglia circuits and has implications for understanding STN's role in both physiological and pathological conditions.

Glucocorticoids modulate expression of perineuronal net component genes and parvalbumin during development of mouse cortical neurons
Severe prenatal maternal stress is a risk factor for schizophrenia in offspring. Since parvalbumin-containing GABAergic interneuron function in cortex and hippocampus is compromised in schizophrenia, and perineuronal nets (PNNs) facilitate the functioning of these cells, we tested the hypothesis that glucocorticoids, as stress mediators that can access the foetal compartment, might influence the expression of PNN component genes. In cultured mouse cortical neurons, we detected effects of hydrocortisone on many PNN component genes, via diverse mechanisms. A rapid (<4h), glucocorticoid receptor (GR)-mediated suppression of neurocan and hyaluronan synthase (Has) 1 and 3 mRNAs was observed at 7 days in vitro (DIV), whereas at 14DIV, brevican and versican expression was reduced by hydrocortisone without GR involvement, while GR inhibition elevated Has1 and Has2 mRNA levels and suppressed aggrecan mRNA levels. Tenascin R expression was rapidly suppressed by hydrocortisone at 7DIV but not at 14. At 21DIV, PNN component gene expression had become insensitive to hydrocortisone, although parvalbumin expression was reduced after 24h but not 4h exposure. Additionally, effects on protein levels were observed that were sometimes consistent with the mRNA changes (e.g. Has3, Gad1) and sometimes unrelated to them (e.g. elevated TnR levels at 14DIV after glucocorticoid receptor antagonism). We found that hydrocortisone could directly inhibit proteasome activity, potentially explaining the ability of hydrocortisone to increase Has2 levels. As expected from these results, the overall structure of the PNN was compromised by hydrocortisone exposure, with the length of proximal dendrite covered by PNN being reduced. Overall, the data demonstrate a complex and profound, but developmental stage-dependent, regulation of PNN component gene expression by glucocorticoids. This may contribute to the action of severe prenatal or perinatal stress to increase schizophrenia risk.

A paradigm to study the learning of muscle activity patterns outside of the natural repertoire
The acquisition of novel muscle activity patterns is a key aspect of motor skill learning which can be seen at play, for example, when beginner musicians learn new guitar or piano chords. To study this process, here we introduce a new paradigm that requires quick and synchronous flexion and extension of multiple fingers. First, participants practiced all the 242 possible combinations of isometric finger flexion and extension around the metacarpophalangeal joint (i.e., chords). We found that some chords were initially extremely challenging, but participants could eventually achieve them with practice, showing that the initial difficulty did not reflect hard biomechanical constraints imposed by the interaction of tendons and ligaments. In a second experiment we found that chord learning was largely chord-specific and did not generalize to untrained chords. Finally, we explored which factors make some chords more difficult than others. Difficulty was well predicted by the muscle activity pattern required by that chord. Interestingly, difficulty related to how similar chords' muscle activity patterns are to the muscle activity patterns required by everyday hand use, and to the overall size of the muscle activity. Together, our results suggest that the new paradigm introduced in this work may provide a valuable tool to study the neural processes underlying the acquisition of novel muscle activity patterns in the human motor system.

Integrated Cross-Disease Atlas of Human And Mouse Astrocytes Reveals Heterogeneity and Conservation of Astrocyte Subtypes in Neurodegeneration
Astrocytes play a pivotal role in central nervous system homeostasis and neuroinflammation. Despite advancements in single-cell analyses, the heterogeneity of reactive astrocytes in neurodegenerative diseases, particularly across species, remains understudied. Here, we present an integrated atlas of 187,000 astrocytes from mouse models of Alzheimer's (AD) and multiple sclerosis (MS) alongside 438,000 astrocytes from AD, MS, and Parkinson's (PD) patients. Our analysis identified four distinct mouse astrocyte clusters, including two disease-associated astrocyte (DAA) clusters, DAA1 and DAA2. DAA1 displayed reactivity resembling responses to acute stimuli, including endotoxemia, while DAA2 expressed well-known AD risk genes. In an AD model, DAA1 and DAA2 exhibited distinct spatial relationships to amyloid plaques. In humans, we identified eight distinct astrocyte clusters, encompassing homeostatic and disease-associated subtypes. Cross-species analysis linked disease-associated clusters while also highlighting divergent expression in others. Our astrocyte atlas is available through a user-friendly, searchable website: http://research-pub.gene.com/AstroAtlas/.

Nano-organization of synapses defines synaptic release properties at cortical neuron dendritic spines
Visualization of the submicron organization of excitatory synapses has revealed an unexpectedly ordered architecture consisting of nanocolumns of synaptic proteins that group into nanomodules which scale in number as spine size increases. How these features are related to synaptic function has remained unclear. Here, using super-resolution followed by live-cell line-scan imaging, we find that the size of the smallest miniature calcium and glutamate events are the same, regardless of whether spines have one or two nanopuncta of PSD-95, and that miniature synaptic response in all spines are best fit by a three term Poisson. Two nanomodule spines exhibit more large events without a significant change in event frequency, with the number of the largest events increasing disproportionately. These data support a model where nanomodules define sites of synaptic release and where the nanoarchitecture of synaptic proteins specifies subtypes of excitatory synapses, with increasing numbers of nanomodules increasing coordinated multivesicular release.

