bioRxiv Subject Collection: Neuroscience's Journal

AKAP150-anchored PKA regulation of synaptic transmission and plasticity, neuronal excitability and CRF neuromodulation in the lateral habenula
Numerous studies of hippocampal synaptic function in learning and memory have established the functional significance of the scaffolding A-kinase anchoring protein 150 (AKAP150) in kinase and phosphatase regulation of synaptic receptor and ion channel trafficking/function and hence synaptic transmission/plasticity and neuronal excitability. Emerging evidence also suggests that AKAP150 signaling may play a critical role in brain's processing of rewarding/aversive experiences. Here we focused on an unexplored role of AKAP150 in the lateral habenula (LHb), a diencephalic brain region that integrates and relays negative reward signals from forebrain striatal and limbic structures to midbrain monoaminergic centers. LHb aberrant activity (specifically hyperactivity) is also linked to depression. Using whole cell patch clamp recordings in LHb of male wildtype (WT) and {Delta}PKA knockin mice (with deficiency in AKAP-anchoring of PKA), we found that the genetic disruption of PKA anchoring to AKAP150 significantly reduced AMPA receptor (AMPAR)-mediated glutamatergic transmission and prevented the induction of presynaptic endocannabinoid (eCB)-mediated long-term depression (LTD) in LHb neurons. Moreover, {Delta}PKA mutation potentiated GABAA receptor (GABAAR)-mediated inhibitory transmission postsynaptically while increasing LHb intrinsic neuronal excitability through suppression of medium afterhyperpolarizations (mAHPs). Given that LHb is a highly stress-responsive brain region, we further tested the effects of corticotropin releasing factor (CRF) stress neuromodulator on synaptic transmission and intrinsic excitability of LHb neurons in WT and {Delta}PKA mice. As in our earlier study in rat LHb, CRF significantly suppressed GABAergic transmission onto LHb neurons and increased intrinsic excitability by diminishing small-conductance potassium (SK) channel-mediated mAHPs. {Delta}PKA mutation-induced suppression of mAHPs also blunted the synaptic and neuroexcitatory actions of CRF in mouse LHb. Altogether, our data suggest that AKAP150 complex signaling plays a critical role in regulation of AMPAR and GABAAR synaptic strength, glutamatergic plasticity and CRF neuromodulation possibly through AMPAR and potassium channel trafficking and eCB signaling within the LHb.

Therapeutic validation of MMR-associated genetic modifiers in a human ex vivo model of Huntington's disease
The pathological huntingtin (HTT) trinucleotide repeat underlying Huntington's disease (HD) continues to expand throughout life. Repeat length correlates both with earlier age at onset (AaO) and faster progression, making slowing its expansion an attractive therapeutic approach. Genome-wide association studies have identified candidate variants associated with altered AaO and progression, with many found in DNA mismatch repair (MMR) associated genes. We examine whether lowering expression of these genes affects the rate of somatic expansion in human ex vivo models using HD iPSCs and HD iPSC-derived striatal neurons. We have generated a stable CRISPR interference HD iPSC line in which we can specifically and efficiently lower gene expression from a donor carrying over 125 CAG repeats. Lowering expression of each member of the MMR complexes MutS (MSH2, MSH3 & MSH6), MutL (MLH1, PMS1, PMS2 & MLH3) and LIG1 resulted in characteristic MMR deficiencies. Reduced MSH2, MSH3 and MLH1 slowed repeat expansion to the largest degree, while lowering either PMS1, PMS2 and MLH3 slowed it to a lesser degree. These effects were recapitulated in iPSC derived striatal cultures where MutL factor expression was lowered. Here, reducing the expression of MMR factors by CRISPRi to levels typically reached by current therapeutics effectively slows the pathogenic expansion of the HTT CAG repeat tract. We highlight members of the MutL family as potential therapeutic targets to slow repeat expansion with the aim to delay onset and progression of HD, and potentially other repeat expansion disorders exhibiting somatic instability.

Loss of PV interneurons in the BLA contributes to altered network and behavioral states in chronically epileptic mice
Psychiatric disorders, including anxiety and depression, are highly comorbid in people with epilepsy. However, the mechanisms mediating the shared pathophysiology are currently unknown. There is considerable evidence implicating the basolateral amygdala (BLA) in the network communication of anxiety and fear, a process demonstrated to involve parvalbumin-positive (PV) interneurons. The loss of PV interneurons has been well described in the hippocampus of chronically epileptic mice and in postmortem human tissue of patients with temporal lobe epilepsy (TLE). We hypothesize that a loss of PV interneurons in the BLA may contribute to comorbid mood disorders in epilepsy. To test this hypothesis, we employed a ventral intrahippocampal kainic acid (vIHKA) model of chronic epilepsy in mice, which exhibits profound behavioral deficits associated with chronic epilepsy. We demonstrate a loss of PV interneurons and dysfunction of remaining PV interneurons in the BLA of chronically epileptic mice. Further, we demonstrate altered principal neuron function and impaired coordination of BLA network and behavioral states in chronically epileptic mice. To determine whether these altered network and behavioral states were due to the loss of PV interneurons, we ablated a similar percentage of PV interneurons observed in chronically epileptic mice by stereotaxically injecting AAV-Flex-DTA into the BLA of PV-Cre mice. Loss of PV interneurons in the BLA is sufficient to alter behavioral states, inducing deficits in fear learning and recall of fear memories. These data suggest that compromised inhibition in the BLA in chronically epileptic mice contributes to behavioral deficits, suggesting a novel mechanism contributing to comorbid anxiety and epilepsy.

Cue-Invariant Geometric Structure of the Population Codes in Macaque V1 and V2
We investigated the cue-invariant representation of visual patterns associated with surface boundaries in V1 and V2. We found that individual neurons exhibited a modest degree of tuning invariance in their responses to these patterns. This tuning invariance is stronger in V1 than in V2. At a population activity level, we studied the performance of a decoder trained with one cue in decoding patterns defined in another cue. We found that cue-transfer decoding is greatly enhanced when a geometric transform is first performed to align the population activities across cues. With this geometric transform, transfer decoding can be successful even when the tuning invariance of the individual neurons are destroyed by shuffling and when the neurons are from distinct populations. These findings suggest that abstract representation of these boundary-related patterns in V1 and V2 might be primarily encoded in the geometric structures of the population codes, rather than the cue-invariant tuning properties of the individual neurons.

