bioRxiv Subject Collection: Neuroscience's Journal

Mitochondrial Biomarkers and Metabolic Syndrome in Bipolar Disorder
Importance: Examining translatable mitochondrial blood-based biological markers to identify its association with metabolic diseases in bipolar disorder. Objective: To test whether mitochondrial metabolites, mainly lactate, and cell-free circulating mitochondrial DNA are associated with markers of metabolic syndrome in bipolar disorder, hypothesizing higher lactate but unchanged cell-free circulating mitochondrial DNA levels in bipolar disorder patients with metabolic syndrome. Design: In a cohort study, primary testing from the FondaMental Advanced Centers of Expertise for bipolar disorder was conducted, including baseline plasma samples and blinded observers for all experimentation and analysis. Setting: The FondaMental Foundation coordinate a multicenter, multidisciplinary French networks aiming at creation of cohorts to improve identification of homogeneous subgroups of psychiatric disorders toward personalized treatments. Participants: The FACE-BD primary testing cohort includes 837 stable bipolar disorder patients. The I-GIVE validation cohort consists of 235 participants: stable and acute bipolar patients, non-psychiatric controls, and acute schizophrenia patients. Participants were randomly selected based on biosample availability. Exposures: All patients underwent the standard primary care within their center. No intentional exposures were part of this study. Main Outcome and Measures: The primary outcome modelled an association with lactate and metabolic syndrome in this population. Reflective a priori hypothesis. Results: Multivariable regression analyses show lactate association with triglycerides (Est= 0.072(0.023), p = 0.0065,), fasting glucose (Est = 12(0.025), p= 0.000015) and systolic (Est= 0.003(0.0013), p= 0.031) and diastolic blood pressure (Est = 0.0095(0.0017), p= 1.3e-7). Significantly higher levels of lactate were associated with presence of metabolic syndrome (Est = 0.17(0.049), p=0.00061) after adjusting for potential confounding factors. Mitochondrial-targeted metabolomics identified distinct metabolite profiles in patients with lactate presence and metabolic syndrome, differing from those without lactate changes but with metabolic syndrome. Circulating cell-free mitochondrial DNA was not associated with metabolic syndrome. Conclusion & Relevance: This thorough analysis mitochondrial biomarkers indicate the associations with lactate and metabolic syndrome, whereas circulating cell-free mitochondrial DNA is limited in the context of metabolic syndrome. This study is relevant to improve the identification and stratification of bipolar patients with metabolic syndrome and provide potential personalized-therapeutic opportunities.

Long-range action of an HDAC inhibitor treats chronic pain in a spared nerve injury rat model
Histone deacetylase inhibitors (HDACi) that modulate epigenetic regulation and are approved for treating rare cancers have, in disease models, also been shown to mitigate neurological conditions, including chronic pain. They are of interest as non-opioid treatments, but achieving long-term efficacy with limited dosing has remained elusive. Here we utilize a triple combination formulation (TCF) comprised of a pan-HDACi vorinostat (Vo at its FDA-approved daily dose of 50mg/Kg), the caging agent 2-hydroxypropyl-{beta}-cyclodextrin (HPBCD) and polyethylene glycol (PEG) known to boost plasma and brain exposure and efficacy of Vo in mice and rats, of various ages, spared nerve injury (SNI) model of chronic neuropathic pain. Administration of the TCF (but not HPBCD and PEG) decreased mechanical allodynia for 4 weeks without antagonizing weight, anxiety, or mobility. This was achieved at less than 1% of the total dose of Vo approved for 4 weeks of tumor treatment and associated with decreased levels of major inflammatory markers and microglia in ipsilateral (but not contralateral) spinal cord regions. A single TCF injection was sufficient for 3-4 weeks of efficacy: this was mirrored in repeat injections, specific for the injured paw and not seen on sham treatment. Pharmacodynamics in an SNI mouse model suggested pain relief was sustained for days to weeks after Vo elimination. Doubling Vo in a single TCF injection proved effectiveness was limited to male rats, where the response amplitude tripled and remained effective for > 2 months, an efficacy that outperforms all currently available chronic pain pharmacotherapies. Together, these data suggest that through pharmacological modulation of Vo, the TCF enables single-dose effectiveness with extended action, reduces long-term HDACi dosage, and presents excellent potential to develop as a non-opioid treatment option for chronic pain.

Magnetogenetic stimulation inside MRI induces spontaneous and evoked changes in neural circuits activity in rats
The ability to modulate specific neural circuits and simultaneously visualize and measure brain activity with MRI would greatly impact understanding brain function in health and disease. The combination of neurostimulation methods and MRI in animal models have already shown promise in elucidating fundamental mechanisms associated with brain activity. We developed an innovative magnetogenetics neurostimulation technology that can trigger neural activity through magnetic fields. Similar to other genetic-based neuromodulation methods, magnetogenetics offers cell-, area- and temporal-specific control of neural activity. However, the magnetogenetics protein (Electromagnetic Preceptive Gene (EPG)) are activated by non-invasive magnetic fields, providing a unique way to target neural circuits by the MRI gradients while simultaneously measure their effect on brain activity. EPG was expressed in rat visual cortex and the amplitude of low-frequency fluctuation (fALFF), resting-state functional connectivity (FC), and sensory activation was measured using a 7T MRI. The results demonstrate that EPG-expressing rats had significantly higher signal fluctuations in the visual areas and stronger FC in sensory areas consistent with known anatomical visuosensory and visuomotor connections. This new technology complements the existing neurostimulation toolbox and provides a mean to study brain function in a minimally-invasive way which was not possible previously.