Dynamics of infant white matter maturation from birth to 6 months
The first months after a baby's birth encompass the most rapid period of postnatal change in the human lifespan, but longitudinal trajectories of white matter maturation in this period remain uncharted. Using densely sampled diffusion tensor images collected longitudinally at a mean rate of 1 scan per 1.55 days, we measured non-linear growth and growth rate trajectories of major white matter tracts from birth to 6 months. Growth rates at birth were 6 to 11 times faster than at 6 months, with tracts less mature at birth developing fastest. When matched on chronological age, shorter gestation infants had less mature white matter at birth but faster growth rates than their longer gestation peers; however, growth trajectories were highly similar when corrected for gestational age. This is the first study to estimate white matter trajectories using dense sampling in the first 6 post-natal months, which can inform the study of neurodevelopmental disorders beginning in infancy.

Informational Complexity as a Neural Marker of Cognitive Reserve
In Alzheimer's disease (AD), a mismatch between neurological damage and cognitive functioning often is attributed to individual differences in cognitive reserve. Understanding the neural mechanism of cognitive reserve could help assessing the therapeutic effectiveness of interventions in AD. To address this, here, 38 elderly participants performed a sustained attention task during high-density EEG while awake and during drowsiness. Operationally, the degree to which performance was impaired under drowsiness signalled the extent of cognitive reserve, with less impairment indicating a higher level of cognitive reserve. Investigating performance variations during the active management of neural challenges offers a novel approach to studying cognitive reserve, capturing dynamics that mirror everyday cognitive demand. We related cognitive reserve to various measures, including informational complexity using the Lempel-Ziv (LZSUM) algorithm. We found a significant interaction effect between arousal and performance, where LZSUM values increased in high performers when drowsy but decreased in low performers. This effect was most pronounced in the frontal and central areas. Our findings suggest LZSUM to be indicative of a compensatory mechanism and thus show potential for LZSUM as a neural marker in assessing cognitive reserve. However, we found no consistent relationship between performance and structural brain measures, and proxies of cognitive reserve. Critically, our findings present a counterexample to the prevailing view that informational complexity purely reflects conscious level. Further research, such as a study with the same paradigm in patients with mild cognitive impairment (MCI) and AD, may lead to additional insights of whether we are truly measuring cognitive reserve.

Field-Programmable Gate Array Based Ultra-Low Power Discrete Fourier Transforms for Closed-Loop Neural Sensing
Digital implementations of discrete Fourier transforms (DFT) are a mainstay in feature assessment of recorded biopotentials, particularly in the quantification of biomarkers of neurological disease state for adaptive deep brain stimulation. Fast Fourier transform (FFT) algorithms and architectures present a substantial power demand from onboard batteries in implantable medical devices, necessitating the development of ultra-low power Fourier transform methods in resource-constrained environments. Numerous FFT architectures aim to optimize power and resource demand through computational efficiency; however, prioritizing the reduction of logic complexity at the cost of additional computations can be equally or more effective. This paper introduces a minimal architecture single-delay feedback discrete Fourier transform (mSDF-DFT) for use in ultra-low-power field programmable gate array applications and shows energy and power improvements over state-of-the-art FFT methods. We observe a 33% reduction in dynamic power and 4% reduction in resource utilization in a neural sensing application when compared to state-of-the-art FFT algorithms. While designed for use in closed-loop deep brain stimulation and medical device implementations, the mSDF-DFT is also easily extendable to any ultra-low power embedded application.

Evidence accumulation from experience and observation in the cingulate cortex
We use our experiences to form and update beliefs about the hidden states of the world. When possible, we also gather evidence by observing others. However, how the brain integrates experiential and observational evidence is not understood. We studied the dynamics of evidence integration in a two-player game with volatile hidden states. Both humans and monkeys successfully updated their beliefs while playing the game and observing their partner, though less effectively when observing. Electrophysiological recordings in animals revealed that the anterior cingulate cortex (ACC) integrates independent sources of experiential and observational evidence into a coherent neural representation of dynamic belief about the environment's state. The geometry of population activity revealed the computational architecture of this integration and provided a neural account of the behavioral asymmetry between experiential and observational evidence accumulation. This work lays the groundwork for understanding the neural mechanisms underlying evidence accumulation in social contexts within the primate brain.

Phasic and tonic pain serve distinct functions during adaptive behaviour
Pain drives self-protective behaviour, and evolutionary theories suggest it acts over different timescales to serve distinct functions. Whilst phasic pain provides a teaching signal to drive avoidance of new injury, tonic pain is argued to support recuperative behaviour, for instance by reducing motivational vigour. We test this hypothesis in an immersive virtual reality EEG foraging task where subjects harvested fruit in a forest: some fruit elicited brief phasic pain to the grasping hand, and this reduced choice probability. Simultaneously, tonic pressure pain to the contralateral upper arm was associated with reduced action velocities. This could be explained by a free-operant computational framework that formalises and quantifies the function of tonic and phasic pain in terms of motivational vigour and decision value, and model parameters correlated with EEG responses. Overall, the results show how tonic and phasic pain subserve distinct objective motivational functions that support harm minimisation during ongoing adaptive behaviour.

