bioRxiv Subject Collection: Neuroscience's Journal

Metabolic reprogramming and altered ATP content impair neuroprotective functions of microglia in β-glucocerebrosidase deficiency models
Mutations in the GBA gene, which reduce beta-glucocerebrosidase (GCase) activity, represent the most significant genetic risk factor for Parkinson's disease (PD). Decreased GCase activity has also been observed in sporadic PD cases, supporting a broader role for GCase in the poorly understood mechanisms underlying PD etiopathogenesis. While most studies on the relationship between GBA mutations and PD have focused on neurons, evidence suggests that PD pathology promoted by GCase deficiency involves other cell types and, in particular, interactions between neuronal and glial cells. Here, we identify microglia as primary players undergoing significant alterations at early stages of the pathological processes triggered by a GCase impairment. Using both pharmacological and genetic mouse models of GCase deficiency, we observed microglial morphological, transcriptional and metabolic changes. Interestingly, these changes were associated with a cell-specific, significant reduction of microglial ATP levels. When microglial ATP depletion was reproduced in an in vitro system of co-cultured microglial and neuronal cells, the neuroprotective properties of microglia were compromised and neuronal susceptibility to oxidative stress was enhanced. These findings underscore the role of microglia in PD pathogenesis and point to a pathogenetic mechanism by which microglial metabolic disturbances leading to ATP depletion enhance neuronal vulnerability to injury and neurodegeneration. This mechanism could be targeted for therapeutic intervention aimed at mitigating PD risk and counteracting the development of PD pathology.

Machine Learning Discovers Numerous New Computational Principles Supporting Elementary Motion Detection
Motion direction detection is a fundamental visual computation that transforms spatial luminance patterns into directionally tuned outputs. Classical models of direction selectivity rely on temporal asymmetry, where motion detection arises through either delayed excitation or inhibition. Here, I used biologically inspired machine learning applied to retinal and cortical circuits to uncover multiple novel feedforward architectures capable of direction selectivity. These include mechanisms based on asymmetric synaptic properties, spatial receptive field variations, new roles for pre- and postsynaptic inhibition, and previously unrecognized kinetic implementations. Conceptually, these circuit architectures cluster into eight computational primitives underlying motion detection, four of which are newly discovered. Many of the solutions rival or outperform classical models in both robustness and precision, and several exhibit enhanced noise tolerance. All mechanisms are biologically plausible and correspond to known physiological and anatomical motifs, offering fresh insights into motion processing and illustrating how machine learning can uncover general principles of neural computation.

Olfactory bulb and cortex activity reflects subjective odor intensity perception rather than concentration
Understanding stimulus intensity processing is fundamental in sensory science, yet this question remains largely unexplored in human olfaction. We investigated how the human olfactory bulb (OB) and piriform cortex (PC) process odor concentration versus subjective perceived intensity. We demonstrate that OB-PC network oscillatory dynamics are predominantly driven by perceived intensity, not physical concentration. The OB initially processes and communicates perceived intensity to the PC via early gamma-band oscillations (bottom-up feedback). The PC then refines and sends this percept back to the OB via later beta-band oscillations (top-down feedback), updating the OB's gamma activity for subsequent odorants. Critically, analyses of phase-amplitude coupling and beta burst activity demonstrate that transient beta patterns from the PC update OB gamma activity, providing the OB with an updated internal representation of the odor percept. These results reveal an oscillatory mechanism by which the olfactory system maintains perceptual constancy and adaptability despite fluctuations in environmental odor concentrations.

Counteracting memories acquired in parallel by limited pairings in eyeblink conditioning
Animals adapt to environments by learning even when experience is limited, which is critical for their survival in nature. However, the relationship between experience richness and learning efficiency remains elusive. Here we investigated this issue using mice eyeblink conditioning with three distinct training paradigms. Surprisingly, 1/9th of sensory pairings than conventional ones were not only sufficient for acquisition of conditioned responses, relying on the activity of deep cerebellar nuclei as usual, but also resulted in more efficient learning. On the other hand, the learning established by limited training was more susceptible for extinction. Modeling of establishment and extinction processes, coupled with experimental validation of the model prediction, unveiled that memories for execution of eyeblink and its suppression are independently formed irrespective of their temporal order.

