bioRxiv Subject Collection: Neuroscience's Journal

Neocortical Layer-5 tLTD Relies on Non-Ionotropic Presynaptic NMDA Receptor Signaling
In the textbook view, NMDA receptors (NMDARs) act as coincidence detectors in Hebbian plasticity by fluxing Ca2+ when simultaneously depolarized and glutamate bound. Hebbian coincidence detection requires that NMDARs be located postsynaptically, but enigmatic presynaptic NMDARs (preNMDARs) also exist. It is known that preNMDARs regulate neurotransmitter release, but precisely how remains poorly understood. Emerging evidence suggest that NMDARs can also signal non-ionotropically, without the need for Ca2+ flux. At synapses between developing visual cortex layer-5 (L5) pyramidal cells (PCs), preNMDARs rely on Mg2+ and Rab3-interacting molecule 1{beta} (RIM1{beta}) to regulate evoked release during periods of high-frequency firing, but they signal non-ionotropically via c-Jun N-terminal kinase 2 (JNK2) to regulate spontaneous release regardless of frequency. At the same synapses, timing-dependent long-term depression (tLTD) depends on preNMDARs but not on frequency. We therefore tested if tLTD relies on non-ionotropic preNMDAR signaling. We found that tLTD at L5 PC[->]PC synapses was abolished by pre- but not postsynaptic NMDAR deletion, cementing the view that tLTD requires preNMDARs. In agreement with non-ionotropic NMDAR signaling, tLTD prevailed after channel pore blockade with MK-801, unlike tLTP. Homozygous RIM1{beta} deletion did not affect tLTD, but wash-in of the JNK2 blocker SP600125 abolished tLTD. Consistent with a presynaptic need for JNK2, a peptide blocking the interaction between JNK2 and Syntaxin-1a (STX1a) abolished tLTD if loaded pre- but not postsynaptically, regardless of frequency. Finally, low-frequency tLTD was not blocked by the channel pore blocker MK-801, nor by 7-CK, a non-competitive NMDAR antagonist at the co-agonist site. We conclude that neocortical L5 PC[->]PC tLTD relies on non-ionotropic preNMDAR signaling via JNK2/STX1a. Our study brings closure to long-standing controversy surrounding preNMDARs and highlights how the textbook view of NMDARs as ionotropic coincidence detectors in plasticity needs to be reassessed.

Stable Actin As Synaptic Tag
According to the tagging and capture hypothesis, long-lasting long-term potentiation (LTP) requires protein synthesis and a synaptic tag, which is a synapse specific memory of the synapse with a so far unclear molecular or biophysical identity. Here we use an interdisciplinary approach to explore the hypothesis that interaction between the dynamics of actin and the spine geometry can provide such a memory. Using a mathematical model, we demonstrate that this implementation of the tag requires an increase in the stable, cross-linked pool of actin filaments, and is not possible without this stable pool. Using FRAP experiments, we show that such an increase in stable actin can indeed be observed after cLTP in vitro. Thus, the interaction between actin dynamics and spine geometry could indeed serve as a synaptic tag for LTP.

Fixation shifts in a novel 'no-report' binocular rivalry paradigm induce saccade-related perceptual switches
No-report paradigms help to avoid report-related confounds in conscious perception studies. A novel no-report binocular rivalry paradigm by Hesse and Tsao (2020) tracks conscious content using eye position as subjects follow fixation points linked to the rivaling stimuli. However, it remains unclear whether perceptual changes arise spontaneously or are induced by external factors such as fixation shifts and saccades. We found an increased probability of perceptual switches time-locked to fixation point shifts, indicating that some switches are externally driven. To disentangle the effects of visual fixation shifts and saccades, we implemented a two-factorial design and found that saccades play a larger role in eliciting perceptual changes. We estimate that 14% of saccades trigger a switch, accounting for 24% of all perceptual transitions. Our findings provide an analysis framework and guidelines for excluding externally driven perceptual switches, enabling a clearer focus on internally generated perceptual dynamics.

Non-invasive peripheral delivery of CDNF fragment protects neurons in models of Parkinsons and ALS
Non-invasive delivery of brain therapeutics is a key challenge for treating neurodegenerative diseases. Here, we discovered a novel carboxy (C)-terminal fragment of cerebral dopamine neurotrophic factor (C-CDNF) that protects dopamine (DA) and motoneurons (MNs) in rodent models of Parkinsons disease (PD) and amyotrophic lateral sclerosis (ALS). C-CDNF retains the same structure as CDNF and similarly to CDNF regulates cell stress pathways but unorthodoxly enters cultured neurons and passes through the blood-brain barrier. In vivo, intracranially or peripherally delivered C-CDNF improves motor deficits, protects DA neurons, and restores motor behavior in a rat model of PD. Subcutaneous C-CDNF also protects MNs and reduces microglial activation in an ALS model. Based on our findings, beginning C-CDNF treatment soon after diagnosis is anticipated to delay progression of PD and ALS, thereby improving treatment outcome. Thus, systemic delivery of C-CDNF should simplify the administration of protein-based therapeutics to patients while reducing treatment risk and financial burden for patients and families.

Functional connectivity of thalamic nuclei during sensorimotor task-based fMRI at 9.4 Tesla
The thalamus is the brain's central communication hub, playing a key role in processing and relaying sensorimotor and cognitive information between the cerebral cortex and other brain regions. It consists of specific and non-specific nuclei, each with a different role. Specific thalamic nuclei relay sensory and motor information to specific cortical and subcortical regions to ensure precise communication. In contrast, non-specific thalamic nuclei are involved in general functions such as attention or consciousness through broader and less targeted connections. In the present study, we aimed to investigate the functional connectivity patterns of the thalamic nuclei identified in our previous study as being involved in motor (finger-tapping) and sensory (finger-touch) tasks. The results of this study show that thalamic nuclei are not static hubs with a predefined role in neural signal processing, as they show different task-specific functional connectivity patterns in the anterior, middle, lateral, and posterior thalamic nuclei. Instead, they are all functional hubs that can flexibly change their connections to other brain regions in response to task demands. This work has important implications for understanding task-dependent functional connectivity between thalamic nuclei and different brain regions using task-based fMRI at 9.4 Tesla.