Neurexins control the strength and precise timing of glycinergic inhibition in the auditory brainstem
Neurexins play diverse functions as presynaptic organizers in various glutamatergic and GABAergic synapses. However, it remains unknown whether and how neurexins are involved in shaping functional properties of the glycinergic synapses, which mediate prominent inhibition in the brainstem and spinal cord. To address these issues, we examined the role of neurexins in a model glycinergic synapse between the principal neuron in the medial nucleus of the trapezoid body (MNTB) and the principal neuron in the lateral superior olive (LSO) in the auditory brainstem. Combining RNAscope with stereotactic injection of AAV-Cre in the MNTB of neurexin1/2/3 conditional triple knockout mice, we showed that MNTB neurons highly express all isoforms of neurexins although their expression levels vary remarkably. Selective ablation of all neurexins in MNTB neurons reduced not only the amplitude but also the kinetics of the glycinergic synaptic transmission at LSO neurons. The synaptic dysfunctions were primarily caused by impaired Ca2+ sensitivity of release and tightness of coupling between voltage-gated Ca2+ channels and synaptic vesicles. Together, our current findings demonstrate that neurexins are essential in controlling the strength and temporal precision of the glycinergic synapse, which therefore corroborates the role of neurexins as key presynaptic organizers in all major types of fast chemical synapses.

An in vitro model of acute horizontal basal cell activation reveals dynamic gene regulatory networks underlying the acute activation phase
Horizontal basal cells (HBCs) activate only in response to severe olfactory epithelium (OE) injury. This activation is mediated by the loss of the transcription factor TP63. Using the compound phorbol 12-myristate 13-acetate (PMA), we find that we can model the process of acute HBC activation. First, we find that PMA treatment induces a rapid loss in TP63 protein and induces the expression of HOPX and the nuclear translocation of RELA, previously identified to mediate HBC activation. Using bulk RNA sequencing, we find that PMA-treated HBCs pass through various stages of acute activation identifiable by transcriptional regulatory signatures that mimic stages identified in vivo. These temporal stages are associated with varying degrees of engraftment and differentiation potential in transplantation assays. Together, this data shows that our model can model physiologically relevant features of HBC activation and identifies new candidates for mechanistic testing.

Assessing adverse effects and unspecific effects of transcutaneous spinal direct current stimulation (tsDCS)
Background: Transcutaneous spinal direct current stimulation (tsDCS) is a relatively recent method for non-invasively modulating neuronal activity in the human spinal cord. Despite its growing prominence, comprehensive studies addressing its potential adverse effects (AEs) and unspecific effects (UEs) are lacking. Objective: In this study, we conducted a systematic investigation into the potential AEs and UEs of tsDCS in healthy volunteers. Methods: We used a randomized double-blind within-participant design, employing anodal, cathodal and sham tsDCS of the thoracolumbar spinal cord. Our approach involved a newly-developed structured questionnaire (to assess subjectively-reported AEs) in combination with tsDCS-concurrent recording of skin conductance, cardiac and respiratory activity (to assess UEs in bodily state). Results: The most frequently participant-reported AEs were sensations of burning, tingling, and itching, although they were largely described as mild; skin redness (experimenter-reported) occurred even more frequently. Importantly, when comparing AEs between active and sham tsDCS via frequentist and Bayesian analysis approaches, the results were largely in favour of no difference between conditions (with the exception of skin redness). A similar picture emerged for most UE metrics, suggesting that tsDCS does not induce changes in bodily state, at least as measured by our autonomic nervous system metrics. Conclusion: We believe that the strategy employed here could serve as a starting point for a systematic AE and UE assessment in clinical populations, longitudinal designs and when stimulating different spinal sites. Taken together, our results contribute to assessing the tolerability, safety and specificity of tsDCS, in order to further the investigation of spinal cord function in health and disease.

Memory reactivation during sleep does not act holistically on object memory
Memory reactivation during sleep is thought to facilitate memory consolidation. Most sleep reactivation research has examined how reactivation of specific facts, objects, and associations benefits their overall retention. However, our memories are not unitary, and not all features of a memory persist in tandem over time. Instead, our memories are transformed, with some features strengthened and others weakened. Does sleep reactivation drive memory transformation? We leveraged the Targeted Memory Reactivation technique in an object category learning paradigm to examine this question. Participants learned three categories of novel objects, where each object had unique, distinguishing features as well as features shared with other members of its category. We used a real-time EEG protocol to cue the reactivation of these objects during sleep at moments optimized to generate reactivation events. We found that reactivation improved memory for distinguishing features while worsening memory for shared features, suggesting a differentiation process. The results indicate that sleep reactivation does not act holistically on object memories, instead supporting a transformation process where some features are enhanced over others.

Striatal dopamine reflects individual long-term learning trajectories
Learning from naive to expert occurs over long periods of time, accompanied by changes in the brain's neuronal signals. The principles governing behavioural and neuronal dynamics during long-term learning remain unknown. We developed a psychophysical visual decision task for mice that allowed for studying learning trajectories from naive to expert. Mice adopted sequences of strategies that became more stimulus-dependent over time, showing substantial diversity in the strategies they transitioned through and settled on. Remarkably, these transitions were systematic; the initial strategy of naive mice predicted their strategy several weeks later. Longitudinal imaging of dopamine release in dorsal striatum demonstrated that dopamine signals evolved over learning, reflecting stimulus-choice associations linked to each individual's strategy. A deep neural network model trained on the task with reinforcement learning captured behavioural and dopamine trajectories. The model's learning dynamics accounted for the mice's diverse and systematic learning trajectories through a hierarchy of saddle points. The model used prediction errors mirroring recorded dopamine signals to update its parameters, offering a concrete account of striatal dopamine's role in long-term learning. Our results demonstrate that long-term learning is governed by diverse yet systematic transitions through behavioural strategies, and that dopamine signals exhibit key characteristics to support this learning.