Aversive memories can be weakened during human sleepvia the reactivation of positive interfering memories
Recollecting painful or traumatic experiences can be deeply troubling. Sleep may offer an opportunity to reduce such suffering. Accordingly, we developed a procedure to weaken older aversive memories by reactivating newer positive memories during sleep, thereby producing interference. Participants viewed 48 nonsense words each paired a unique aversive image, followed by overnight sleep. The next day, participants learned additional associations between half of the words and positive images, creating interference. During non-rapid-eye-movement sleep that night, memory cues were unobstruisvely delivered. Upon waking, presenting cues associated with both aversive and positive images during sleep, as opposed to not presenting cues, weakened aversive memory recall while increasing positive memory intrusions. Substantiating these memory benefits, computational modeling revealed that cueing facilitated evidence accumulation toward positive affect judgments. Moreover, cue-elicited theta brain rhythms during sleep predominantly predicted recall of positive memories. A noninvasive sleep intervention can thus modify aversive recollection and affective responses.

Sex-specific role for the long noncoding RNA Pnky in mouse behavior
The human brain expresses thousands of different long noncoding RNAs (lncRNAs), and aberrant expression of specific lncRNAs has been associated with cognitive and psychiatric disorders. While a growing number of lncRNAs are now known to regulate neural cell development and function, relatively few have been shown to underlie animal behavior, particularly with genetic strategies that establish lncRNA function in trans. Pnky is an evolutionarily conserved, neural lncRNA that regulates brain development. Using mouse genetic strategies, we show that Pnky has sex-specific roles in mouse behavior and that this lncRNA underlies specific behavior by functioning in trans. Male Pnky-knockout (KO) mice have deficits in cued fear recall, a type of Pavlovian associative memory. In female Pnky-KO mice, the acoustic startle response (ASR) is increased and accompanied by a decrease in prepulse inhibition (PPI), both of which are behaviors altered in affective disorders. Remarkably, expression of Pnky from a bacterial artificial chromosome (BAC) transgene reverses the ASR phenotype of female Pnky-KO mice, demonstrating that Pnky underlies specific animal behavior by functioning in trans. More broadly, these data provide genetic evidence that a lncRNA gene and its function in trans can play a key role in the behavior of adult mammals, contributing fundamental knowledge to our growing understanding of the association between specific lncRNAs and disorders of cognition and mood.

The Effects of Physical and Mental Fatigue on Time Perception
The subjective perception of time holds a foundational significance within the realm of human psychology and our conceptualizations of reality and how we elucidate the chronological progression of events within our lives. While there have been some studies examining the effects of exercise on time perception during the exercise period, there are no studies investigating the effects of fatiguing exercise on time perception after the exercise intervention. This study aimed to investigate the effects of physical and mental fatigue on time estimates over 30-seconds (5-, 10-, 20-, and 30-seconds) immediately after exercise and 6-minutes after the post-test. Seventeen healthy and recreationally active volunteers (14 males, 3 females) were subjected to three conditions: physical fatigue, mental fatigue, and control. All participants completed a familiarization and three 30-minute experimental conditions (control, physical fatigue (cycling at 65% peak power output), and mental fatigue (Stroop task for 1100 trials) on separate days. Heart rate and body temperature were recorded at the pre-test, start, 5-, 10-, 20-, 30- seconds of the interventions, post-test, and 6-min follow-up. Rating of perceived exertion (RPE) was recorded four times during the intervention. Time perception was measured prospectively (at 5-, 10-, 20-, and 30-seconds) at the pre-test, post-test, and 6-minute follow-up. Physical fatigue significantly (p=0.001) underestimated time compared to mental fatigue and control conditions at the post-test and follow-up, with no significant differences between mental fatigue and control conditions. Heart rate, body temperature, and RPE were significantly (all p=0.001) higher with physical fatigue compared to mental fatigue and control conditions during the intervention and at the post-test. This study demonstrated that cycling-induced fatigue led to time underestimation compared to mental fatigue and control conditions. It is crucial to consider that physical fatigue has the potential to lengthen an individual's perception of time estimates in sports or work environments.

Neuronal connectivity, behavioral, and transcriptional alterations associated with the loss of MARK2
Neuronal connectivity is essential for adaptive brain responses and can be modulated by dendritic spine plasticity and the intrinsic excitability of individual neurons. Dysregulation of these processes can lead to aberrant neuronal activity, which has been associated with numerous neurological disorders including autism, epilepsy, and Alzheimer's disease. Nonetheless, the molecular mechanisms underlying aberrant neuronal connectivity remains unclear. We previously found that the serine/threonine kinase Microtubule Affinity Regulating Kinase 2 (MARK2), also known as Partitioning Defective 1b (Par1b), is important for the formation of dendritic spines in vitro. However, despite its genetic association with several neurological disorders, the in vivo impact of MARK2 on neuronal connectivity and cognitive functions remains unclear. Here, we demonstrate that loss of MARK2 in vivo results in changes to dendritic spine morphology, which in turn leads to a decrease in excitatory synaptic transmission. Additionally, loss of MARK2 produces substantial impairments in learning and memory, anxiety, and social behavior. Notably, MARK2 deficiency results in heightened seizure susceptibility. Consistent with this observation, RNAseq analysis reveals transcriptional changes in genes regulating synaptic transmission and ion homeostasis. These findings underscore the in vivo role of MARK2 in governing synaptic connectivity, cognitive functions, and seizure susceptibility.