A lateralized pathway for associating nutrients with flavors
Animals learn about the external world, in part, via interoceptive signals. For example, the nutrient content of food is first estimated in the mouth, in the form of flavor, and then measured again via slower signals from the gut. How these signals from the mouth and gut are integrated to drive learning is unknown. Here we identify a lateralized dopamine pathway that is specialized for learning about the nutrient content of food. We show that dopamine neurons in the ventral tegmental area (VTADA) are necessary for associating nutrients with flavors, but that this does not involve canonical dopamine targets in the striatum. Instead, post-ingestive nutrients trigger DA release selectively in a small region of the anterior basolateral amygdala (BLA). Remarkably, this hotspot for nutrient-triggered DA release is localized to the left side of the brain in both mice and humans, revealing that the DA system is functionally lateralized. We identify the gut sensors that are responsible for nutrient-triggered DA release; show that they selectively activate BLA-projecting DA neurons defined by expression of cholecystokinin (CCK); and demonstrate that stimulation of DA axon terminals in the anterior BLA drives flavor-nutrient learning but not other aspects of behavior. Two-photon imaging of neurons in the left anterior BLA reveals that they encode food flavors, and these representations are strengthened by post-ingestive nutrients, whereas silencing of these neurons prevents flavor-nutrient learning. These findings establish a neural basis for how animals learn about the nutrient content of their food. They also reveal unexpectedly that post-ingestive nutrients are differentially represented on the right and left sides of the brain.

The sensorimotor basis of subjective experience in social synchronization behavior
The sensorimotor contingency (SMC) theory addresses action and perception as constitutive factors to one another - what is being perceived depends on what we do and vice versa. Accordingly, perception is seen as an active process of probing the environment and receiving feedback from it. These action- effect patterns may also be a predominant factor in social interactions. They can manifest in the phenomenon of synchronization, for example, when applause, gait, or posture in conversations synchronize unintentionally and form the basis of the concept of socially deployed sensorimotor contingencies (socSMCs). In this study, we introduce and compare measures used to study complex systems in order to quantify the information-theoretic basis of socSMCs. Two human participants had to synchronize arm movements in a full body virtual reality (VR) environment with each other. We aimed to evaluate the information-theoretic measures transfer entropy and mutual information in this complex motion synchronization task. Furthermore, in our experiment participants shared a mutual sensation of synchronicity and creativity for their interaction solely based on their movements. These subjective ratings of synchronicity and creativity can be predicted using transfer entropy and mutual information, showing that informational coupling between agents is relevant to subjective experience of interaction as described by the concept of socSMCs.

Impact of Aging on Theta-Phase Gamma-Amplitude Coupling During Learning: A Multivariate Analysis
Aging is associated with cognitive decline and memory impairment, but the underlying neural mechanisms remain unclear. Phase-amplitude coupling (PAC) between mid-frontal theta (5 Hz) and occipital gamma (>30 Hz) oscillations is a proposed marker for parallel storage of multiple items in working memory. However, research has mainly focused on young individuals and epilepsy patients, with only a few studies on aging populations. Moreover, these studies have relied on univariate PAC methods, which can be flawed by potential spurious or biased PAC estimates due to non-stationarity of EEG signals. Additionally, these methods typically assess PAC at the level of individual electrodes, potentially overlooking the broader functional significance of theta-gamma coupling in coordinating neural activity across distant brain regions. To address these gaps, we employed multivariate PAC (mPAC) through generalized eigendecomposition (GED) analysis, which avoids the pitfalls of non-sinusoidal oscillations. 113 young and 117 older healthy participants engaged in a sequence learning paradigm (6423 sequence repetitions, 55'944 stimuli), in which they learned a fixed sequence of visual stimuli over repeated observations, allowing us to track the mPAC during the incremental process of learning. Behavioral results revealed that younger participants learned significantly faster than older participants. Neurophysiological data showed that mPAC increased over the course of learning in both age groups and could identify fast and slow learners. However, older participants exhibited lower mPAC compared to younger counterparts, which suggest compromised parallel storage of items in working memory in older age. Finally, stratification analysis revealed that mPAC effects persist across performance groups with similar mid-frontal theta levels, suggesting that theta alone does not account for these effects. These findings shed light on the age-related differences in memory formation processes and may guide interventions to enhance memory performance in older adults and slow learners.

GsMTx-4 Reduces Mechanosensitivity in a Model of Schwannomatosis-related Pain
Patients with schwannomatosis (SWN) develop multiple tumors along major peripheral nerves, with most experiencing significant pain, though each patient's symptoms are unique. Neuropathic, nociceptive, and inflammatory pain types have been reported, but many patients describe severe pain when a schwannoma is palpated or even lightly touched. Currently, the only effective treatment for pain relief is surgical removal. We are investigating the root causes of tumor-induced pain. In some cases, tumor growth increases pressure on nearby nerves, resulting in pain. Additionally, schwannoma cells in culture secrete proinflammatory cytokines into the surrounding medium. This conditioned medium (CM) sensitizes sensory neurons to painful stimuli both in vitro and in vivo. When injected into the glabrous skin of a mouse hindpaw, CM from painful schwannomas increases neuron sensitivity to light touch, as demonstrated by a fourfold reduction in paw withdrawal threshold (measured using the Von Frey assay) one hour post-injection (p = 0.006), with effects persisting for 24 hours (p = 0.002).We hypothesize that this increase in sensitivity is linked to mechanosensitive ion channels (MSCs), which detect pressure and stretch. These channels can be blocked by the peptide GsMTx-4. This peptide penetrates deeper into cell membranes under mechanical pressure to block MSCs from opening without affecting other ion channels. When co-injected with CM into the mouse hindpaw, 10 M GsMTx-4 prevents heightened sensitivity to light touch. Moreover, GsMTx-4 can reverse hyperalgesia, restoring withdrawal thresholds to baseline levels. Thus, local injection of GsMTx-4 near painful tumors presents a promising, minimally invasive therapeutic approach for SWN patients.