'Are you even listening?' - EEG-based detection of absolute auditory attention to natural speech with application to neuro-steered hearing devices
In this study, we use electroencephalography (EEG) recordings to perform absolute auditory attention detection (aAAD), i.e., determine whether a subject is actively listening to a presented speech stimulus or not. More precisely, we aim to discriminate between an active listening condition, and a distractor condition where subjects passively listen to the speech stimulus while performing another cognitive task. To this end, we re-use an existing EEG dataset where the subjects watch a silent movie as a distractor condition, and introduce a new EEG dataset with two other distractor conditions (silently reading a text and performing arithmetic exercises). We focus on two EEG features, namely neural envelope tracking (NET) and spectral entropy (SE). We find significantly higher NET and lower SE in the active listening condition compared to the distractor conditions, which for the SE is the reverse of what was previously found for an active listening versus passive listening condition (without any distractors). In addition, aAAD is used in the context of a selective auditory attention decoding (sAAD) task, where the goal is to decode to which of two competing speakers the subject is attending, which is a core task in the context of so-called neuro steered hearing devices. We show that evaluating sAAD performance only on segments of active listening improves sAAD performance when detecting these active listening segments as having higher NET, whereas the reverse trend is observed when detecting these segments as having lower SE. We conclude that NET is a more reliable metric for aAAD as it is consistently higher for the active listening condition, whereas the relation of the SE between the active listening and passive listening conditions seems to depend on the nature of the distractor task. Consequently, NET shows the most promise for aAAD and to detect auditory inattentive segments in neuro-steered hearing devices.

Gene replacement therapy for Lafora disease in the Epm2a-/- mouse model
Lafora disease is a rare and fatal form of progressive myoclonic epilepsy typically occurring early in adolescence. Common symptoms include seizures, dementia, and a progressive neurological decline leading to death within 5-15 years from onset. The disease results from mutations transmitted with autosomal recessive inheritance in the EPM2A gene, encoding laforin, a dual-specificity phosphatase, or the EPM2B gene, encoding malin, an E3-ubiquitin ligase. Laforin has glucan phosphatase activity, is an adapter of enzymes involved in glycogen metabolism, is involved in endoplasmic reticulum-stress and protein clearance, and acts as a tumor suppressor protein. Laforin and malin work together in a complex to control glycogen synthesis and prevent the toxicity produced by misfolded proteins via the ubiquitin-proteasome system. Disruptions in either protein can lead to alterations in this complex, leading to the formation of Lafora bodies that contain abnormal, insoluble, and hyperphosphorylated forms of glycogen called polyglucosans. We used the Epm2a-/- knock-out mouse model of Lafora disease to apply a gene replacement therapy by administering intracerebroventricular injections of a recombinant adeno-associated virus carrying the human EPM2A gene. We evaluated the effects of this treatment by means of neuropathological studies, behavioral tests, video-electroencephalography recording, and proteomic/phosphoproteomic analysis. Gene therapy with recombinant adeno-associated virus containing the EPM2A gene ameliorated neurological and histopathological alterations, reduced epileptic activity and neuronal hyperexcitability, and decreased the formation of Lafora bodies. Differential quantitative proteomics and phosphoproteomics revealed beneficial changes in various molecular pathways altered in Lafora disease. Improvements were observed for up to nine months following a single intracerebroventricular injection. In conclusion, gene replacement therapy with human EPM2A gene in the Epm2a-/- knock-out mice shows promise as a potential treatment for Lafora disease.

Common anesthetic used in preclinical PET imaging inhibits metabolism of the PET tracer 3F4AP
PET imaging studies in laboratory animals are almost always performed under isoflurane anesthesia to ensure that the subject stays still during the image acquisition. Isoflurane is effective, safe, and easy to use, and it is generally assumed to not have an impact on the imaging results. Motivated by marked differences observed in [18F]3F4AP brain uptake and metabolism between human and nonhuman primate studies, this study investigates the possible effect of isoflurane on [18F]3F4AP metabolism and brain uptake. Isoflurane was found to largely abolish tracer metabolism in mice resulting in a 3.3-fold higher brain uptake in anesthetized mice at 35 min post radiotracer administration, which replicated the observed effect in unanesthetized humans and anesthetized monkeys. This effect is attributed to isoflurane's interference in the CYP2E1-mediated breakdown of [18F]3F4AP, which was confirmed by reproducing a higher brain uptake and metabolic stability upon treatment with the known CYP2E1 inhibitor disulfiram. These findings underscore the critical need to examine the effect of isoflurane in PET imaging studies before translating tracers to humans that will be scanned without anesthesia.