Neural correlates of category learning in monkey inferior temporal cortex
We trained two monkeys implanted with multi-electrode arrays to categorize natural images of cats and dogs, in order to observe changes in neural activity related to category learning. We recorded neural activity from area TE, which is required for normal learning of visual categories based on perceptual similarity. Neural activity during a passive viewing task was compared pre- and post-training. After the category training, the accuracy of abstract category decoding improved. Specifically, the proportion of single units with category selectivity increased, and units sustained their category-specific responses for longer. Visual category learning thus appears to enhance category separability in area TE by driving changes in the stimulus selectivity of individual neurons and by recruiting more units to the active network.

Searchlight-based trial-wise fMRI decoding in the presence of trial-by-trial correlations
In multivariate pattern analysis (MVPA) for functional magnetic resonance imaging (fMRI) signals, trial-wise response amplitudes are sometimes estimated using a general linear model (GLM) with one onset regressor for each trial. When using rapid event-related designs with trials closely spaced in time, those estimates can be highly correlated due to the temporally smoothed shape of the hemodynamic response function. In previous work (Soch, J., Allefeld, C., & Haynes, J.-D. (2020). Inverse transformed encoding models - a solution to the problem of correlated trial-by-trial parameter estimates in fMRI decoding. NeuroImage, 209, 116449, 1-19. https://doi.org/10.1016/j.neuroimage.2019.116449), we have proposed inverse transformed encoding modelling (ITEM), a principled approach for trial-wise decoding from fMRI signals in the presence of trial-by-trial correlations. Here, we (i) perform simulation studies addressing its performance for multivariate signals and (ii) present searchlight-based ITEM analysis - which allows to predict a variable of interest from the vicinity of each voxel in the brain. We empirically validate the approach by confirming a priori plausible hypotheses about the well-understood visual system.

Sord deficient rats develop a motor-predominant peripheral neuropathy unveiling novel pathophysiological insights
Biallelic SORD mutations cause one of the most frequent forms of recessive hereditary neuropathy, estimated to affect approximately 10,000 patients in North America and Europe alone. Pathogenic SORD loss-of-function changes in the encoded enzyme sorbitol dehydrogenase result in abnormally high sorbitol levels in cells and serum. How sorbitol accumulation leads to peripheral neuropathy remains to be elucidated. A reproducible animal model for SORD neuropathy is essential to illuminate the pathogenesis of SORD deficiency and for preclinical studies of potential therapies. Therefore, we have generated a Sord knockout (KO), Sord-/-, Sprague Dawley rat, to model the human disease and to investigate the pathophysiology underlying SORD deficiency. We have characterized the phenotype in these rats with a battery of behavioral tests as well as biochemical, physiological, and comprehensive histological examinations. Sord-/- rats had remarkably increased levels of sorbitol in serum, cerebral spinal fluid (CSF), and peripheral nerve. Moreover, serum from Sord-/- rats contained significantly increased levels of neurofilament light chain, NfL, an established biomarker for axonal degeneration. Motor performance significantly declined in Sord-/- animals starting at ~7 months of age. Gait analysis evaluated with video motion tracking confirmed abnormal gait patterns in the hindlimbs. Motor nerve conduction velocities of the tibial nerves were slowed. Light and electron microscopy of the peripheral nervous system revealed degenerating myelinated axons, de- and remyelinated axons, and a likely pathognomonic finding, enlarged ballooned myelin sheaths. These findings mainly affected myelinated motor axons; myelinated sensory axons were largely spared. In summary, Sord-/- rats develop a motor-predominant neuropathy that closely resembles the human phenotype. Our studies revealed novel significant aspects of SORD deficiency, and this model will lead to an improved understanding of the pathophysiology and the therapeutic options for SORD neuropathy.

Multilayer meta-matching: translating phenotypic prediction models from multiple datasets to small data
Resting-state functional connectivity (RSFC) is widely used to predict phenotypic traits in individuals. Large sample sizes can significantly improve prediction accuracies. However, for studies of certain clinical populations or focused neuroscience inquiries, small-scale datasets often remain a necessity. We have previously proposed a "meta-matching" approach to translate prediction models from large datasets to predict new phenotypes in small datasets. We demonstrated large improvement of meta-matching over classical kernel ridge regression (KRR) when translating models from a single source dataset (UK Biobank) to the Human Connectome Project Young Adults (HCP-YA) dataset. In the current study, we propose two meta-matching variants ("meta-matching with dataset stacking" and "multilayer meta-matching") to translate models from multiple source datasets across disparate sample sizes to predict new phenotypes in small target datasets. We evaluate both approaches by translating models trained from five source datasets (with sample sizes ranging from 862 participants to 36,834 participants) to predict phenotypes in the HCP-YA and HCP-Aging datasets. We find that multilayer meta-matching modestly outperforms meta-matching with dataset stacking. Both meta-matching variants perform better than the original "meta-matching with stacking" approach trained only on the UK Biobank. All meta-matching variants outperform classical KRR and transfer learning by a large margin. In fact, KRR is better than classical transfer learning when less than 50 participants are available for finetuning, suggesting the difficulty of classical transfer learning in the very small sample regime. The multilayer meta-matching model is publicly available at GITHUB_LINK.

Statistical diversity distinguishes global states of consciousness
Application of complexity measures to neurophysiological time series has seen increased use in recent years to identify neural correlates of global states of consciousness. Lempel-Ziv complexity is currently the de-facto complexity measure used in these investigations. However, by simply counting the number of patterns, this measure theoretically takes its maximum value for data that are completely random. Recently, a measure of statistical complexity - which calculates the diversity of statistical interactions - has been devised which aims to account for and remove randomness seen in data. It was recently found that this measure decreases during anaesthesia in fruit flies. This paper investigates this statistical complexity measure on human neurophysiology data from different stages of sleep, and from individuals under the effects of three psychedelic substances: ketamine, lysergic acid diethylamide (LSD), and psilocybin. Results indicate that statistical complexity: (i) differentiates the different stages of sleep analogously to Lempel-Ziv complexity; (ii) increases relative to placebo for all three psychedelic substances. Thus, statistical complexity is a useful alternative measure for investigating the complexity of neural activity associated with different states of consciousness.