Microglial SWELL1 Deficiency Drives Male-Specific Seizure Vulnerability but Paradoxical Neuroprotection through Impaired Phagocytosis
The discovery of genes encoding the volume-regulated anion channel (VRAC) has enabled detailed exploration of its cell type-specific roles in the brain. LRRC8A (SWELL1) is the essential VRAC subunit. We observed seizure-induced, subunit-specific changes in microglial VRAC expression and investigated its function using conditional knockout (cKO) of LRRC8A in microglia. SWELL1 cKO mice exhibited a male-specific increase in kainate-induced seizure severity yet showed paradoxical neuroprotection against seizure-associated neuronal loss. Mechanistically, SWELL1 deletion led to a cell-autonomous reduction in microglial density and decreased release of VRAC-permeable neuroactive metabolites, including taurine, GABA, and glutamate. Additionally, impaired phagocytic kinetics and reduced lysosomal biogenesis contributed to the observed neuroprotection. These findings reveal novel roles for microglial VRAC in regulating seizure outcomes and microglia-neuron interactions.

Prolonged Single Neuron Voltage Imaging in Behaving Mammals
Long-term recording of single-neuron membrane voltage dynamics during behavior is highly desirable. Using ElectraOFF, a fully genetically encoded, photostable, high-performance fluorescence voltage indicator, we achieve routine cellular-resolution imaging over tens of minutes, occasionally up to eighty minutes, in behaving mice with minimal signal loss across neuron types and brain regions. This extended recording capability reveals plasticity changes throughout intracranial electrical neuromodulation, highlighting the novel insights enabled by prolonged voltage imaging.

Neural integration of acoustic statistics enables detecting acoustic targets in noise
Sound detection amidst noise presents an important challenge in audition. Many naturally occurring sounds (rain, wind) can be described and predicted only statistically, so-called sound textures. Previous research has demonstrated the human ability to leverage this statistical predictability for sound recognition, but the neural mechanisms remain elusive. We trained mice to detect vocalizations embedded in sound textures with different statistical predictability, while recording and optogenetically modulating the neural activity in the auditory cortex. Mice showed improved performance and neural encoding if they could sample the statistics longer per trial. Textures with more exploitable structure, specifically higher cross-frequency correlations improved performance, background encoding and vocalization activity. Activating parvalbumin-positive (PV) interneurons had an asymmetric effect, improving detection and neural encoding of vocalizations for low correlations, and impoverishing them for high cross-frequency correlations. In summary, mice exploit stimulus statistics to improve sound detection in naturalistic background noise, reflected in behavioral performance and neural activity, relying on PV interneurons for temporal integration.

Cholinergic disruption of state-dependent retrosplenial layer 1 activity causes temporal associative memory deficit under stress
Memory functions rely on discrete patterns of neuronal activity emerging from individual neurons, their interconnected circuits, and their alignment with global brain states. We show that such patterns are generated by layer 1 inhibitory neurons in the ventral retrosplenial cortex (vRSPL1), whose activity correlated with immobility and specifically contributed to the formation of temporal associative memories. This state-dependent activity was subject to stress-induced cholinergic modulation through muscarinic 1 receptor (M1R) signaling, leading to selective impairments in temporal but not contextual associative memory. Slice studies showed that these effects were likely due to M1R-mediated inhibition of vRSPL1 neurons, transiently disrupting their local connectivity as well as responsiveness to afferent input. Together, we demonstrate a mechanism by which vRSPL1 activity aligned to immobility coordinate the formation of temporal associations. Through its sensitivity to stress-related cholinergic modulation, this mechanism presents vulnerability to traumatic amnesia, especially for temporal details of episodic memories.