Offset analgesia and onset hyperalgesia: A comparison between women with non-suicidal self-injury and healthy women
Individuals who engage in non-suicidal self-injury (NSSI) exhibit reduced pain sensitivity compared to the general population. It has been argued that this hypoalgesic characteristic may be attributable to hyper-effective pain modulation; however, empirical support for this theory remains inconsistent. The aim of the study was to use a combined offset analgesia (OA) and onset hyperalgesia (OH) protocol to investigate if women with NSSI have a propensity to inhibit nociceptive signals to a higher degree (stronger OA response) and facilitate nociceptive signals to a lesser degree (weaker OH response), compared to a control group. Data was collected from 76 women, 18-35 age (37 with NSSI and 39 healthy controls). The OA and OH protocol was combined with functional magnetic resonance imaging (fMRI). The NSSI group displayed a weaker OH response, compared to the control group. This suggests that women with NSSI do not facilitate nociceptive signals to the same extent as healthy women. However, there were no significant differences between the groups regarding OA. Across all participants, we observed stronger activation in the primary sensory cortex during the OH condition, compared to the control condition. The results offer partial support for the general hypothesis that women with NSSI demonstrate enhanced pain modulation.

Transcriptomic and spatial GABAergic neuron subtypes in zona incerta mediate distinct innate behaviors
Understanding the anatomical connection and behaviors of transcriptomic neuron subtypes is critical to delineating cell type-specific functions in the brain. Here we integrated single-nucleus transcriptomic sequencing, in vivo circuit mapping, optogenetic and chemogenetic approaches to dissect the molecular identity and function of heterogeneous GABAergic neuron populations in the zona incerta (ZI) in mice, a region involved in modulating various behaviors. By microdissecting ZI for transcriptomic and spatial gene expression analyses, our results revealed two non-overlapping Ecel1- and Pde11a-expressing GABAergic neurons with dominant expression in the rostral and medial zona incerta (ZIrEcel1 and ZImPde11a), respectively. The GABAergic projection from ZIrEcel1 to periaqueductal gray mediates self-grooming, while the GABAergic projection from ZImPde11a to the oral part of pontine reticular formation promotes transition from sleep to wakefulness. Together, our results revealed the molecular markers, spatial organization and specific neuronal circuits of two discrete GABAergic projection neuron populations in segregated subregions of the ZI that mediate distinct innate behaviors, advancing our understanding of the functional organization of the brain.

Brain bases for navigating acoustic features
Whether physical navigation shares neural substrates with mental travel in other behaviourally relevant domains is debated. With respect to sound, pure tone working memory in humans elicits hippocampal as well as auditory cortical and inferior frontal activity, and rodent work suggests that hippocampal cells that usually track an animal's physical location can also map to tone frequency when task-relevant. We generated a sound dimension based on the density of random-frequency tones in a stack, resulting in a percept ranging from low- ("beepy") to high-density ("noisy"). We established that unlike tone frequency, which listeners automatically associate with vertical position, this density dimension elicited no consistent spatial mapping. During functional magnetic resonance imaging, human participants (both sexes) held in mind the density of a series of tone stacks and, after a short maintenance period, adjusted further stacks to match the target ("navigation"). Density was represented most strongly in bilateral non-primary auditory cortex, specifically bilateral planum polare. Encoding and maintenance activity in hippocampus, inferior frontal gyrus, planum polare and posterior cingulate was positively associated with subsequent navigation success. Bilateral inferior frontal gyrus and hippocampus were among regions with elevated activity during navigation, compared to a parity-judgment condition with closely matched acoustics and motor demands. Bilateral orbitofrontal cortex was more active when navigation was toward a target density than when participants adjusted density in a control condition with no particular target. We find that self-initiated travel along a non-spatial auditory dimension engages a brain system overlapping with that supporting physical navigation.

Cell-generated mechanical forces play a role in epileptogenesis after injury
Traumatic brain injury (TBI) is associated with a significantly increased risk of epilepsy. One of the consequences of severe TBI is progressive brain atrophy, which is frequently characterized by organized tissue retraction. Retraction is an active process synchronized by mechanical interactions between surviving cells. This results in unbalanced mechanical forces acting on surviving neurons, potentially activating mechanotransduction and leading to hyperexcitability. This novel mechanism of epileptogenesis was examined in organotypic hippocampal cultures, which develop spontaneous seizure-like activity in vitro. Cell-generated forces in this model resulted in contraction of hippocampal tissue. Artificial imbalances in mechanical forces were introduced by placing cultured slices on surfaces with adhesive and non-adhesive regions. This modeled disbalance in mechanical forces that may occur in the brain after trauma. Portions of the slices that were not stabilized by substrate adhesion underwent increased contraction and compaction, revealing the presence of cell-generated forces capable of shaping tissue geometry. Changes in tissue geometry were followed by excitability changes that were specific to hippocampal sub-region and orientation of contractile forces relative to pyramidal cell apical-basal axis. Results of this study suggest that imbalanced cell-generated forces contribute to development of epilepsy, and that force imbalance may represent a novel mechanism of epileptogenesis after trauma.

Real-world visual search goes beyond eye movements: Active searchers select 3D scene viewpoints too
Visual search is a ubiquitous task; people search for objects on a daily basis. However, the majority of the existing visual search literature focuses on passive search on a 2D computer screen, a far cry from emulating a real-world environment. Search is a real-world task that involves active observation. Search targets may be occluded, completely out of the observer's line of sight, or oriented in unconventional ways. This is typically mitigated by actively selecting viewpoints, an important aspect of search behaviour with limited scope on a computer screen. Our goal was to explore viewpoint selection in active visual search. Subject eye and head movements were tracked as they moved freely while searching for toy objects in a controlled 3-dimensional environment, yielding the first such record of search-driven viewpoint selection. We found that subjects utilized their full range of eye and head motion to move from viewpoint to viewpoint, apparently employing a variety of objectives including changing viewing height and pose depending on object 3D pose. Subjects were also adept at selecting unobstructed views to search through otherwise occluded areas with objects. Furthermore, subjects completed the search task with high accuracy, even with no training on the environment. Although no learning was found in terms of accuracy over the duration of the experiment, increases in efficiency were found for other metrics such as response time, number of fixations, and distance travelled, particularly in target present trials where the target was not visible from the starting location. These results paint the story of a visual system that selects and moves to useful and informative views to facilitate the successful execution of an active visual search task, and stresses the significance of active vision research in understanding how vision is used in naturalistic environments.