Aperiodic EEG predicts variability of visual temporal processing
The human brain exhibits both oscillatory and aperiodic, or 1/f, activity. Although a large body of research has focused on the relationship between brain rhythms and sensory processes, aperiodic activity has often been dismissed as noise and functionally irrelevant. Prompted by recent findings linking aperiodic activity to the balance between neural excitation and inhibition, we investigated its effects on the temporal resolution of perception. We recorded EEG from participants during resting state and a task in which they detected the presence of two flashes separated by variable inter-stimulus intervals. Two-flash discrimination accuracy typically follows a sigmoid function whose steepness reflects perceptual variability or inconsistent integration/segregation of the stimuli. We found that individual differences in the steepness of the psychometric function correlated with EEG aperiodic exponents over posterior scalp sites. In other words, participants with higher levels of aperiodic activity (i.e., neural excitation) exhibited increased sensory noise, resulting in shallower psychometric curves. Our finding suggest that aperiodic EEG is linked to sensory integration processes usually attributed to the rhythmic inhibition of neural oscillations. Overall, this correspondence between aperiodic neural excitation and behavioral measures of sensory noise provides a more comprehensive explanation of the relationship between brain activity and sensory integration and represents an important extension to theories of how the brain samples sensory input over time.

Inhibition of the melanocortin-3 receptor (MC3R) causes generalized sensitization to anorectic agents
The melanocortin-3 receptor (MC3R) acts presynaptically to regulate GABA release from agouti-related protein (AgRP) nerve terminals and thus may be a negative regulator of multiple circuits involved in feeding behavior and energy homeostasis. Here, we examined the role of MC3R in regulating the response to various anorexigenic agents. Our findings reveal that genetic deletion or pharmacological inhibition of MC3R improves the dose responsiveness to Glucagon-like peptide 1 (GLP1) agonists, as assayed by inhibition of food intake and weight loss. An enhanced anorectic response to other agents, including the acute satiety factors peptide YY (PYY3-36) and cholecystokinin (CCK) and the long-term adipostatic factor, leptin, demonstrated that increased sensitivity to anorectic agents is a generalized result of MC3R antagonism. Enhanced neuronal activation in multiple nuclei, including ARH, VMH, and DMH, was observed using Fos immunohistochemistry following low-dose liraglutide in MC3R knockout mice (Mc3r-/-), supporting the hypothesis that the MC3R is a negative regulator of circuits regulating multiple aspects of feeding behavior. The enhanced anorectic response in Mc3r-/- mice after administration of GLP1 analogs was also independent of the incretin effects and malaise induced by GLP1R analogs, suggesting that MC3R antagonists may have value in enhancing the dose-response range of obesity therapeutics.

Cognitive load dissonance and personality factors: an empirical analysis in organizational settings
Previous research that aimed at characterizing the importance of workers' personological traits in coping with stress in organizational settings is often biased by the potential inconsistency (i.e., dissonance) between subjective perception and objective experience of workload. This study explored the relationship between the subjective, self-reported and objective physiological measures of cognitive load, and the possible confounding role of personality traits on a representative population (call center operators) in ecological settings (daily working routines consisting in inbound and outbound calls). With this aim, the personality traits of 30 call center operators were preliminarily characterized using the Ten Item Personality Inventory. Then, objective heart rate variability and electrodermal activity were measured during their inbound and outbound calls. Finally, a subjective self-evaluation of the experienced cognitive load was acquired. No significant correlations were found between subjective and objective measures of cognitive load except when controlling for personality traits. In particular, a negative correlation was found between the subjective perception of cognitive load and the psychophysiological indices of the parasympathetic tone. Specifically, the personality factors of Openness to Experience, Agreeableness and Emotional Stability have a significant influence on the subjective perception of cognitive load, without predicting the objective psychophysiological expression. Our study emphasizes the importance of studying the dissonance between subjectively-perceived feelings and their objective physiological instantiation, along with the influence of personality factors, in organizational settings.

Adaptation with naturalistic textures in macaque V1 and V2
Adaptation affects neuronal responsivity and selectivity throughout the visual hierarchy. However, because most prior studies have tailored stimuli to a single brain area of interest, we have a poor understanding of how exposure to a particular image alters responsivity and tuning at different stages of visual processing. Here we assess how adaptation with naturalistic textures alters neuronal responsivity and selectivity in primary visual cortex (V1) and area V2 of macaque monkeys. Neurons in both areas respond to textures, but V2 neurons are sensitive to high-order statistics which do not strongly modulate V1 responsivity. We tested the specificity of adaptation in each area with textures and spectrally-matched 'noise' stimuli. Adaptation reduced responsivity in both V1 and V2, but only in V2 was the reduction dependent on the presence of higher-order texture statistics. Despite this specificity, the texture information provided by single neurons and populations was reduced after adaptation, in both V1 and V2. Our results suggest that adaptation effects for a given feature are induced at the stage of processing that tuning for those features first arises and that stimulus-specific adaptation effects need not result in improved sensory coding.

Axon guidance genes are regulated by TDP-43 and RGNEF through the rate of long-intron processing.
Rho guanine nucleotide exchange factor (RGNEF) is a guanine nucleotide exchange factor (GEF) mainly involved in regulating the activity of Rho-family GTPases. Previous work has shown that RGNEF inclusions in the spinal motor neurons of ALS patients co-localise with TDP-43, the major RNA binding protein aggregating in the brain and spinal cord of ALS patients. To further characterise their relationship, we have compared the transcriptomic profiles of neuronal-like cells depleted of TDP-43 and RGNEF and show that these two factors predominantly act in an antagonistic manner when regulating the expression of axon guidance genes. From a mechanistic point of view, our experiments show that the effect of these factors on the processivity of long introns can explain their mode of action. Our findings highlight that neurodegenerative processes at the RNA level can often represent the result of combinatorial interactions between different RNA binding factors, leading to a better understanding of pathogenic mechanisms occurring in patients where more than one specific protein may be aggregating in their neurons.