Hidden hearing loss in a hereditary peripheral neuropathy: evidence from a mouse model of Charcot Marie Tooth type 1A.
Hidden hearing loss (HHL), a recently described auditory neuropathy characterized by normal audiometric thresholds but reduced sound-evoked potentials, has been proposed to underlie hearing difficulties in noisy environments. Animal studies showing that HHL is associated with loss of inner hair cell (IHC) synapses in response to mild noise exposures and aging led to most studies of HHL focusing on IHC synaptopathy. More recently we showed that transient auditory nerve (AN) demyelination also causes HHL but without affecting IHC synapses, suggesting that demyelinating peripheral neuropathies could be a second HHL mechanism. To test this in a clinically relevant model, we studied a mouse model of Charcot-Marie-Tooth type 1A (CMT1A), the most prevalent hereditary peripheral neuropathy in humans. CMT1A mice exhibit HHL, i.e., normal auditory threshold but reduced amplitudes and longer latencies of sound-evoked compound action potential. The AN structural phenotypes of CMT1A mice are also remarkably similar to those found in mice with transient demyelination, i.e., disorganization of AN heminodes near the IHCs with minor loss of AN fibers. Our results support the hypothesis that mild disruptions of AN myelination can cause HHL and that heminodal defects contribute to the alterations in action potential amplitudes and latencies seen in these models. Also, these findings suggest that patients with CMT1A or other peripheral neuropathies like Guillain-Barre Syndrome, are likely to suffer from HHL. Furthermore, these results suggest that studies of hearing in CMT1A patients might help in the development of robust clinical tests for HHL, which are currently lacking.

Corticopostural functional and effective connectivity reveal cortical control of postural sway velocity during quiet standing
Background: Despite a large body of evidence showing the involvement of the sensorimotor cortex in postural control, its exact role remains unclear. Models of postural control outcomes suggested that the velocity of the center of pressure is a crucial parameter to maintain balance. Inspired by corticokinematic coherence, we hypothesized that cortical oscillations and the velocity of the center of pressure (CoP) would synchronize and that this synchronization would increase with postural task difficulty during quiet standing. Methods: We compared the magnitude of coherence and Granger causality computed between brain oscillations recorded with electroencephalography and the center of pressure velocity in the Delta and Theta frequency bands obtained from 23 participants performing four quiet standing tasks with various levels of difficulty. The effect of postural task difficulty and information flow direction were tested with a linear mixed model while non-parametric correlations were computed between coherence magnitude and postural performance measured by 95% confidence ellipse area and mean center of pressure velocity. Results: We found significant coherence between the Cz EEG electrode and CoP velocity in the Delta and Theta frequency bands. This EEG-CoP velocity coherence significantly increased with task difficulty in the Delta (F = 18.8, p < 0.001) and Theta (F = 7.83, p < 0.001) bands. Granger causality significantly increased with task difficulty (F = 12.5, p < 0.001) and was higher in the efferent than afferent direction (F = 78, p < 0.001). The 95% confidence ellipse area was correlated to coherence magnitude in the most difficult condition. Participants showing significant Granger causality in the afferent direction showed more stable postural outcomes. Conclusion: Our results confirm that the CoP velocity has a crucial role in postural control through its synchronization with sensorimotor cortex oscillations. The efferent information predominance suggests that posture is partly controlled by the sensorimotor cortex by a mechanism named corticopostural coherence. Our results show that this corticopostural coherence could represent a mechanism for controlling balance during quiet standing.

GSK3 inhibition reduces ECM production and prevents age-related macular degeneration-like pathology
Malattia Leventinese/Doyne Honeycomb Retinal Dystrophy (ML/DHRD) is an age-related macular degeneration (AMD)-like retinal dystrophy caused by an autosomal dominant R345W mutation in the secreted glycoprotein, fibulin-3 (F3). To identify new small molecules that reduce F3 production from retinal pigmented epithelium (RPE) cells, we knocked-in a luminescent peptide tag (HiBiT) into the endogenous F3 locus which enabled simple, sensitive, and high throughput detection of the protein. The GSK3 inhibitor, CHIR99021 (CHIR), significantly reduced F3 burden (expression, secretion, and intracellular levels) in immortalized RPE and non-RPE cells. Low-level, long-term CHIR treatment promoted remodeling of the RPE extracellular matrix (ECM), reducing sub-RPE deposit-associated proteins (e.g., amelotin, complement component 3, collagen IV, and fibronectin), while increasing RPE differentiation factors (e.g., tyrosinase, and pigment epithelium derived factor). In vivo, treatment of 8 mo R345W+/+ knockin mice with CHIR (25 mg/kg i.p., 1 mo) was well tolerated and significantly reduced R345W F3-associated AMD-like basal laminar deposit number and size, thereby preventing the main pathological feature in these mice. This is the first demonstration of small molecule-based prevention of AMD-like pathology in ML/DHRD mice and may herald a rejuvenation of interest in GSK3 inhibition for the treatment of neurodegenerative diseases, including, potentially AMD itself.

Acute Neuropixels recordings in the marmoset monkey
High-density linear probes, like Neuropixels, provide an unprecedented opportunity to understand how neural populations within specific laminar compartments contribute to behavior. Marmoset monkeys, unlike macaque monkeys, have a lissencephalic (smooth) cortex that enables recording perpendicular to the cortical surface, thus making them an ideal animal model for studying laminar computations. Here we present a method for acute Neuropixels recordings in the common marmoset (Callithrix jacchus). The approach replaces the native dura with an artificial silicon-based dura that grants visual access to the cortical surface, which is helpful in avoiding blood vessels, ensures perpendicular penetrations, and could be used in conjunction with optical imaging or optogenetic techniques. The chamber housing the artificial dura is simple to maintain with minimal risk of infection and could be combined with semi-chronic microdrives and wireless recording hardware. This technique enables repeated acute penetrations over a period of several months. With occasional removal of tissue growth on the pial surface, recordings can be performed for a year or more. The approach is fully compatible with Neuropixels probes, enabling the recording of hundreds of single neurons distributed throughout the cortical column.