Self-supervised image restoration in coherent X-rayneuronal microscopy
Coherent X-ray microscopy is emerging as a transformative technology for neuronal imaging, with the potential to offer a scalable solution for reconstruction of neural circuits in millimeter sized tissue volumes. Specifically, X-ray holographic nanotomography (XNH) brings together outstanding capabilities in terms of contrast, spatial resolution and data acquisition speed. While recent XNH developments already enabled generating valuable datasets for neurosciences, a major challenge for reconstruction of neural circuits remained overcoming resolving power limits to distinguish smaller neurites and synapses in the reconstructed volumes. Here we present a self-supervised image restoration approach that improves simultaneously spatial resolution, contrast, and data acquisition speed. This enables revealing synapses with XNH, marking a major milestone in the quest for generating connectomes of full mammalian brains. We demonstrate that this method is effective for various types of neuronal tissues and acquisition schemes. We propose a scalable implementation compatible with multi-terabyte image volumes. Altogether, this work brings large scale X-ray nanotomography to a new precision level.

Feedback and feedforward control are differentially delayed in cerebellar ataxia
Damage to the cerebellum can cause ataxia, a condition associated with impaired movement coordination. Typically, coordinated movement relies on a combination of anticipatory mechanisms (specifically, feedforward control) and corrective mechanisms (embodied by feedback control). Here, we show that in 3D reaching in VR, ataxia preserves the visuomotor feedforward and feedback control structure compared to the control group. However, the ataxia group exhibits a small increase in feedback delay (~20 ms) and a substantial increase in feedforward delay (~70 ms) together with a reduced feedback gain (~25% lower). Our results suggest that the feedforward and feedback pathways remain largely intact in ataxia, but that time delay deficits and temoral discoordination amongst these control pathways may contribute to the disorder. We also find that providing a preview---analogous to driving on a clear night and seeing the road ahead vs.~driving in the fog---improves tracking performance in the ataxia group, although the control group was significantly better able to exploit this preview information. Overall, our results indicate that the feedforward control and preview utilization are relatively well-preserved in individuals with cerebellar ataxia, and that preview could potentially be leveraged to enhance the feedforward performance of those with ataxia.

SEX-DEPENDENT MODULATION OF BEHAVIORAL ALLOCATION VIA VENTRAL TEGMENTAL AREA-NUCLEUS ACCUMBENS SHELL CIRCUITRY
Diagnostic criteria for substance use disorder, cocaine type (i.e., cocaine use disorder), outlined in the 5th edition of the Diagnostic and Statistical Manual, imply that the disorder arises, at least in part, from the maladaptive allocation of behavior to drug use. To date, however, the neural circuits involved in the allocation of behavior have not been systematically evaluated. Herein, a chemogenetics approach (i.e., designer receptors exclusively activated by designer drugs (DREADDs)) was utilized in combination with a concurrent choice self-administration experimental paradigm to evaluate the role of the mesolimbic neurocircuit in the allocation of behavior. Pharmacological activation of hM3D(Gq) DREADDs in neurons projecting from the ventral tegmental area (VTA) to the nucleus accumbens (AcbSh) induced a sex-dependent shift in the allocation of behavior in rodents transduced with DREADDs. Specifically, male DREADDs animals exhibited a robust increase in responding for a natural (i.e., sucrose) reward following pharmacological activation of the VTA-AcbSh circuit; female DREADDs rodents, in sharp contrast, displayed a prominent decrease in drug-reinforced (i.e., cocaine) responding. The sequential activation of hM3D(Gq) and KORD DREADDs within the same neuronal population validated the role of the VTA-AcbSh circuit in reinforced responding for concurrently available natural and drug rewards. Collectively, the VTA-AcbSh circuit is fundamentally involved in behavioral allocation affording a key target for the development of novel pharmacotherapies.

3-D Reconstruction of Fingertip Deformation during Contact Initiation
Dexterous manipulations rely on tactile feedback from the fingertips, which provides crucial information about contact events, object geometry, interaction forces, friction, and more. Accurately measuring skin deformations during tactile interactions can shed light on the mechanics behind such feedback. To address this, we developed a novel setup using 3-D digital image correlation (DIC) to both reconstruct the bulk deformation and local surface skin deformation of the fingertip under natural loading conditions. Here, we studied the local spatiotemporal evolution of the skin surface during contact initiation. We showed that, as soon as contact occurs, the skin surface deforms very rapidly and exhibits high compliance at low forces (<0.05 N). As loading and thus the contact area increases, a localized deformation front forms just ahead of the moving contact boundary. Consequently, substantial deformation extending beyond the contact interface was observed, with maximal amplitudes ranging from 5% to 10% at 5 N, close to the border of the contact. Furthermore, we found that friction influences the partial slip caused by these deformations during contact initiation, as previously suggested. Our setup provides a powerful tool to get new insights into the mechanics of touch and opens avenues for a deeper understanding of tactile afferent encoding.

Calcium channel-coupled transcription factors facilitate direct nuclear signaling
VGCCs play crucial roles within the CNS, in maintaining cell excitability, enabling activity-dependent neuronal development, and forming long-term memory by regulating Ca2+ influx. The intracellular carboxyl-terminal domains of VGCC 1 subunits help regulate VGCC function. Emerging evidence suggests that some VGCC C-termini have functions independent of channel gating and exist as stable proteins. Here, we demonstrate that all VGCC gene family members express bicistronic mRNA transcripts that produce functionally distinct C-terminal proteins (CTPs) in tandem with full-length VGCC 1 subunits. Two of these CTPs, 1CCT and 1ACT, cycle to and from the nucleus in a Ca2+- and calmodulin-dependent fashion. 1CCT, 1ACT, and 1HCT regulate chromatin accessibility and/or bind directly to genes, regulating gene networks involved in neuronal differentiation and synaptic function in a Ca2+-dependent manner. This study elucidates a conserved process of coordinated protein expression within the VGCC family, coupling the channel function with VGCC C-terminal transcription factors.