Make or break: The influence of expected challenges and rewards on the motivation and experience associated with cognitive effort exertion
Challenging goals can induce harder work but also greater stress, in turn potentially undermining goal achievement. We sought to examine how mental effort and subjective experiences thereof interact as a function of challenge level and the size of the incentives at stake. Participants performed a task that rewarded individual units of effort investment (correctly performed Stroop trials) but only if they met a threshold number of correct trials within a fixed time interval (challenge level). We varied this challenge level (Study 1, N = 40), and the rewards at stake (Study 2, N = 79), and measured variability in task performance and self-reported affect across task intervals. Greater challenge and higher rewards facilitated greater effort investment but also induced greater stress, while higher rewards (but less challenge) simultaneously induced greater positive affect. Current findings further our understanding of task demands and incentives on mental effort exertion and wellbeing.

Rapid TetOn-mediated gene expression in neuronsacross the lifespan using uTTOP
Conditional expression of genes of interest is essential for interrogation of cellular development and function. Although tools exist for conditional gene expression, techniques for rapid-onset, temporally precise expression are lacking. The doxycycline-inducible TetOn expression system allows for this in numerous organ systems, however, transgenic TetOn expression cassettes become silenced in the nervous system during postnatal development. Here, we circumvent this silencing with uTTOP: in utero electroporation of Transposable TetOn Plasmids. When electroporated as transposable elements that integrate into the genome, the TetOn system allowed for robust DOX-dependent induction of expression across the postnatal lifespan of the mouse. We demonstrated induction in neurons of sensorimotor and retrospleninal cortex, hippocampus and the olfactory bulb. Latency to peak induction was [≤]12 hours, a several fold increase in induction kinetics over existing methodology for in vivo conditional expression. To demonstrate the utility of uTTOP, we induced ectopic expression of Sonic hedgehog in adult mouse layer 2/3 cortical neurons, demonstrating that its expression can diversify expression of Kir4.1 in surrounding astrocytes. The rapid induction kinetics of uTTOP allowed us to show that Kir4.1 upregulation significantly lags onset of Shh expression by ~2 days, a difference in expression time course that is likely not resolvable with current methods. Together, these data demonstrate that uTTOP is a powerful and flexible system for conditional gene expression in multiple brain areas across the mouse lifespan.

Speech-induced suppression and vocal feedback sensitivity in human cortex
Across the animal kingdom, neural responses in the auditory cortex are suppressed during vocalization, and humans are no exception. A common hypothesis is that suppression increases sensitivity to auditory feedback, enabling the detection of vocalization errors. This hypothesis has been previously confirmed in non-human primates, however a direct link between auditory suppression and sensitivity in human speech monitoring remains elusive. To address this issue, we obtained intracranial electroencephalography (iEEG) recordings from 35 neurosurgical participants during speech production. We first characterized the detailed topography of auditory suppression, which varied across superior temporal gyrus (STG). Next, we performed a delayed auditory feedback (DAF) task to determine whether the suppressed sites were also sensitive to auditory feedback alterations. Indeed, overlapping sites showed enhanced responses to feedback, indicating sensitivity. Importantly, there was a strong correlation between the degree of auditory suppression and feedback sensitivity, suggesting suppression might be a key mechanism that underlies speech monitoring. Further, we found that when participants produced speech with simultaneous auditory feedback, posterior STG was selectively activated if participants were engaged in a DAF paradigm, suggesting that increased attentional load can modulate auditory feedback sensitivity.

CELSR3 deficiency leads to tic-related behaviors and dopaminergic alterations in the striatum
The gene CELSR3 (cadherin EGF LAG seven-pass-G-type receptor 3) has been recently recognized as a high-confidence risk factor for Tourette syndrome (TS) Here, we characterized the behavioral phenotypes of Celsr3 mutant mice to verify whether the deficiency of this gene is associated with TS predisposition. Celsr3 mutant mice displayed tic-like grooming stereotypies and jerks, as well as sensorimotor gating deficits, which were opposed by TS therapies. Single-nucleus transcriptomic analyses revealed that Celsr3 mutants featured a unique group of eccentric striatal projection neurons. Notably, the Drd3 gene, encoding the dopamine D3 receptor, was significantly upregulated in these cells as well as striosomal D1-positive neurons, while it was reduced in calretinin-positive GABAergic interneurons. Activating and blocking D3 receptors amplified or decreased tic-like jerks and stereotypies in Celsr3-deficient mice, respectively. These findings suggest that modifications of D3 receptor distribution across various striatal cell populations contribute to the tic-like responses associated with Celsr3 deficiency.

Segmentation of supragranular and infragranular layers in ultra-high resolution 7T ex vivo MRI of the human cerebral cortex
Accurate labeling of specific layers in the human cerebral cortex is crucial for advancing our understanding of neurodevelopmental and neurodegenerative disorders. Leveraging recent advancements in ultra-high resolution ex vivo MRI, we present a novel semi-supervised segmentation model capable of identifying supragranular and infragranular layers in ex vivo MRI with unprecedented precision. On a dataset consisting of 17 whole-hemisphere ex vivo scans at 120 microns, we propose a multi-resolution U-Nets framework (MUS) that integrates global and local structural information, achieving reliable segmentation maps of the entire hemisphere, with Dice scores over 0.8 for supra- and infragranular layers. This enables surface modeling, atlas construction, anomaly detection in disease states, and cross-modality validation, while also paving the way for finer layer segmentation. Our approach offers a powerful tool for comprehensive neuroanatomical investigations and holds promise for advancing our mechanistic understanding of progression of neurodegenerative diseases.