Prenatal exposure to valproic acid reduces synaptic δ-catenin levels and disrupts ultrasonic vocalization in neonates
Valproic acid (VPA) is an effective and commonly prescribed drug for epilepsy and bipolar disorder. However, children born from mothers treated with VPA during pregnancy exhibit an increased incidence of autism spectrum disorder (ASD). Although VPA may impair brain development at the cellular level, the mechanism of VPA-induced ASD has not been completely addressed. A previous study has found that VPA treatment strongly reduces {delta}-catenin mRNA levels in cultured human neurons. {delta}-catenin is important for the control of glutamatergic synapses and is strongly associated with ASD. VPA inhibits dendritic morphogenesis in developing neurons, an effect that is also found in neurons lacking {delta}-catenin expression. We thus hypothesize that prenatal exposure to VPA significantly reduces {delta}-catenin levels in the brain, which impairs glutamatergic synapses to cause ASD. Here, we found that prenatal exposure to VPA markedly reduced {delta}-catenin levels in the brain of mouse pups. VPA treatment also impaired dendritic branching in developing mouse cortical neurons, which was reversed by elevating {delta}-catenin expression. Prenatal VPA exposure significantly reduced synaptic AMPA receptor levels and postsynaptic density 95 (PSD95) in the brain of mouse pups, indicating dysfunctions in glutamatergic synaptic transmission. VPA exposure also significantly altered ultrasonic vocalization (USV) in newly born pups when they were isolated from their nest. Moreover, VPA-exposed pups show impaired hypothalamic response to isolation, which is required to produce animals' USVs following isolation from the nest. Therefore, these results suggest that VPA-induced ASD pathology can be mediated by the loss of {delta}-catenin functions.

Neuroanatomical changes observed over the course of a human pregnancy
Pregnancy is a period of profound hormonal and physiological change experienced by millions of women annually, yet the neural changes unfolding in the maternal brain throughout gestation have not been studied in humans. Leveraging precision imaging, we mapped neuroanatomical changes in an individual from preconception through two years postpartum. Pronounced decreases in gray matter volume and cortical thickness paired with increases in white matter microstructure were evident across the brain, with few regions untouched by the transition to motherhood.

Engram Reactivation Mimics Cellular Signatures of Fear
Engrams, or the physical substrate of memory in the brain, recruit heterogeneous cell-types. Targeted reactivation of neurons processing discrete memories drives the behavioral expression of memory, though the underlying landscape of recruited cells and their real-time responses remain elusive. To understand how artificial stimulation of fear affects intra-hippocampal neuronal and astrocytic dynamics as well as their behavioral consequences, we expressed channelrhodopsin-2 in an activity-dependent manner in dentate gyrus neurons while performing fiber photometry of both cell types in ventral CA1 across learning and memory. Neurons and astrocytes were shock-responsive, while astrocytic calcium events were uniquely modulated by fear conditioning. Notably, optogenetic stimulation of a hippocampus-mediated engram recapitulated coordinated calcium signatures time-locked to freezing that were also observed during natural fear memory recall, suggesting that engram activation alters activity across different cell types within hippocampal circuits during the behavioral expression of fear. Together, our data reveals cell-type specific hippocampal dynamics during freezing behavior and points to neuronal-astrocytic coupling as a shared mechanism enabling the natural and artificial recall of a memory.

Choice anticipation as gated accumulation of sensory expectations
Expectations are combined with sensory information when making choices. Some models of the choice process have conceptualized expectations as trial by trial updates to baseline evidence in an accumulator framework. These models have been successful in explaining the influence of choice history across trials on reaction times and choice probabilities, however they do not account for variability in the delay interval within trials. Here, we derive a gated accumulator that models the onset of evidence accumulation as a combination of delayed sensory information and an expectation of sensory timing that changes within trials. To test how the delay intervals interact with trial by trial expectations, we designed a free choice saccade task where participants directed eye movements, as fast as possible, to either of two targets that appeared with variable delays and asynchronies. Despite being instructed to not to anticipate, participants responded prior to target onset on a subset of trials. We reasoned that anticipatory responses may reflect a trade-off between gating the onset of evidence accumulation prior to target onset and releasing this gate as target appearance became more likely. Using a choice history analysis, we found that anticipatory responses were more likely following repeated choices, despite task randomization, suggesting that the balance between anticipatory and sensory responses was driven by an expectation of sensory timing. By fitting the gated accumulator model to reaction time data, we determined that within-trial fluctuations in baseline evidence occurring prior to the onset of the target could explain the joint increase of anticipatory responses and faster sensory-guided responses as the delay period increased. Thus, we conclude that an expectation of sensory timing can shape how the accumulation of choice evidence is gated, allowing us to balance the costs of anticipation with lowering the amount of evidence required to trigger movement.