How Autism Impacts Children's Working Memory for Faces
This study investigates how visual working memory (WM) functions in children with Autism Spectrum Disorder (ASD) compared to typically developing (TD) children aged 7 to 12, focusing on their ability to remember faces. The primary goal was to understand the differences in visual WM performance between these two groups and identify the sources of memory recall errors. Findings indicate that children with ASD demonstrated significantly poorer visual WM compared to their TD counterparts, particularly in the precision of memory recall. Despite this difference, both groups exhibited similar rates of random guessing, suggesting that the challenges faced by children with ASD are more about accuracy than guessing strategy. This research sheds light on the specific deficits in visual WM associated with ASD, enhancing our understanding of the cognitive mechanisms at play and highlighting potential areas for targeted interventions to support these children.

Rigid Control of Motor Unit Firing Rates in the Human Tibialis Anterior Muscle Persists during Neurofeedback
The conventional framework of motor-unit (MU) control assumes that MUs in a MU pool are constrained by a fixed recruitment order and a common input. This rigid-control framework has been challenged by recent findings suggesting that MU activity could be flexibly modulated, potentially mediated by descending cortical inputs. In this study, rather than evaluating flexibility from the perspective of recruitment thresholds, we investigated control flexibility by assessing if human participants can voluntarily modulate MU firing rates beyond rigid-control constraints. Specifically, we examined whether participants could voluntarily modulate the firing rates of a pair of MUs from the tibialis anterior muscle during real-time feedback. Two tasks involving target-reach with different visual feedback derived from the MUs firing rates were conducted. In both tasks, there was no evidence that participants were able to change MU firing rates in a way that would violate rigid control robustly. Our findings demonstrate limited flexibility in MU control in human tibialis anterior muscle within single-session training, even when real-time MU activity feedback was provided. The results suggest that MU flexibility is not inherently present in the human lower limb.

Sex Specific Attenuation of Reward Preference
Estradiol receptor signaling has a sex-specific impact on the brain's reward pathways, enhancing cocaine reinforcement in females but not in males. Selective activation of G-Protein Coupled Estradiol Receptor 1 (GPER-1) in the dorsolateral striatum (DLS) attenuates the reinforcing effects of 0.1% saccharin (SACC) and cocaine in males but not females. This study investigated GPER-1 activation in the DLS and systemically using the GPER-1 agonist, G1, to assess its effect on SACC and cocaine preference in male and female rats. Five experiments were conducted using gonad-intact and gonadectomized animals to determine dose-response effects and the influence of circulating hormones. Intra-DLS GPER-1 activation with 20% G1 selectively reduced SACC preference in intact males but not females, while higher and lower concentrations had no effect. Systemic G1 administration attenuated cocaine-induced conditioned place preference (CPP) in both sexes in a dose-dependent way. Interestingly, systemic administration of G1 did not alter SACC preference in either sex, regardless of the presence or absence of gonadal hormones. These findings suggest that GPER-1 activation influences reward processing in a site, reward, and sex-dependent manner.

Altered Functional Network Energy Across Multiscale Brain Networks in Preterm vs. Full-Term Subjects: Insights from the Adolescent Brain Cognitive Development (ABCD) Study
Infants born premature, or preterm, can experience altered brain connectivity, due in part to incomplete brain development at the time of parturition. Research has also shown structural and functional differences in the brain that persist in these individuals as they enter adolescence when compared to peers who were fully mature at birth. In this study, we examined functional network energy across multiscale functional connectivity in approximately 4600 adolescents from the Adolescent Brain Cognitive Development (ABCD) study who were either preterm or full term at birth. We identified three key brain networks that show significant differences in network energy between preterm and full-term subjects. These networks include the visual network (comprising the occipitotemporal and occipital subnetworks), the sensorimotor network, and the high cognitive network (including the temporoparietal and frontal subnetworks). Additionally, it was demonstrated that full-term subjects exhibit greater instability, leading to more dynamic reconfiguration of functional brain information and increased flexibility across the three identified canonical brain networks compared to preterm subjects. In contrast, those born prematurely show more stable networks but less dynamic and flexible organization of functional brain information within these key canonical networks. In summary, measuring multiscale functional network energy offered insights into the stability of canonical brain networks associated with subjects born prematurely. These findings enhance our understanding of how early birth impacts brain development.

Pallidal prototypic neuron and astrocyte activities regulate flexible reward-seeking behaviors
Behavioral flexibility allows animals to adjust actions to changing environments. While the basal ganglia are critical for adaptation, the specific role of the external globus pallidus (GPe) is unclear. This study examined the contributions of two major GPe cell types--prototypic neurons projecting to the subthalamic nucleus (ProtoGPe[->]STN neurons) and astrocytes--to behavioral flexibility. Using longitudinal operant conditioning with context reversals, we found that ProtoGPe[->]STN neurons dynamically represent contextual information correlating with behavioral optimality. In contrast, GPe astrocytes exhibited gradual contextual encoding independent of performance. Deleting ProtoGPe[->]STN neurons impaired adaptive responses to changing action-outcome contingencies without altering initial reward-seeking acquisition, highlighting their specific role in enabling behavioral flexibility. Furthermore, we discovered that ProtoGPe[->]STN neurons integrate inhibitory striatal and excitatory subthalamic inputs, modulating downstream basal ganglia circuits to support flexible behavior. This research elucidates the complementary roles of ProtoGPe[->]STN neurons and astrocytes in cellular mechanisms of flexible reward-seeking behavior.

Molecularly-guided spatial proteomics captures single-cell identity and heterogeneity of the nervous system
Single-cell proteomics is an emerging field with significant potential to characterize heterogeneity within biological tissues. It offers complementary insights to single-cell transcriptomics by revealing unbiased proteomic changes downstream of the transcriptome. Recent advancements have focused on enhancing proteome coverage and depth, mostly in cultured cell lines, and a few recent studies have explored the potential of analyzing tissue micro-samples but were limited to homogenous peripheral tissues. In this current work, we utilize the power of spatial single cell-proteomics through immunostaining-guided laser capture microdissection (LCM) coupled with LC-MS to investigate the heterogenous central nervous system. We used this method to compare neuronal populations from cortex and substantia nigra, two brain regions associated with motor and cognitive function and various neurological disorders. Moreover, we used the technique to understand the neuroimmune changes associated with stab wound injury. Finally, we focus our application on the peripheral nervous system, where we compare the proteome of the myenteric plexus cell ganglion to the nerve bundle. This study demonstrates the utility of spatial single-cell proteomics in neuroscience research toward understanding fundamental biology and the molecular drivers of neurological conditions.