Endogenous opioid receptor system mediates costly altruism in the human brain
Functional neuroimaging studies suggest that a large-scale brain network transforms others' pain into its vicarious representation in the observer, potentially modulating helping behaviour. The neuromolecular basis of individual differences in vicarious pain and helping have however remain poorly understood. Here we investigated the role of the endogenous mu-opioid receptor (MOR) system - known for its role in analgesia and sociability - in altruistic costly helping. MOR density was measured with high-affinity agonist radioligand [11C]carfentanil. In a separate fMRI experiment, participants could choose to donate money to reduce the pain of the confederate who was subjected to electric shocks of varying intensity. We found people were in general willing to engage in costly helping, and haemodynamic activity in amydala, aIns, anterior cingulate cortex, striatum, primary motor cortex, primary somatosensory cortex, and thalamus increased when participants witnessed the pain of others. These haemodynamic responses were negatively associated with MORs availability in the striatolimbic and cortical emotion circuits. In turn, haemodynamic responses during helping were positively associated with MOR availability in the anterior cingulate cortex and hippocampus. Altogether these data suggest that endogenous MOR system modulates processing of altruistic behaviour and costly helping in the human brain.

The Role of Attention in Multi Attribute Decision Making
Real-life decisions typically involve multiple options, each with multiple attributes affecting value. In such complex cases, sequential shifts of attention to specific options and attributes are thought to guide the decision process. We designed a task that allowed us to monitor attention in monkeys engaged in such multi-attribute decisions. We recorded pre-supplementary motor area neurons encoding action value signals reflecting the decision process. Attention guides this process through two mechanisms. First, attention enhances the activity of neurons representing the currently sampled option, independent of the attended option value. Second, attention up-regulates the gain of information integration towards the evolving value estimate for the attended option. In contrast, we found no evidence for a third suggested mechanism, in which only the attended option is represented. Instead, attention influences the ongoing information accumulation and competition between the options by modulating the strength of the value information that drives this circuit.

Unravelling neuronal and glial differences in ceramide composition, synthesis, and sensitivity to toxicity
Ceramides are lipids that play vital roles in complex lipid synthesis, membrane function, and cell signaling. Disrupted ceramide homeostasis is implicated in cell-death and several neurologic diseases. Ceramides are often analyzed in tissue, but this approach fails to resolve cell-type differences in ceramide homeostasis that are likely essential to understanding cell and non-cell autonomous contributions to neurodegeneration. We show that human iPSC-derived neurons and glia differ in their rate of ceramide synthesis, ceramide isoform composition, and responses to altered ceramide levels. RNA-sequencing of cells treated to increase or decrease ceramides revealed connections to inflammation, ER stress, and apoptosis. Moreover, introducing labeled sphinganine showed that glia readily synthesize ceramide de novo and that neurons are relatively more sensitive to ceramide toxicity. Our findings provide a framework for understanding neurologic diseases with sphingolipid alternations and insights in to designing therapeutics that target ceramide for treating them.

A phosphorylation-dependent switch of the lysosomal V-ATPase assembly regulates α-synuclein clearance in glia
Glia serve as double-edged swords to modulate neuropathology in Parkinson's disease (PD), but how they react opposingly to be beneficial or detrimental under pathological conditions, like promote or eliminate alpha-synuclein (alpha-syn) inclusions, remains elusive. Here we present evidence that the PD risk factor Cyclin G-associated kinase (GAK)/dAuxilin (dAux) regulates the lysosomal degradation of alpha-syn in glia. In addition to a broad spectrum of parkinsonian symptoms in Drosophila and mice, lack of glial Gak/dAux causes abnormally higher alpha-syn levels in fly and mouse brains, and further enhances the alpha-syn preformed fibril levels in mouse brains. dAux phosphorylates at the serine 543 of Vha44, the V1C subunit of the vacuolar H+-ATPase (V-ATPase), regulates the V-ATPase assembly, and controls the lysosomal acidification and hydrolase activity to set an acidic milieu for alpha-syn degradation in glia. Importantly, blocking Vha44 serine 543 phosphorylation disrupts V-ATPase assembly, lysosome acidification, and hydrolase activity, leading to DA neurodegeneration and locomotor deficits in flies. Our findings identify a phosphorylation-dependent switch controlling the V-ATPase assembly for lysosomal alpha-syn degradation in glia. Targeting the clearance of glial alpha-syn inclusions via this lysosomal pathway could potentially be a therapeutical approach to ameliorate the disease progression in PD.

Low-dimensional neural geometry underlies distributed goal representations in working memory
Successful goal-directed behavior requires the maintenance and implementation of abstract task goals on concrete stimulus information in working memory. Previous working memory research has revealed distributed neural representations of task information across cortex. However, how the distributed task representations emerge and communicate with stimulus-specific information to implement flexible goal-directed computations is still unclear. Here, leveraging EEG and fMRI along with state space analyses, we provided converging evidence in support of a low-dimensional neural geometry of goal information consistent with a designed task space, which first emerged in frontal cortex during goal maintenance and then transferred to posterior sensory cortex through frontomedial-to-posterior theta coherence for implementation on stimulus-specific representations. Importantly, the fidelity of the goal geometry predicted memory performance. Collectively, our findings suggest that abstract goals in working memory are represented in an organized, low-dimensional neural geometry for communications from frontal to posterior cortex to enable computations necessary for goal-directed behaviors.

Stimulus selection influences prediction of individual phenotypes in naturalistic conditions
Understanding individual differences and brain-behaviour relationships is an essential goal of human neuroscience. Recent studies have shown the great potential of naturalistic stimuli, e.g., movie clips, in advancing this pursuit. While the use of naturalistic stimuli attracts increasing interest, the influence of stimulus selection remains largely unclear. In this study, we show that brain activity is generally sensitive to the choice of movie stimuli at both group and individual subject levels. Using sex classification as an example, we demonstrate that brain activity elicited by different stimuli can lead to distinct prediction performance and unique predictive features. The stimuli that yield better classification performance often elicit stronger synchrony of brain activity across all subjects and are mostly derived from Hollywood films with rich social content and cohesive narratives. Our results highlight the importance of stimulus selection and provide practical guidance for choosing appropriate stimuli, opening up new avenues for future studies on individual differences and brain-behaviour relationships.