NMDAR Phosphoproteome Controls Synaptic Growth and Learning
In the mammalian brain, NMDA receptors (NMDARs) activation triggers a calcium-dependent signal transduction cascade resulting in postsynaptic remodeling and behavioral learning. However, the phosphoprotein signal flow through this transduction network is poorly understood. Here, we show that NMDAR-dependent phosphorylation drives the assembly of protein signaling complexes that regulate synaptic morphology and behavior. We performed large-scale phosphoproteomic analyses of protein kinase target proteins in successive layers of the signaling network in mouse striatal/accumbal slices. NMDARs activation resulted in the phosphorylation of 194 proteins, including Rho GTPase regulators. CaMKII-mediated phosphorylation of ARHGEF2 increased its RhoGEF activity, thereby activating the RhoA-Rho-kinase pathway. Subsequent phosphoproteomics of Rho-kinase revealed 221 protein targets, including SHANK3. Experimental validation revealed a pathway from NMDAR-dependent calcium influx through CaMKII, ARHGEF2, Rho-kinase, and SHANK3 to coordinate assembly of an actin-tethered postsynaptic complex of SHANK3/NMDAR/PSD95/DLGAP3 for spine growth and aversive learning. These findings show that NMDARs initiate metabolic phosphorylation for learning.

Dissociable effects of urgency and evidence accumulation during reaching revealed by dynamic multisensory integration
When making perceptual decisions, humans combine information across sensory modalities dependent on their respective uncertainties. However, it remains unknown how the brain handles multisensory integration during movement, and which factors besides sensory uncertainty might influence the contribution of different modalities. We performed two reaching experiments on healthy adults to investigate whether movement corrections to combined visual and mechanical perturbations scale with visual uncertainty. To describe the dynamics of multimodal feedback responses, we further varied movement speed and duration of visual feedback during the movement. The results of our first experiment (N=16, 11 females) show that the contribution of visual feedback decreased with uncertainty. Interestingly, we observed a transient phase during which visual feedback responses were stronger during faster movements. In a follow-up experiment (N=16, 10 females), we found that the contribution of vision increased more quickly during slow movements when we presented the visual feedback for a longer time. Using an optimal feedback control model, we show that the increased response to visual feedback during fast movements can be explained by an urgency-dependent increase in control gains. Further, the fact that viewing duration increased the visual contributions suggests that the brain indeed performs a continuous state-estimation as expected in the optimal control model featuring a Kalman filter. Hence, both uncertainty and urgency determine how the sensorimotor system responds to multimodal perturbation during reaching control. We highlight similarities between reaching control and decision-making, both of which appear to be influenced by the accumulation of sensory evidence as well as response urgency.

Metabolic sensing in AgRP regulates sucrose preference and dopamine release in the nucleus accumbens
Hunger increases the motivation for calorie consumption, often at the expense of low taste appeal. However, the neural mechanisms integrating calorie-sensing with increased motivation for calorie consumption remain unknown. Agouti-related peptide neurons in the arcuate nucleus of the hypothalamus sense hunger, and the ingestion of caloric solutions promote dopamine release in the absence of sweet taste perception. Therefore, we hypothesized that metabolic-sensing of hunger by AgRP neurons would be essential to promote dopamine release in the nucleus accumbens in response to caloric, but not non-caloric solutions. Moreover, we examined whether metabolic sensing in AgRP neurons affected taste preference to bitter solutions under conditions of energy need. Here we show that impaired metabolic sensing in AgRP neurons attenuated nucleus accumbens dopamine release in response to sucrose, but not saccharin, consumption. Further, metabolic sensing in AgRP neurons was essential to distinguish nucleus accumbens dopamine response to sucrose consumption when compared with saccharin. Under conditions of hunger, metabolic sensing in AgRP neurons increased the preference of sucrose solutions laced with the bitter tastant, quinine, to ensure calorie consumption whereas mice with impaired metabolic sensing in AgRP neurons maintained a strong aversion to sucrose/quinine solutions despite ongoing hunger. In conclusion, we demonstrate normal metabolic sensing in AgRP neurons drives the preference for calorie consumption, primarily when needed, by engaging dopamine release in nucleus accumbens.

Individual variations in McGurk illusion susceptibility reflect different integration-segregation strategies of audiovisual speech perception
The McGurk illusion is a widely used indicator of audiovisual speech integration. In this illusion, incongruent visual articulations bias the auditory speech percepts, resulting in illusory percepts that differ from auditory and visual inputs. Despite its widespread use, its validity for measuring audiovisual integration has been questioned due to substantial individual variations. Classical forced fusion theories propose that variations in McGurk illusion susceptibility reflect differences in multisensory integration ability, whereas Bayesian causal inference (BCI) theory proposes that these variations reflect different audiovisual integration-segregation strategies used according to unisensory accuracy and inferred causal structures of the auditory and visual signals. To test these two proposals, we investigated the relationships between variations in McGurk illusion susceptibility and unisensory accuracy across testing time (Experiment 1, N =161), task types (Experiment 2, N = 88), and stimuli (Experiment 3, N = 37). We found stable negative correlations between McGurk illusion susceptibility and unisensory accuracy both at the group level and single participant level. Participants with weak illusion susceptibility had higher unisensory accuracy than participants with strong illusion susceptibility. Conversely, participants with strong illusion susceptibility were more likely to incorrectly categorize the auditory and visual stimuli as illusory percepts. Moreover, participants with similar unisensory accuracy showed similar McGurk illusion susceptibility. Consistent with the BCI theory, our results indicate that individual variations in McGurk illusion susceptibility reflect different integration-segregation strategies used by participants due to variations in unisensory accuracy. Failing to perceive an illusion does not indicate a failure to integrate audiovisual signals; instead, it indicates a successful segregation of incongruent signals. These findings suggest that caution is necessary when generalizing variations in McGurk illusion susceptibility to differences in audiovisual integration ability.