Increased Complement C4 in a Sparse Neuronal Subset Induces Network-Wide Transcriptomic Alterations in the Prefrontal Cortex
The complement (C) pathway, a vital part of innate immunity, defends against pathogens and supports tissue surveillance. While local activation in the periphery enhances immune protection, dysregulation can trigger a self-amplifying cascade that spreads beyond the initial site, resulting in tissue injury. In the brain, complement proteins regulate synaptic plasticity and connectivity, raising the possibility that similar mechanisms of maladaptive propagation may disrupt neural circuits under pathological conditions. Although complement dysregulation is linked to neurological and psychiatric disorders, the impact of localized upregulation in a small subset of cells on broader cortical networks remains unclear. To examine this, we overexpressed the schizophrenia (SCZ) risk gene C4 (C4-OE) in approximately 2% of prefrontal cortex neurons in mice using in utero electroporation. Bulk RNA sequencing of microdissected tissue revealed widespread transcriptional changes in a network predominantly comprising untransfected cells. C4-OE induced the upregulation of genes involved in cholesterol biosynthesis, axon guidance, synaptic plasticity, cytoprotection, and neurogenesis, suggesting a response consistent with compensatory remodeling or aberrant plasticity. Co-expression analysis identified a C4b-containing module enriched for dendritic development and cell cycle regulation, indicating alterations in circuit maturation. In contrast, immune and inflammatory genes were broadly downregulated, suggesting homeostatic suppression in response to sustained complement activity. Comparisons with human SCZ proteomic datasets revealed conserved gene signatures, highlighting the potential of our model to capture disease-relevant mechanisms. These findings demonstrate that sparse C4 overexpression can trigger widespread transcriptional changes across neural circuits, suggesting that local complement dysregulation may spread through cortical networks and drive broader functional disruption.

Neural connectivity of a computational map for fly flight control.
Nervous systems rely on sensory feature maps, where the tuning of neighboring neurons for some ethologically-relevant parameter varies systematically, to control behavior1,2. Such maps can be organized topographically or based on some computational principle. However, it is unclear how the central organization of a sensory system corresponds to the functional logic of the motor system. This problem is exemplified by insect flight, where sub-millisecond modifications in wing-steering muscle activity are necessary for stability and maneuverability. Although the muscles that control wing motion are anatomically and functionally stratified into distinct motor modules3-7, comparatively little is known about the architecture of the sensory circuits that regulate their precise firing times. Here, we leverage an existing volume of an adult female VNC of the fruit fly Drosophila melanogaster8,9 to reconstruct the complete population of afferents in the haltere--nature's only biological ''gyroscope''10,11--and their synaptic partners. We morphometrically classify these neurons into distinct subtypes and design split-GAL4 lines that help us determine the peripheral locations from which each subtype originates. We find that each subtype, rather than originating from the same anatomical location, is comprised of multiple regions on the haltere. We then trace the flow of rapid mechanosensory feedback from the peripheral haltere receptors to the central motor circuits that control wing kinematics. Our work demonstrates how a sensory system's connectivity patterns construct a neural map that may facilitate rapid processing by the motor system.

Multifaceted brain representation of numerosity across the senses and presentation formats
Humans can extract numerosity from different senses and a variety of context. How and where the brain abstract numerical information from low-level sensory inputs remains debated. Using multivariate pattern analysis (MVPA) and representational similarity analysis (RSA) applied to fMRI data, we comprehensively investigate how the brain represents numerical information (range 2-5) across different modalities (auditory, visual) and formats (sequential, simultaneous; symbolic, non-symbolic). We identify a set of brain regions along the dorsal pathway - from early visual cortex to the intraparietal and frontal regions - that encode specific non-symbolic numerical information across formats and modalities. The numerical distance effect, a hallmark of magnitude encoding, was observed in most of these regions. We found aligned representation of numerical information across visual and auditory modalities in intraparietal and frontal regions, but only when they shared a sequential presentation format. RSA further revealed a posterior-to-anterior gradient in the intraparietal sulcus (IPS) showing that the dominant factors influencing distributed numerical representations shifted from sensory modality in the posterior parietal regions to presentation format in the anterior parietal areas. Our study reveals multifaceted organization of numerical processing from various sensory and format inputs.