Modeling an ultra-rare epilepsy variant in wildtype mice with in utero prime editing
Generating animal models for individual patients within clinically relevant time frames holds the potential to revolutionize personalized medicine for rare genetic epilepsies. By incorporating patient-specific genomic variants into model animals, capable of replicating elements of the patient's clinical manifestations, a range of applications would be enabled: from preclinical platforms for rare disease drug screening, to bedside surrogates for tailoring pharmacotherapy without subjecting the patient to excessive trial medications. Here, we present the conceptual framework and proof-of-principle modeling of an individual epilepsy patient with an ultra-rare variant of the NMDA receptor subunit GRIN2A. Using in utero prime editing in the embryonic brain of wildtype mice, our approach demonstrated high editing precision and induced frequent, spontaneous seizures in prime editor-treated mice, reflecting key features of the patient's clinical presentation. Leveraging the speed and versatility of this approach, we introduce PegAssist, a generalizable 7-week workflow for bedside-to-bench modeling of patients using in utero prime editing. These individualized animal models can allow for widely-accessible personalized medicine for rare neurological conditions, as well as accelerate the drug development pipeline by providing an efficient and versatile preclinical platform for screening compounds against ultra-rare genetic diseases.

Brain-wide topographic coordination of traveling spiral waves
Traveling waves of activity are a prevalent phenomenon within neural networks of diverse brain regions and species, and have been implicated in myriad brain functions including sensory perception, memory, spatial navigation and motor control. However, the anatomical basis for these waves, and whether they are locally confined versus distributed across the brain, remains unclear. Here we use cortex-wide imaging and large-scale electrophysiology in awake mice to reveal the organization of traveling waves across spatial scales. Traveling waves formed spiral patterns predominantly centered on somatosensory cortex. Strikingly, the local axonal architecture of neurons in sensory cortex exhibited a matching circular arrangement. At the cortex-wide scale, these spiral waves were mirrored between hemispheres and between sensory and motor cortex, reflecting topographic long-range axons. Finally, at the brain-wide scale, cortical spiral waves were coordinated with subcortical spiking patterns in the thalamus, striatum and midbrain. These results establish that traveling waves are structured by axonal pathways and globally impact neural activity across diverse brain systems.

Metabolomics of Mouse Embryonic CSF Following Maternal Immune Activation
The cerebrospinal fluid (CSF) provides a protective fluid cushion for the brain (Fame & Lehtinen, 2020; Lacey et al, 2023) and delivers neuroactive proteins, peptides, and small molecules that are critical for normal brain development (Saunders et al, 2023; Gelb & Lehtinen, 2023). In the case of the developing cerebral cortex, whose neural progenitor cells line the brain's ventricles and extend primary cilia into the CSF, age-appropriate cocktails of CSF growth factors help ensure that progenitors remain healthy, acquire the correct identity, and proliferate in a developmentally appropriate manner (Chau et al, 2018; Lehtinen et al, 2011; Kim et al, 2010). Conversely, the CSF can be used as a readout of different stages of brain development including metabolic state (Chau et al, 2018; Fame et al, 2019). Indeed, CSF is commonly sampled for biomarkers of infections and neurologic diseases.

Knockdown of NeuroD2 leads to seizure-like behaviour, brain neuronal hyperactivity and a leaky blood-brain barrier in a Xenopus laevis tadpole model of DEE75
Developmental and Epileptic Encephalopathies (DEE) are a genetically diverse group of severe, early onset seizure disorders. DEE are normally identified clinically in the first six months of life by the presence of frequent, difficult to control seizures and accompanying stalling or regression of development. DEE75 results from de novo mutations of the NEUROD2 gene that result in loss of activity of the encoded transcription factor, and the seizure phenotype was shown to be recapitulated in Xenopus tropicalis tadpoles. We used CRISPR/Cas9 to make a DEE75 model in Xenopus laevis, to further investigate the developmental aetiology. NeuroD2.S CRISPR/Cas9 edited tadpoles were more active, swam faster on average, and had more unprovoked escape responses (C-starts) than their sibling controls. Live imaging of Ca2+ signalling revealed prolongued, strong signals sweeping through the brain, indicative of neuronal hyperactivity. While the resulting tadpole brain appeared grossly normal, the blood-brain barrier was found to be leakier than that of controls. Additionally, the TGF antagonist Losartan was shown to have a short-term protective effect, reducing neuronal hyperactivity and reducing permeability of the blood-brain barrier. Severity of the behavioral phenotype correlated with increased with editing efficiency. Our results support a haploinsufficiency model of DEE75 resulting from reduced NeuroD2 activity during vertebrate brain development, and indicate that a leaky blood-brain barrier contributes to epileptogenesis.

Multi-omics analyses reveal novel effects of PLCγ2 deficiency in the mouse brain
Phospholipase C gamma-2 (PLCg2) catalyzes the hydrolysis of the membrane phosphatidylinositol-4,5-bisphosphate (PIP2) to form diacylglycerol (DAG) and inositol trisphosphate (IP3), which subsequently feed into numerous downstream signaling pathways. PLCG2 polymorphisms are associated with both reduced and increased risk of Alzheimer's disease (AD) and with longevity. In the brain, PLCG2 is highly expressed in microglia, where it is proposed to regulate phagocytosis, secretion of cytokines/chemokines, cell survival and proliferation. We analyzed the brains of three-month-old PLCg2 knockout (KO), heterozygous (HET), and wild-type (WT) mice using multiomics approaches, including shotgun lipidomics, proteomics, and gene expression profiling, and immunofluorescence. Lipidomic analyses revealed sex-specific losses of total cerebrum PIP2 and decreasing trends of DAG content in KOs. In addition, PLCg2 depletion led to significant losses of myelin-specific lipids and decreasing trends of myelin-enriched lipids. Consistent with our lipidomics results, RNA profiling revealed sex-specific changes in the expression levels of several myelin-related genes. Further, consistent with the available literature, gene expression profiling revealed subtle changes on microglia phenotype in mature adult KOs under baseline conditions, suggestive of reduced microglia reactivity. Immunohistochemistry confirmed subtle differences in density of microglia and oligodendrocytes in KOs. Exploratory proteomic pathway analyses revealed changes in KO and HET females compared to WTs, with over-abundant proteins pointing to mTOR signaling, and under-abundant proteins to oligodendrocytes. Overall, our data indicate that loss of PLCg2 has subtle effects on brain homeostasis that may underlie enhanced vulnerability to AD pathology and aging via novel mechanisms in addition to regulation of microglia function.