Dopaminergic mechanisms of dynamical social specialization in mouse microsocieties
Social organization and division of labor are fundamental to animal societies, but how do these structures emerge from individual interactions, and what role does neuromodulation play in shaping them? Using behavioral tracking in a semi-natural environment, neural recordings, and computational models integrating reinforcement and social learning, we show that groups of three isogenic mice spontaneously develop specialized roles while solving a foraging task requiring individual decisions under social constraints. Moreover, these roles are shaped by dopaminergic activity in the ventral tegmental area. Strikingly, despite minor sex-differences in behavior when mice were tested alone, male triads formed stable worker-scrounger relationships driven by competition, whereas female triads adopted uniform, cooperative strategies. Model analysis revealed how intra- and inter-sex parameter differences in resource exploitation, combined with contingent and dynamic social interactions, drive behavioral specialization and labor division. Most notably, it highlighted how contingency, amplified by competition, magnifies individual differences and shapes social profiles. The plastic, adaptive nature of social organization within triads was confirmed by manipulating dopaminergic cell activity, which reshaped social roles and altered group structure. Our findings support a feedback loop where social context shapes neural states, which in turn reinforce behavioral specialization and stabilize social structures.

Generation of C9orf72 repeat knock-in iPSC lines for modelling ALS and FTD
Induced pluripotent stem cell (iPSC) models are powerful tools for neurodegenerative disease modelling, as they allow mechanistic studies in a human genetic environment and they can be differentiated into a range of neuronal and non-neuronal cells. However, these models come with inherent challenges due to line-to-line and clonal variability. To combat this issue, the iPSC Neurodegenerative Disease Initiative (iNDI) has generated an iPSC repository using a single clonal reference line, KOLF2.1J, into which disease-causing mutations and revertants are introduced via gene editing. Here we describe the generation and validation of lines carrying the most common causative mutation for amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD), a repeat expansion in the C9orf72 gene, for the iNDI collection of neurodegenerative iPSC models. We demonstrate that these C9orf72 knock-in lines differentiate efficiently into neurons and display characteristic C9orf72-associated pathologies, including reduced C9orf72 levels and the presence of dipeptide repeat proteins (DPRs) and RNA foci, which increase in abundance over time in culture. These pathologies are not present in revertant cells lacking the repeat expansion. These repeat expansion and revertant cell lines are now available to academic and for-profit institutions through the JAX iPS cell repository and will help to facilitate and standardise iPSC-based ALS/FTD research.

From ancient fears to airborne threats: fMRI insights into neural fear responses
Threat perception is a fundamental aspect of human cognition, shaped by evolutionary pressures and modern environmental demands. While ancestral threats (e.g., snakes) have been shown to elicit stronger neural responses than modern threats (e.g., guns), less is known about how the brain processes airborne threats, such as depictions of individuals wearing face masks. This fMRI study investigates neural responses to ancestral, modern, and airborne threats to identify shared and distinct activation patterns. Sixty participants viewed visual stimuli from the three categories while undergoing fMRI scanning. Results showed heightened activation in the fear-processing network for all affective stimuli. In addition, activation of the ventral attention network was found for the ancestral threats. Modern threats elicited less intense responses, primarily engaging cortical regions associated with context-specific analysis. Notably, airborne threats elicited neural responses of similar intensity to ancestral threats but activated cortical regions overlapping with those for modern threats. This dual pattern highlights the brain's capacity to integrate evolutionary biases with socially constructed threat awareness. These findings underscore the importance of recognizing airborne threats as a unique category of threat processing, with implications for public health and mental well-being.

Aseptic, semi-sealed cranial chamber implants for chronic multi-channel neurochemical and electrophysiological neural recording in nonhuman primates
We developed an implantable neural interface for monitoring both chemical and electrical forms of brain activity in monkeys that maintains aseptic properties for year-long periods while leveraging the modular functions (e.g., sensor moveability) provided by a chamber system. Invasive electrophysiological recordings, especially in subcortical structures from nonhuman primates usually involves implanting electrodes into the brain through a skull-mounted chamber. These electrodes may be attached temporarily for several hours of recording, or permanently. Permanent attachments are favorable to allow for sealing the chamber completely from externally originating pathogenic species that can infiltrate and compromise the health of the animal. A sealed chamber also reduces the need for frequent chamber cleaning required to minimize the accumulation of pathogenic organisms. However, neurochemical measurements require specialized electrodes with extremely fragile carbon fiber tips and are not compatible with recently developed sealed chamber systems. Here, we leveraged osseointegrating materials and hermetic sealing strategies to enable both neurochemical and electrical neural activity measurements from a sealed chamber with an aspirating port for culturing chamber fluid to ensure an aseptic environment. The system was shown to provide successful recordings of neural activity in two monkeys while maintaining negative bacteria culture results for over a year post-implant.

SNARE Protein Ykt6 Drives AMPAR Insertion at Synaptic Terminals During LTP
Ykt6 is a SNARE protein essential for vesicular fusion along the secretory pathway. It has been implicated in -synuclein (Syn) pathology and is critically involved in -synuclein-dependent dementias. Syn disrupts AMPA receptor (AMPAR) expression at synaptic terminals, which is key for long-term potentiation (LTP) - the cellular basis of memory and learning. AMPAR expression at synaptic terminals relies on the secretory pathway; however, the role of Ykt6 in LTP and its perturbation under Syn pathological conditions remain unexplored. Here, we demonstrate that Ykt6 is highly expressed in the hippocampus of the mammalian brain, relocalizes to synaptic terminals during LTP, and regulates GluA1 surface expression. Moreover, we found that Ykt6 modulates the synaptic vesicular pools and dendritic arborization. Taken together, our findings establish Ykt6 as an essential SNARE for hippocampal neural function during LTP, with significant implications for -synuclein-dependent dementias.