A Multi-Region Brain Model to Elucidate the Role of Hippocampus in Spatially Embedded Decision-Making
Brains excel at robust decision-making and data-efficient learning. Understanding the architectures and dynamics underlying these capabilities can inform inductive biases for deep learning. We present a multi-region brain model that explores the normative role of structured memory circuits in a spatially embedded binary decision-making task from neuroscience. We counterfactually compare the learning performance and neural representations of reinforcement learning (RL) agents with brain models of different interaction architectures between grid and place cells in the entorhinal cortex and hippocampus, coupled with an action-selection cortical recurrent neural network. We demonstrate that a specific architecture--where grid cells receive and jointly encode self-movement velocity signals and decision evidence increments--optimizes learning efficiency while best reproducing experimental observations relative to alternative architectures. Our findings thus suggest brain-inspired structured architectures for efficient RL. Importantly, the models make novel, testable predictions about organization and information flow within the entorhinal-hippocampal-neocortical circuit: we predict that grid cells must conjunctively encode position and evidence for effective spatial decision-making, directly motivating new neurophysiological experiments.

Heightened Sensitivity to Voice Loudness Changes in Parkinson's Disease
Parkinson's disease (PD) affects voice and speech production, often resulting in reduced vocal intensity and monotonous speech. Recent studies have suggested that these changes can be partially explained by altered sensory feedback processing when producing speech. Individuals with PD (IwPD) may fail to monitor sensory feedback from their own voice, impairing their ability to adjust voice and speech when sensory input differs from expectations. In this study, we investigated sensory feedback processing in PD by looking at the sensory attenuation typically observed in event-related responses (ERP) to the self-generated voice. When sensory feedback processing is intact, the P50, N100, and P200 ERP responses to the self-generated voice are more attenuated than to an externally-generated voice. Twenty-three IwPD and 23 healthy controls (HCs) participated in a voice playback study that comprised three conditions: self-generated voice (auditory-motor condition; AMC), externally-generated voice (auditory-only condition; AOC), and motor-only. The AMC and AOC conditions also included an amplitude modulation of the voice (0/+15dB). Linear mixed models assessed group differences in ERP morphology. While groups did not differ in their P50 and P200 responses, there was a significant group-condition-loudness interaction for the N100. Follow-up analyses showed that IwPD displayed much larger N100 error responses for unexpected loudness modulations as compared to HC. This observation suggests that IwPD may process voice modulations differently than HC. The hypersensitivity to loudness changes may underlie IwPD's difficulties in processing and adapting their voice acoustics.

Cognitive and Synaptic Impairment Induced by Deficiency of Autism Risk Gene Smarcc2 and its Rescue by Histone Deacetylase Inhibition
SMARCC2, which encodes BAF170, a core subunit of chromatin remodeling BAF complex, is one of the top-ranking risk genes for autism spectrum disorder (ASD). However, the mechanisms linking SMARCC2 haploinsufficiency to ASD remain poorly understood. SMARCC2 binds to many other ASD risk genes involved in transcriptional regulation. Knockdown (KD) of Smarcc2 in prefrontal cortex (PFC) of adolescent mice led to impaired working memory, with largely intact social and anxiety-like behaviors. Genome-wide RNA-seq analysis revealed that the downregulated genes by Smarcc2 KD were enriched in synaptic transmission. Significant reduction of GABA and glutamate-related genes was also found in PFC of Smarcc2-deficient mice by qPCR profiling. In parallel, electrophysiological recordings uncovered the significant impairment of GABAergic and glutamatergic synaptic currents in PFC pyramidal neurons by Smarcc2 KD. Smarcc2 binds to HDAC2, and Smarcc2 KD reduced global histone acetylation and H3K9ac enrichment at synaptic gene Slc1a3 (EAAT1), Slc6a1 (GAT1), and Slc32a1 (VGAT) promoters. Treatment of Smarcc2-deficient mice with romidepsin, a class I histone deacetylase (HDAC) inhibitor, restored H3K9ac level, working memory and synaptic gene expression. These findings highlight the critical role of Smarcc2 in regulating cognitive and synaptic function, suggesting that targeting HDAC could alleviate deficits in Smarcc2-associated neurodevelopmental disorders.