What pupil size can and cannot tell about math anxiety
Math Anxiety (MA) consists of excessive fear and worry about math-related situations. It represents a major barrier to numerical competence and the pursuit of STEM careers. Yet it is still poorly evaluated, mostly through self-reports. Here we sought to probe Pupil Size (PS) as a viable biomarker of MA by administering arithmetic problems to young adults (N= 70) with various levels of MA. We found that arithmetic competence and performance are indeed negatively associated with MA, and this is accurately tracked by PS. When performance is accounted for, MA does not further modulate PS (before, during, or after calculation). However, the latency of PS peak dilation could add a significant contribution to predicting MA scores, indicating that high MA may be accompanied by more prolonged cognitive effort. Results show that MA and mathematical competence may be too crystalized in young adults to be discernible, calling for early educational interventions.

Task dependent coarticulation of movement sequences
Classical theories propose that multiple goals of a sequence are specified simultaneously in the motor system, enabling each element's execution to be influenced by others in the sequence, a process known as coarticulation. However, recent neural recordings suggest independent execution within circuits generating motor commands, with each individual goal separately specified to the controller. Consequently, the manner in which movement sequences are produced remains unclear. In a two-reaches sequence task, we tested the idea that both coarticulation and separation are related to flexible feedback control. More precisely, simulations revealed that an optimal feedback controller planning for multiple reaches at once can produce both separated and coarticulated sequences based on constraints imposed at intermediate goals. Human experiments confirmed that the coarticulation of sequential reaches was flexibly modulated by task instructions, matching simulation results closely. Moreover, the location of the second goal impacted long-latency muscle stretch responses to mechanical perturbations applied prior to the first goal, indicating that sequence combination was expressed in the sensorimotor network mediating fast feedback control. Overall, our results uncover the role of long-latency feedback pathway as a flexible control policy, supporting efficient sequence production without the need to individually specify each target one-by-one to the controller.

The use of threshold methods to detect beta bursts may inadequately characterize the beta band
In neurophysiological research, the traditional view of beta band activity as sustained oscillations is being reinterpreted as transient bursts. Bursts are characterized by a distinct wavelet shape, high amplitude, and, most importantly, brief temporal occurrence. The primary method for their detection relies on a threshold-based analysis of spectral power, and this presents two fundamental issues. First, the threshold selection is effectively arbitrary, being influenced by both local and global factors in the signal. Second, the method necessarily detects temporal events, as such it is susceptible to misidentifying sustained signals as transient bursts. To address these issues, this study systematically explores burst detection through simulations, shedding light on the method's robustness across various scenarios. Although the method is effective in detecting transients in numerous cases, it can be overly sensitive, leading to spurious detections. Moreover, when applied to simulations featuring exclusively sustained events, the method frequently yields events exhibiting characteristics consistent with a transient burst profile. This capacity to produce misleading outcomes challenges the reinterpretation of sustained beta oscillations as transient bursts and prompts a critical reassessment of the existing literature.

AAV-mediated interneuron-specific gene replacement for Dravet syndrome
Dravet syndrome (DS) is a devastating developmental epileptic encephalopathy marked by treatment-resistant seizures, developmental delay, intellectual disability, motor deficits, and a 10-20% rate of premature death. Most DS patients harbor loss-of-function mutations in one copy of SCN1A, which has been associated with inhibitory neuron dysfunction. Here we developed an interneuron-targeting AAV human SCN1A gene replacement therapy using cell class-specific enhancers. We generated a split-intein fusion form of SCN1A to circumvent AAV packaging limitations and deliver SCN1A via a dual vector approach using cell class-specific enhancers. These constructs produced full-length NaV1.1 protein and functional sodium channels in HEK293 cells and in brain cells in vivo. After packaging these vectors into enhancer-AAVs and administering to mice, immunohistochemical analyses showed telencephalic GABAergic interneuron-specific and dose-dependent transgene biodistribution. These vectors conferred strong dose-dependent protection against postnatal mortality and seizures in two DS mouse models carrying independent loss-of-function alleles of Scn1a, at two independent research sites, supporting the robustness of this approach. No mortality or toxicity was observed in wild-type mice injected with single vectors expressing either the N-terminal or C-terminal halves of SCN1A, or the dual vector system targeting interneurons. In contrast, nonselective neuronal targeting of SCN1A conferred less rescue against mortality and presented substantial preweaning lethality. These findings demonstrate proof-of-concept that interneuron-specific AAV-mediated SCN1A gene replacement is sufficient for significant rescue in DS mouse models and suggest it could be an effective therapeutic approach for patients with DS.