Neurons in the inferior colliculus use multiplexing to encode features of frequency-modulated sweeps
Within the central auditory pathway, the inferior colliculus (IC) is a critical integration center for ascending sound information. Previous studies have shown that many IC neurons exhibit receptive fields for individual features of auditory stimuli, such as sound frequency, intensity, and location, but growing evidence suggests that some IC neurons may multiplex features of sound. Here, we used in vivo juxtacellular recordings in awake, head-fixed mice to examine how IC neurons responded to frequency-modulated sweeps that varied in speed, direction, intensity, and frequency range. We then applied machine learning methods to determine how individual IC neurons encode features of FM sweeps. We found that individual IC neurons multiplex FM sweep features using various strategies including spike timing, distribution of inter-spike intervals, and first spike latency. In addition, we found that decoding accuracy for sweep direction can vary with sweep speed and frequency range, suggesting the presence of mixed selectivity in single neurons. Accordingly, using static receptive fields for direction alone yielded poor predictions of neuron responses to vocalizations that contain simple frequency changes. Lastly, we showed that encoding strategies varied across individual neurons, resulting in a highly informative population response for FM sweep features. Together, our results suggest that multiplexing sound features is a common mechanism used by IC neurons to represent complex sounds.

Astrocyte gap junctions and Kir channels contribute to K+ buffering and regulate neuronal excitability
Astrocytes are connected in a functional syncytium via gap junctions, which is thought to contribute to maintenance of extracellular K+ homeostasis. The prevailing hypothesis is that K+ released during neuronal firing is taken up by astrocytes via Kir channels and then distributed among neighboring astrocytes via gap junctions. Previous reports examining the role of Kir channels and gap junctions have shown both hyperexcitability and depression when each mechanism is blocked. Here, we tested the effect of blocking Kir channels and gap junctions, both independently and simultaneously, on field activity of cortical slices in response to a 3 s, 20 Hz stimulation train. Independently blocking either Kir channels or gap junctions increased the amplitude of the first fEPSC (field excitatory post-synaptic current) in response to a stimulation train, followed by suppression of fEPSCs during sustained stimulation. Surprisingly, blocking both gap junctions and Kir channels enhanced the suppression of neuronal activity, resulting in a ~75% decrease in fiber volley (pre-synaptic action potentials) amplitude in the first response, followed by a fast and strong suppression of sustained fEPSCs. Our results demonstrate that blocking Kir channels and gap junctions can increase the excitability of neurons when firing is sparse, but suppression results when the firing frequency is increased to cortical physiological ranges. This suggest that K+ buffering via Kir and gap junctions, likely mediated by astrocytes, together play a critical role in maintaining neuronal excitability, particularly during sustained activity.

Sex specific disruptions in Protein Kinase Cγ signaling in a mouse model of Spinocerebellar Ataxia Type 14
Spinocerebellar Ataxia Type 14 (SCA14) is an autosomal dominant neurodegenerative disease caused by mutations in the gene encoding protein kinase C gamma (PKC{gamma}), a Ca2+/diacylglycerol (DG)-dependent serine/threonine kinase dominantly expressed in cerebellar Purkinje cells. These mutations impair autoinhibitory constraints to increase the basal activity of the kinase, resulting in deficits in the cerebellum that are not observed upon simple deletion of the gene, and severe ataxia. To better understand the phenotypic impact of aberrant PKC{gamma} signaling in disease pathology, we developed a knock-in murine model of the SCA14 mutation {Delta}F48 in PKC{gamma}. This fully penetrant mutation is severe in humans and is mechanistically informative as it has high basal activity but is unresponsive to agonist stimulation. Genetic, behavioral, and molecular testing revealed that {Delta}F48 PKC{gamma} SCA14 mice have ataxia related phenotypes and an altered cerebellar phosphoproteome, effects that are more severe in male mice. Analysis of existing human data reveal that SCA14 has a significantly earlier age of onset for males compared with females. Our data from this clinically relevant mutation suggest that enhanced basal activity of PKC{gamma} is necessary and sufficient to cause ataxia and that treatment strategies to modulate aberrant PKC{gamma} may be particularly beneficial in males.

Post-Synaptic Density Proteins in Oligodendrocytes are Required for Activity-Dependent Myelin Sheath Growth
Compelling evidence demonstrates a functional link between neuronal activity and myelination, highlighting the vital importance of axon-oligodendrocyte crosstalk in myelin physiology and function. However, how neuronal activity is relayed to oligodendroglia to regulate myelin formation remains not fully understood. Here, we aimed to characterize how that myelination is regulated by glutamate vesicular release in zebrafish spinal cord. We compared oligodendrocyte precursor cells (OPCs) and myelinating oligodendrocytes (mOLs) for their close apposition with pre-synaptic boutons and found that these are increased in number on mOLs during myelin internode elongation. Consistently, mOLs show more pre-synaptic boutons during myelin internode elongation compared to OPCs. In addition, we also found that oligodendroglial cells express the post-synaptic density protein 95 (PSD-95) along punctated domains, regardless of their differentiation stage. Genetically targeted PSD-95-GFP expression in oligodendroglia revealed post-synaptic-like domains along their processes and sheaths, which are contacted by axonal pre-synaptic varicosities. These contacts are increased in mOLs. Importantly, CRISPR-Cas9 mediated deletion of dlg4 in oligodendroglia impairs myelin sheath growth, in vivo. Overall, our data indicate that PSD-95 is a key component of axons to oligodendrocytes neurotransmission that regulates myelin sheath growth.

Joint visual-vestibular computation of head direction and reflexive eye movement
Several cognitive maps have been identified, but what sensory signals drive them and how these are combined are not well understood. One such map, the head-direction representation, is believed to be primarily driven by vestibular motion signals in mammals. Here, we combine in vivo imaging of neuronal activity, genetic perturbation of neuronal circuits, behavioral testing, and theoretical modeling to show that the representation of head direction in mouse is driven by not only vestibular but also visual motion signals: both are essential, and the latter, originating in direction-selective retinal ganglion cell activity, dominates at low speeds. We show that, correspondingly, visual perturbations alter navigational behavior that relies on head-direction computation. Finally, we find that head-direction representation and the slow phase of reflexive eye movement are tightly correlated, and we propose a theoretical model that elucidates their emergence from coupled visual and vestibular processes. Our results suggest that the brain's estimate of head direction is built on an oculomotor reflex pathway driven by both visual and vestibular signals.