An increased excitation and inhibition onto CA1 pyramidal cells sets the path to Alzheimer s disease
Synapses are critical targets of Alzheimer s disease (AD), a highly prevalent neurodegenerative disease associated with accumulation of extracellular amyloid-{beta} peptides. Although amyloidosis and aggregation of the 42-amino acid amyloid-{beta} (A{beta}42) have long been considered pathogenic triggers for AD, clinical evidence linking high levels of A{beta}42 with normal cognition challenges this hypothesis. To resolve this conundrum on the role of A{beta}42 in regulating synaptic activity, we used an adeno-associated viral vector approach that triggers extracellular accumulation of A{beta}42 and spatial memory impairment. We show that A{beta}42 leads to an early increase in excitatory and proximal inhibitory synaptic transmission onto hippocampal CA1 pyramidal cells, and an increased expression of the glutamate transporter GLT-1 in these cells. A{beta}42 accumulation does not cause early cognitive deficits unless accompanied by an increased neuronal GLT-1 expression, suggesting this transporter is a critical mediator of A{beta}42 s effects. These findings unveil key molecular and cellular mechanisms implicated with AD pathogenesis.

Gene dosage effects of 22q11.2 copy number variants on in-vivo measures of white matter axonal density and dispersion
22q11.2 deletion (22qDel) and duplication (22qDup) carriers have an increased risk of neurodevelopmental disorders and exhibit altered brain structure, including white matter microstructure. However, the underlying cellular architecture and age-related changes contributing to these white matter alterations remain poorly understood. Neurite orientation dispersion and density imaging (NODDI) was used on mixed cross-sectional and longitudinal data to examine group differences and age-related trajectories in measures of axonal density (i.e., intracellular volume fraction; ICVF), axonal orientation (orientation dispersion index; ODI) and free water diffusion (isotropic volume fraction; ISO) in 50 22qDel (n scans = 69, mean age = 21.7, age range = 7.4-51.1, 65.2% female) and 24 22qDup (n scans = 34, mean age = 23.3, age range = 8.3-49.4, 55.0% female) carriers, and 890 controls (n scans = 901, mean age = 21.9, age range = 7.8-51.1, 54.5%). The results showed widespread gene dosage effects, with higher ICVF in 22qDel and lower ICVF in 22qDup compared to controls, and region-specific effects of the 22qDel and 22qDup on ODI and ISO measures. However, 22qDel and 22qDup carriers did not exhibit an altered age-related trajectory relative to controls. Observed differences in ICVF suggest higher white matter axonal density in 22qDel and lower axonal density in 22qDup compared to controls. Conversely, differences in ODI are highly localized, indicating region-specific effects on axonal dispersion in white matter. We do not find evidence for altered developmental trajectories of axonal density or dispersion among 22q11.2 CNV carriers, suggesting stable disruptions to neurodevelopmental events before childhood.

Beyond the first glance: How human presence enhances visual entropy and promotes spatial learning
Spatial learning emerges not only from static environmental cues but also from the social and semantic context embedded in our surroundings. This study investigates how human agents influence visual exploration and spatial knowledge acquisition in a controlled Virtual Reality (VR) environment, focusing on the role of contextual congruency. Participants freely explored a 1 km{superscript 2} virtual city while their eye movements were recorded. Agents were visually identical across conditions but placed in locations that were either congruent, incongruent, or neutral with respect to the surrounding environment. Using Bayesian hierarchical modeling, we found that incongruent agents elicited longer fixations and significantly higher gaze transition entropy (GTE), a measure of scanning variability. Crucially, GTE emerged as the strongest predictor of spatial recall accuracy. These findings suggest that human-contextual incongruence promotes more flexible and distributed visual exploration, thereby enhancing spatial learning. By showing that human agents shape not only where we look but how we explore and encode space, this study contributes to a growing understanding of how social meaning guides attention and supports navigation.