A CAG repeat threshold for therapeutics targeting somatic instability in Huntington's disease
The Huntington's disease mutation is a CAG repeat expansion in the huntingtin gene that results in an expanded polyglutamine tract in the huntingtin protein. The CAG repeat is unstable, and expansions of hundreds of CAGs have been detected in Huntington's disease post-mortem brains. The age of disease onset can be predicted partially from the length of the CAG repeat as measured in blood. Onset age is also determined by genetic modifiers, which in six cases involve variation in DNA mismatch repair pathways genes. Knocking-out specific mismatch repair genes in mouse models of Huntington's disease prevents somatic CAG repeat expansion. Taken together, these results have led to the hypothesis that somatic CAG repeat expansion in Huntington's disease brains is required for pathogenesis. Therefore, the pathogenic repeat threshold in brain is longer than (CAG)40, as measured in blood, and is currently unknown. The mismatch repair gene MSH3 has become a major focus for therapeutic development, as unlike other mismatch repair genes, nullizygosity for MSH3 does not cause malignancies associated with mismatch repair deficiency. Potential treatments targeting MSH3 currently under development include gene therapy, biologics and small molecules, which will be assessed for efficacy in mouse models of Huntington's disease. The zQ175 knock-in model carries a mutation of approximately (CAG)185 and develops early molecular and pathological phenotypes that have been extensively characterised. Therefore, we crossed the mutant huntingtin allele onto heterozygous and homozygous Msh3 knock-out backgrounds to determine the maximum benefit of targeting Msh3 in this model. Ablation of Msh3 prevented somatic expansion throughout the brain and periphery, and reduction of Msh3 by 50% decreased the rate of expansion. This had no effect on the deposition of huntingtin aggregation in the nuclei of striatal neurons, nor on the dysregulated striatal transcriptional profile. This contrasts with ablating Msh3 in knock-in models with shorter CAG repeat expansions. Therefore, further expansion of a (CAG)185 repeat in striatal neurons does not accelerate the onset of molecular and neuropathological phenotypes. It is striking that highly expanded CAG repeats of a similar size in humans cause disease onset before 2 years of age, indicating that somatic CAG repeat expansion in the brain is not required for pathogenesis. Given that the trajectory for somatic CAG expansion in the brains of Huntington's disease mutation carriers is unknown, our study underlines the importance of administering treatments targeting somatic instability as early as possible.

Development And Characterization Of A Non-Human Primate Model Of Disseminated Synucleinopathy
The presence of a widespread cortical synucleinopathy is the main neuropathological hallmark underlying clinical entities such as Parkinson disease with dementia (PDD) and dementia with Lewy bodies (DLB). There currently is a pressing need for the development of non-human primate (NHPs) models of PDD and DLB to further overcome existing limitations in drug discovery. Here we took advantage of a retrogradely-spreading adeno-associated viral vector serotype 9 coding for the alpha-synuclein A53T mutated gene to induce a widespread synucleinopathy of cortical and subcortical territories innervating the putamen. Four weeks post-AAV deliveries animals were sacrificed and a comprehensive biodistribution study was conducted, comprising the quantification of neurons expressing alpha-synuclein, rostrocaudal distribution and their specific location. In brief, cortical afferent systems were found to be the main contributors to putaminal afferents (superior frontal and precentral gyrus in particular), together with neurons located in the caudal intralaminar nuclei and in the substantia nigra pars compacta (leading to thalamostriatal and nigrostriatal projections, respectively). Obtained data extends current models of synucleinopathies in NHPs, providing a reproducible platform enabling the adequate implementation of end-stage preclinical screening of new drugs targeting alpha-synuclein.

Changes in intra- and interlimb reflexes from hindlimb cutaneous afferents after staggered thoracic lateral hemisections during locomotion in cats
When the foot dorsum contacts an obstacle during locomotion, cutaneous afferents signal central circuits to coordinate muscle activity in the four limbs. Spinal cord injury disrupts these interactions, impairing balance and interlimb coordination. We evoked cutaneous reflexes by electrically stimulating left and right superficial peroneal nerves before and after two thoracic lateral hemisections placed on opposite sides of the cord at 9-13 weeks interval in seven adult cats (4 males and 3 females). We recorded reflex responses in ten hindlimb and five forelimb muscles bilaterally. After the first (right T5-T6) and second (left T10-T11) hemisections, coordination of the fore- and hindlimbs was altered and/or became less consistent. After the second hemisection, cats required balance assistance to perform quadrupedal locomotion. Short-latency reflex responses in homonymous and crossed hindlimb muscles largely remained unaffected after staggered hemisections. However, mid- and long-latency homonymous and crossed responses in both hindlimbs occurred less frequently after staggered hemisections. In forelimb muscles, homolateral and diagonal mid- and long-latency response occurrence significantly decreased after the first and second hemisections. In all four limbs, however, when present, short-, mid- and long-latency responses maintained their phase-dependent modulation. We also observed reduced durations of short-latency inhibitory homonymous responses in left hindlimb extensors early after the first hemisection and delayed short-latency responses in the right ipsilesional hindlimb after the first hemisection. Therefore, changes in cutaneous reflex responses correlated with impaired balance/stability and interlimb coordination during locomotion after spinal cord injury. Restoring reflex transmission could be used as a biomarker to facilitate locomotor recovery.

Markov Spatial Flows in BOLD fMRI: a Novel Lens on the BOLD Signal Reveals Attracting Patterns of Signal Intensity
While the analysis of temporal signal fluctuations and co-fluctuations has long been a fixture of blood oxygenation-level dependent (BOLD) functional magnetic resonance imaging (fMRI) research, the role and implications of spatial propagation within the 4D neurovascular BOLD signal has been almost entirely neglected. As part of a larger research program aimed at capturing and analyzing spatially propagative dynamics in BOLD fMRI, we report here a method that exposes large-scale functional attractors of spatial flows formulated as Markov processes defined at the voxel level. The brainwide stationary distributions of these voxel-level Markov processes represent patterns of signal accumulation toward which we find evidence that the brain exerts a probabilistic propagative undertow. These probabilistic propagative attractors are spatially structured and organized interpretably over functional regions. They also differ significantly between schizophrenia patients and controls.