A novel model of paclitaxel-induced peripheral neuropathy produces a clinically relevant phenotype in mice
One of the most common adverse side effects of chemotherapeutics is chemotherapy-induced peripheral neuropathy (CIPN). Paclitaxel, a highly effective chemotherapeutic, is associated with a high incidence of paclitaxel-induced peripheral neuropathy (PIPN) that persists for over a year in 64% of patients and worsens with cumulative PTX dose. Patients experiencing PIPN may reduce the dosage of chemotherapy or halt treatment due to this pain. Current preclinical models have improved our understanding of PIPN but have been ineffective in generating translational therapeutic options. These models administer a single cycle of PTX to induce a PIPN phenotype of mechanical and cold hypersensitivity that resolves within 28 days. However, this does not mirror the clinical dosing regimen or the patient experience of CIPN. In this study, we conduct a comprehensive and longitudinal behavioral profile of our novel model of PIPN in mice where three consecutive cycles of PTX (4 mg/kg, 4 doses/cycle) are given to mimic the clinical administration. Repeated cycles of PTX caused long-lasting mechanical and cold hypersensitivity in male and female C57Bl/6J mice that mirrors clinical observations of persistent CIPN without causing detrimental effects to rodent overall health, normal rodent behavior, or motor function. Our findings support the use of this translational model to facilitate a better understanding of PIPN and the development of effective treatment options. Improved pain management will enable the completion of cancer treatment, decrease health care expenditure, decrease mortality, and improve the quality of life for cancer patients and survivors.

Molecular and morphological circuitry of the octopus sucker ganglion
The octopus sucker is a profoundly complex sensorimotor structure. Each of the hundreds of suckers that line the octopus arm can move independently or in concert with one another. These suckers also contain an intricate sensory epithelium, enriched with chemotactile receptors. Much of the massive nervous system embedded in the octopus arm mediates control of the suckers. Each arm houses a large axial nerve cord (ANC), which features local enlargements corresponding to each sucker. There is also a sucker ganglion, a peripheral nervous element, situated in the stalk of every sucker. The structure and function of the sucker ganglion remains obscure. We examined the cellular organization and molecular composition of the sucker ganglion in Octopus bimaculoides. The sucker ganglion has an ellipsoid shape and features an unusual organization: the neuropil of the ganglion is distributed as a cap aborally (away from the sucker) and a small pocket orally (towards the sucker), with neuronal cell bodies concentrated in the space between. Using in situ hybridization, we detected positive expression of sensory (PIEZO) and motor (LHX3 and MNX) neuron markers in the sucker ganglion cell bodies. Nerve fibers spread out from the sucker ganglion, targeting the surrounding sucker musculature and the oral roots extending to the ANC. Our results indicate that the sucker ganglion is composed of both sensory and motor elements and suggest that this ganglion is not a simple relay for the ANC but facilitates local reflexes for each sucker.

Persistent Disruptions in Prefrontal Connectivity Despite Behavioral Rescue by Environmental Enrichment in a Mouse Model of Rett Syndrome
Rett Syndrome, a neurodevelopmental disorder caused by loss-of-function mutations in the MECP2 gene, is characterized by severe motor, cognitive and emotional impairments. Some of the deficits may result from changes in cortical connections, especially downstream projections of the prefrontal cortex, which may also be targets of restoration following rearing conditions such as environmental enrichment that alleviate specific symptoms. Here, using a heterozygous Mecp2+/- female mouse model closely analogous to human Rett Syndrome, we investigated the impact of early environmental enrichment on behavioral deficits and prefrontal cortex connectivity. Behavioral analyses revealed that enriched housing rescued fine motor deficits and reduced anxiety, with enrichment-housed Mecp2+/- mice performing comparably to wild-type (WT) controls in rotarod and open field assays. Anatomical mapping of top-down anterior cingulate cortex (ACA) projections demonstrated altered prefrontal cortex connectivity in Mecp2+/- mice, with increased axonal density in the somatosensory cortex and decreased density in the motor cortex compared to WT controls. ACA axons revealed shifts in hemispheric distribution, particularly in the medial network regions, with Mecp2+/- mice exhibiting reduced ipsilateral dominance. These changes were unaffected by enriched housing, suggesting that structural abnormalities in prefrontal cortex connectivity persist despite behavioral improvements. Enriched housing rescued brain-derived neurotrophic factor (BDNF) levels in the hippocampus but failed to restore BDNF levels in the prefrontal cortex, consistent with the persistent deficits observed in prefrontal axonal projections. These findings highlight the focal nature of changes induced by reduction of MeCP2 and by exposure to environmental enrichment, and suggest that environmental enrichment starting in adolescence can alleviate behavioral deficits without reversing abnormalities in large-scale cortical connectivity.

Decoding semantics from natural speech using human intracranial EEG
Brain-computer interfaces (BCIs) hold promise for restoring natural language production capabilities in patients with speech impairments, potentially enabling smooth conversation that conveys meaningful information via synthesized words. While considerable progress has been made in decoding phonetic features of speech, our ability to extract lexical semantic information (i.e. the meaning of individual words) from neural activity remains largely unexplored. Moreover, most existing BCI research has relied on controlled experimental paradigms rather than natural conversation, limiting our understanding of semantic decoding in ecological contexts. Here, we investigated the feasibility of decoding lexical semantic information from stereo-electroencephalography (sEEG) recordings in 14 participants during spontaneous conversation. Using multivariate pattern analysis, we were able to decode word level semantic features during language production with an average accuracy of 21% across all participants compared to a chance level of 10%. This semantic decoding remained robust across different semantic representations while maintaining specificity to semantic features. Further, we identified a distributed left-lateralized network spanning precentral gyrus, pars triangularis, and middle temporal cortex, with low-frequency oscillations showing stronger contributions. Together, our results establish the feasibility of extracting word meanings from neural activity during natural speech production and demonstrate the potential for decoding semantic content from unconstrained speech.