Disentangling objects' contextual associations from perceptual and conceptual attributes using time-resolved neural decoding
Humans effortlessly relate what they see to what they know, drawing on existing knowledge of objects' perceptual, conceptual, and contextual attributes while searching for and recognising objects. While prior studies have investigated the temporal dynamics of perceptual and conceptual object properties in the neural signal, it remains unclear whether and when contextual associations are uniquely represented. In this study, we used representational similarity analysis on electroencephalography (EEG) data to explore how the brain processes the perceptual, conceptual, and contextual dimensions of object knowledge over time. Using human similarity judgments of 190 naturalistic object concepts presented as either as images or words, we constructed separate behavioural models of objects' perceptual, conceptual, and contextual properties. We correlated these models with neural patterns from two EEG datasets, one publicly available and one newly collected, both recorded while participants passively viewed the same object stimuli. Across both datasets, we found that perceptual features dominated the early EEG response to object images, while conceptual features emerged later. Contextual associations were also reflected in neural patterns, but their explanatory power largely overlapped with that of conceptual models, suggesting limited unique representation of objects' contextual attributes under passive viewing conditions. These results highlight the brain's integration of perceptual and conceptual information when processing visual objects. By combining high temporal resolution EEG with behaviourally derived models, this study advances our understanding of how distinct dimensions of object knowledge are encoded in the human brain.

In vivo calcium imaging shows that dorsal root ganglion stimulation predominantly activates large-sized sensory neurons in mice
Dorsal root ganglion (DRG) stimulation is an effective treatment for patients with refractory neuropathic pain. However, the exact mechanisms through which DRG stimulation exerts its analgesic effects, especially in relation to the recruitment of different fiber types present within the DRG, remains largely unknown. Here, we examined the effect of DRG stimulation (20Hz) on sensory neuron fiber recruitment and neuronal responses induced by mechanical hind paw stimulation using in vivo calcium imaging in mice. GCaMP6s was delivered to sensory neurons via an adeno-associated viral vector of serotype 9 (AAV9) injected subcutaneously at P2-5. In adult mice, a stimulation electrode was placed over the L4 DRG, and in vivo calcium imaging was performed. The proportion of responsive neurons and response magnitude was assessed during 30 seconds of DRG stimulation at different stimulation amplitudes (33%, 50%, 66%, 80%, 100% of motor threshold (MT)) for small, medium and large-sized subpopulations. In a second experiment, the effect of 30 minutes of DRG stimulation was assessed at 66% MT on neuronal responses induced by mechanical hind paw stimulation during and following DRG stimulation. We show that 20Hz DRG stimulation at clinically relevant amplitudes mainly activates large-size sensory neurons in mice, both in terms of the proportion of neurons responding as well as response magnitude. There was a small decrease in the proportion of neurons responding to mechanical stimulation during DRG stimulation, which did not recover 30 minutes after stimulation had been switched off. By contrast, DRG stimulation did not alter the response magnitude of responding neurons.

L-selectin shedding regulates functional recovery and neutrophil clearance following spinal cord injury in a sex-dependent manner
During the acute phase of spinal cord injury (SCI), neutrophils infiltrate in large numbers and can exacerbate inflammation, secondary tissue damage, and neurological deficits. L-selectin is a signaling and adhesion receptor that has been shown to facilitate neutrophil recruitment and secondary injury after SCI. During neutrophil activation, L-selectin is typically cleaved or shed from the cell surface and augmenting L-selectin shedding can improve hindlimb recovery and tissue sparing following SCI in male mice. However, it is unclear how endogenous L-selectin shedding regulates neutrophil responses and functional recovery after SCI, particularly when also considering sex as a biological variable. In this study, we investigated the sex-dependent role of endogenous L-selectin shedding in neutrophil function and long-term outcomes in a murine thoracic contusion model of SCI. We found that endogenous L-selectin shedding improves long-term functional recovery and white matter sparing in female, but not male, mice. In addition, we demonstrate that L-selectin shedding alters neutrophil accumulation in a sex-dependent manner. While L-selectin shedding does not mediate neutrophil activation or effector functions, we found that neutrophil clearance is facilitated by L-selectin shedding in female mice alone. These results demonstrate that endogenous L-selectin shedding is a critical and sex-dependent mediator of neutrophil accumulation and clearance, as well as long-term functional outcomes, after SCI.