Juggling the Limits of Lucidity: Searching for Cognitive Constraints in Dream Motor Practice
Lucid dreaming (LD), during which the dreamer becomes aware of the dream state, offers a unique opportunity for a variety of applications, including motor practice, personal well-being, and nightmare therapy. However, these applications largely depend on a dreamer's ability to control their dreams. While LD research has traditionally focused on induction techniques to increase dream frequency, the equally crucial skill of dream control remains underexplored. This study provides an initial investigation into the mechanisms of dream control and its potential influencing factors. We specifically examined whether a complex motor skill -juggling- could be performed within a lucid dream, creating a particularly challenging lucid dream task, which calls for a high level of dream control. Eight healthy participants (aged 24-50) underwent overnight polysomnography (PSG) at the University of Bern's Institute for Sports Science, provided detailed dream reports, and completed questionnaires assessing dream control, self-efficacy, personality traits, mindfulness, motivation, and intention setting. Of these, four participants experienced lucid dreams, and of these, two demonstrated high dream control with successful LD juggling attempts. Trait differences between non-lucid and lucid dreamers in the lab were examined, with a focus on low-to-no dream control versus high dream control among the lucid dreamers. The two lucid dream juggling attempts are described in detail, providing insight into the challenges of executing complex tasks within a lucid dream. While this study lacks in sample size, it highlights the potential roles of many psychological traits, such as belief, motivation, and self-efficacy, in shaping dream control abilities. This study helps to lay the groundwork for future research aimed at investigating lucid dream control and therefore optimizing LD applications in therapy, sports training, and cognitive science.

Transcriptomic analysis of C9orf72-mutated iPSC-derived microglia implicates cell-autonomous upregulation of selected NLRP3 inflammasome genes in motor neuron degeneration
Hexanucleotide repeat expansion (HRE) in the non-coding region of the gene C9orf72 is the most prevalent mutation in Amyotrophic Lateral Sclerosis (ALS) and frontotemporal dementia. The C9orf72 HRE contributes to motor neuron degeneration in ALS both through cell-autonomous mechanisms and non-cell autonomous disease processes involving glial cells such as microglia. The molecular mechanisms by which C9orf72 HRE microglia contribute to motor neuron death in ALS remain to be fully elucidated. In this study, we generated microglia from human C9orf72 HRE and isogenic iPSCs using three different microglia derivation methods that can generate microglia with distinct intrinsic inflammatory profiles. RNA sequencing analysis reveals a consistent dysregulation of specific genes involved in pathways underlying NLRP3 inflammasome activation in C9orf72 HRE microglia independent of priming with any activating cues. In agreement with elevated expression of NLRP3 inflammasome components, conditioned media from C9orf72 HRE microglia enhance the death of C9orf72 HRE motor neurons implicating microglia-secreted molecules in non-cell autonomous mechanisms of C9orf72 HRE pathology. These findings provide evidence that C9orf72 HRE microglia have a cell-autonomous pro-inflammatory phenotype that contributes to non-cell autonomous mechanisms of motor neuron degeneration in ALS through dysregulated expression of specific genes involved in NLRP3 inflammasome-mediated mechanisms.

A zebrafish circuit for behavioral credit assignment
An important question in systems neuroscience is how the brain links choices and their outcomes to modify future behavior. Despite being a fundamental computation, we lack explicit circuit-level models of how this is achieved. Here, we provide evidence for a cellular substrate underlying this computation in a vertebrate brain. We develop an operant thermoregulatory assay in larval zebrafish, and show that adaptive responses depend on the precise temporal coincidence between choices and contingent thermal feedback. Chemogenetic ablations reveal that the dorsal habenula (dHb), the homolog of the mammalian medial habenula, plays an essential role in this adaptive behavior. Functional and anatomical mapping of the downstream circuitry reveals that the intermediate interpeduncular nucleus (iIPN) integrates choice- and temperature-related signals, conveyed by the dHb and a prepontine hindbrain nucleus, respectively. While dHb axons exhibit a diffuse connectivity pattern, hindbrain neurons project onto specific, lateralized iIPN regions. We show that these iIPN glomeruli function as sensorimotor detectors, integrating choices and outcomes. Specifically, when a swim in a given direction is closely followed by a temperature change, iIPN activity shows increased responses. We further show that this coincidence detection, with a precise temporal profile, links an action to its subsequent sensory feedback, relies on a presynaptic mechanism within the terminal segments of dHb axons. This mechanism is mediated by GABAB-dependent modulation of dHb sensory axons by choice-related prepontine GABAergic neurons. In this study we are therefore able to link an adaptive behavior to aspecific circuit mechanism.