(2R,6R)-hydroxynorketamine facilitates extinction and prevents emotional impairment and stress-induced reinstatement in morphine abstinent mice
Opioid addiction is a pressing public health concern marked by frequent relapse during periods of abstinence, perpetuated by negative affective states and anhedonia-driven behaviors. In addition to the current epidemic that was declared in the U.S.A., opioid-related deaths are increasing in other countries around the world. Classical antidepressants, or the currently prescribed opioid substitution pharmacotherapies have limited efficacy to reverse maladaptive behavioral responses, negative affect or prevent relapse in opioid abstinent individuals. Here, by establishing and using novel mouse models for the study of opioid addiction, we demonstrate, for the first time, the therapeutic potential of ketamine's metabolite, (2R,6R)-hydroxynorketamine (HNK). In particular, our studies showcase (2R,6R)-HNK's ability to reverse conditioning to sub-effective doses of morphine in stress-susceptible mice, prevent conditioned-place aversion and mitigate acute somatic withdrawal symptoms in opioid-dependent animals. In addition, we show that this metabolite reverses anhedonia, anxiety-like behaviors, cognitive impairment, and general stress susceptibility associated with protracted opioid withdrawal, thereby presenting a promising therapeutic avenue for opioid relapse prevention. Our results strongly suggest that (2R,6R)-HNK, potentially by augmenting downstream brain-derived neurotrophic factor (BDNF) and GluN2A N-methyl-D-aspartate receptor signaling, effectively reverses maladaptive behavioral responses typical of protracted opioid abstinence. Furthermore, it facilitates the extinction of opioid conditioning and prevents stress-induced reinstatement of opioid-seeking behaviors. Our findings highlight how (2R,6R)-HNK, through an enhancement of synaptic plasticity in mood-regulating brain areas, has the potential to be an effective, next-generation pharmacotherapy for opioid use disorders by addressing emotional disturbances associated with protracted abstinence.

Regional differences in synaptic degeneration are linked to alpha-synuclein burden and axonal damage in Parkinson's disease and Dementia with Lewy bodies
Regional differences in synaptic degeneration may underlie differences in clinical presentation and neuropathological disease progression in Parkinson's Disease (PD) and Dementia with Lewy bodies (DLB). Here, we mapped and quantified synaptic degeneration in cortical brain regions in PD, PD with dementia (PDD) and DLB, and assessed whether regional differences in synaptic loss are linked to axonal degeneration and neuropathological burden. We included a total of 47 brain donors, 9 PD, 12 PDD, 6 DLB and 20 non-neurological controls. Synaptophysin+ and SV2A+ puncta were quantified in eight cortical regions using a high throughput microscopy approach. Neurofilament light chain (NfL) immunoreactivity, Lewy body (LB) density, phosphorylated-tau and amyloid-beta load were also quantified. Group differences in synaptic density, and associations with neuropathological markers and Clinical Dementia Rating (CDR) scores, were investigated using linear mixed models. We found significantly decreased synaptophysin and SV2A densities in the cortex of PD, PDD and DLB cases compared to controls. Specifically, synaptic density was decreased in cortical regions affected at Braak alpha-synuclein stage 5 in PD (middle temporal gyrus, anterior cingulate and insula), and was additionally decreased in cortical regions affected at Braak alpha-synuclein stage 4 in PDD and DLB compared to controls (entorhinal cortex, parahippocampal gyrus and fusiform gyrus). Synaptic loss associated with higher NfL immunoreactivity and LB density. Global synaptophysin loss associated with longer disease duration and higher CDR scores. Synaptic neurodegeneration occurred in temporal, cingulate and insular cortices in PD, as well as in parahippocampal regions in PDD and DLB. In addition, synaptic loss was linked to axonal damage and severe alpha-synuclein burden. These results, together with the association between synaptic loss and disease progression and cognitive impairment, indicate that regional synaptic loss may underlie clinical differences between PD and PDD/DLB. Our results might provide useful information for the interpretation of synaptic biomarkers in vivo.

Glial swip-10 expression controls systemic mitochondrial function, oxidative stress, and neuronal viability via copper ion homeostasis
Cuprous copper (Cu(I)) is an essential cofactor for enzymes supporting many cellular functions including mitochondrial respiration and suppression of oxidative stress. Neurons are particularly dependent on these pathways, with multiple neurodegenerative diseases, including Alzheimers disease (AD), Parkinsons disease, associated with their dysfunction. Key features of Cu(I) contributions to neuronal health in vivo remain to be defined, owing largely to the complex processes involved in Cu(I) production, intracellular transport, and systemic redistribution. Here, we provide genetic and pharmacological evidence that swip-10 is a critical determinant of systemic Cu(I) levels in C. elegans, with deletion leading to systemic deficits in mitochondrial respiration, production of oxidative stress, and neurodegeneration. These phenotypes can be reproduced in wild-type worms by Cu(I)-specific chelation and offset in swip-10 mutants by growth on the Cu(I) enhancing molecule elesclomol, as well as by glial expression of wildtype swip-10. MBLAC1, the most closely related mammalian ortholog to swip-10, encodes for a pre-mRNA processing enzyme for H3 histone, a protein whose actions surprisingly include an enzymatic capacity to produce Cu(I) via the reduction of Cu(II). Moreover, genome-wide association studies and post-mortem molecular studies implicate reductions of MBLAC1 expression in risk for AD with cardiovascular disease comorbidity. Consistent with these studies, we demonstrate that the deposition of b-amyloid plaques, an AD pathological hallmark, in worms engineered to express human Ab1-42, is greatly exaggerated by mutation of swip- 10. Together, these studies identify a novel glial-expressed, and pathway for Cu(I) production that may be targeted for the treatment of AD and other neurodegenerative diseases.