Converging pathways: shared brain circuitry engaged by monoaminergic antidepressants, ketamine and psilocybin
Ketamine has transformed depression treatment by providing therapeutic relief within a single day, unlike monoaminergic antidepressants, which require several weeks to take effect. Here, we conducted whole-brain screening in mice to compare drug-evoked c-fos expression, acting as a marker of brain activity leading to protein synthesis-dependent forms of plasticity, following treatment with monoaminergic antidepressants, ketamine and psilocybin. Our findings reveal a shared limbic brain circuit comprising subcortical and frontocortical regions, with a key distinction: c-fos-based activity in the prelimbic and infralimbic frontal cortex, areas strongly implicated in depression, was acutely induced by ketamine and high-dose psilocybin, but emerged only after chronic dosing with the selective serotonin reuptake inhibitor fluoxetine or microdose psilocybin. These results suggest the existence of a core limbic subcortico-cortical circuit underlying antidepressant efficacy, provide mechanistic insight into the delayed therapeutic effects of monoaminergic antidepressants, and reveal a close similarity in brain activity evoked by monoaminergic antidepressants and psilocybin microdosing.

Lipidomic signatures in microglial extracellular vesicles during acute inflammation: a gateway to neurological biomarkers
Extracellular vesicles (EVs) are membrane bound vesicles released from all cells throughout the body, including the central nervous system, and are known to carry both membrane-bound proteins and cargo reflective of their cell of origin. EVs show promise as neurological disease biomarkers due to their molecular makeup reflecting their parent-cell composition signature and due to their ability to cross the blood-brain barrier. To-date, the vast majority of research in this field has explored the protein profiles of EVs; however, lipids play an important role not only in the formation of EVs, but also in mediating cellular function and the pathological progression of many neurodegenerative conditions. Herein, we take a critical first step in determining the potential utility of EV lipids as biomarkers in neurological disease. In vitro we exposed BV-2 microglia to either control media or media containing lipopolysaccharides (LPS), a known pro-inflammatory stimulus, for 24 hours then isolated both the cells and their EVs and performed LC-MS/MS. For the first time, we reveal distinct lipidomic changes can differentiate resting vs. pro-inflammatory microglia and their EVs, while distinct lipids are preserved between EVs and their parent cell. Moreover, we add to current literature by demonstrating acute pro-inflammatory activation of microglia results in the activation and suppression of distinct lipidomic pathways. Finally, we demonstrate that analysis of lipid-based relationships between parent cells and their EVs may be a useful tool to infer cellular function. This study is the first of its kind to demonstrate that lipidomic analysis can not only differentiate the functional state of cells in vitro but can also differentiate their EVs. We lay the first brick in a foundation to support future research into EV lipids as novel and exciting biomarker candidates in neurological disease.

A PDZ-RapGEF promotes synaptic development in C. elegans through a Rap/Rac signaling pathway
Small G proteins coordinate the development of nerve terminals. The activity of G proteins is finely tuned by GTPase regulatory proteins. Previously, we observed that PXF-1, a Caenorhabditis elegans GTPase regulatory protein, is required for the function of cholinergic motor neurons. Here, we investigated how PXF-1 coordinates the development of presynaptic terminals at the molecular level. We observed that PXF-1 acts through RAP-1 to promote synapse development. Subsequently, we found that pxf-1 mutants display a reduction in RAC-2 activity, which is required for cholinergic synapse development. We observed that RAC-2 acts downstream of RAP-1. Finally, we identified a physical interaction between RAP-1 and TIAM-1, a Rac guanine exchange factor, which links PXF-1 function to the presynaptic actin cytoskeleton through RAC-2 activation. These findings highlight how small G protein signaling pathways interact to coordinate the development of presynaptic terminals.

ECgo: All-Optical Induction of Single Endothelial Cell Injury and Capillary Occlusion in the Brain
The ability to induce endothelial cell (EC) damage in the mouse brain with high spatial precision is invaluable for mechanistic studies of brain capillary injury and repair. Here, we introduce an optical method, termed ECgo, that utilizes a new two-photon-excitable porphyrin-based photosensitizer (Ps2P) to selectively obliterate single ECs within the brain microvascular network. Using the developed approach, we were able to induce occlusions of single capillaries with high spatiotemporal control, while preserving the surrounding tissue. Combined with longitudinal two-photon imaging, ECgo enables studies of morphological and functional consequences of targeted single capillary EC injury in vivo under healthy and diseased conditions.