Biology subjects

Marshall, W.

Publications and source records attributed to Marshall, W..

12 recordsLinked to original sources

bioRxiv2025

Multi-immersion Oblique Plane Microscope (miOPM): A reconfigurable platform for high-resolution Light-Sheet Fluorescence Microscopy.

Light-Sheet Fluorescence Microscopy (LSFM) enables gentle, rapid, and efficient volumetric imaging of biological specimens. Despite its potential, LSFM has fragmented into multiple variants, each narrowly optimized for specific samples or imaging regimes, limiting its broad applicability. To overcome this barrier, we introduce the multi-immersion oblique plane microscope (miOPM), an adaptable LSFM platform for high-resolution imaging ranging from subcellular dynamics to whole organisms and cleared tissues. miOPM uniquely supports seamless interchangeability of oil, water, and air objectives, and maintains diffraction limited performance across a refractive index range of 1.33 - 1.51. It is compatible with standard sample mounting, current clearing protocols and high-throughput 3D imaging. We leverage miOPM to image diverse biological specimens at multiple spatial scales. By greatly expanding the adaptability and ease of use of LSFM, miOPM is poised to democratize advanced three-dimensional imaging.

bioengineering↗

bioRxiv2025

Mitochondrial function regulates cell growth kinetics to actively maintain mitochondrial homeostasis

Mitochondria are not produced de novo in newly divided daughter cells, but are inherited from the mother cell during mitosis. While mitochondrial homeostasis is crucial for living cells, the feedback responses that maintain mitochondrial volume across generations of dividing cells remain elusive. Here, using a microfluidic yeast mother machine, we tracked several generations of fission yeast cells and observed that cell size and mitochondrial volume grew exponentially during the cell cycle. We discovered that while mitochondrial homeostasis relied on the sizer mechanism of cell size maintenance, mitochondrial function was a critical determinant of the timing of cell division: cells born with lower than average amounts of mitochondria grew slower and thus added more mitochondria before they divided. Thus, mitochondrial addition during the cell cycle was tailored to the volume of mitochondria at birth, such that all cells ultimately contained the same mitochondrial volume at cell division. Quantitative modelling and experiments with mitochondrial DNA-deficient rho0 cells additionally revealed that mitochondrial function was essential for driving the exponential growth of cells. Taken together, we demonstrate a central role for mitochondrial activity in dictating cellular growth rates and ensuring mitochondrial volume homeostasis.

cell biology↗

bioRxiv2024

Pearling Drives Mitochondrial DNA Nucleoid Distribution

The mitochondria of most eukaryotes carry an indispensable second genome (mtDNA), encoding genes engaged in oxidative phosphorylation1. The regular positioning and segregation of mtDNA-containing nucleoids is essential for mitochondrial function and inheritance, as well as cellular health2-5. However, the underlying mechanism driving nucleoid distribution and disaggregation remains unknown6,7. Our data reveal that mitochondria frequently undergo reversible pearling, a biophysical instability that undulates tubules into regularly spaced beads 8, typically triggered by calcium influx. We discovered that physiological pearling imposes a characteristic length scale, simultaneously mediating nucleoid disaggregation and establishing inter-nucleoid distancing with near-maximally achievable precision. We found that lamellar cristae invaginations of the inner mitochondrial membrane play a dual role, determining pearling frequency and duration, and preserving the resulting nucleoid spacing after organelle recovery to a tubular form. Thus, disrupting cristae ultrastructure resulted in more frequent pearling, but also aberrant nucleoid clustering. Our results demonstrate that the distribution of mitochondrial genomes is governed by the interplay between rapid and reversible pearling and cristae ultrastructure, establishing a mechanism for this long-puzzling yet fundamental feature of eukaryotic life, and offering insights into its potential modulation.

cell biology↗

bioRxiv2024

Behavioral and cortical arousal from sleep, muscimol-induced coma, and anesthesia by direct optogenetic stimulation of cortical neurons

The cerebral cortex is widely considered part of the neural substrate of consciousness. However, while several studies have demonstrated that stimulation of subcortical nuclei can produce EEG activation and restore consciousness, so far no direct causal evidence has been available for the cortex itself. Here we tested in mice whether optogenetic activation of cortical neurons in posterior parietal cortex (PtA) or medial prefrontal cortex (mPFC) is sufficient for arousal from three behavioral states characterized by progressively deeper unresponsiveness: sleep, a coma-like state induced by muscimol injection in the midbrain, and deep sevoflurane-dexmedetomidine anesthesia. We find that cortical stimulation always awakens the mice from both NREM sleep and REM sleep, with PtA requiring weaker/shorter light pulses than mPFC. Moreover, in most cases light pulses produce both cortical activation (decrease in low frequencies) and behavioral arousal (recovery of the righting reflex) from brainstem coma, as well as cortical activation from anesthesia. These findings provide evidence that direct activation of cortical neurons is sufficient for behavioral and/or cortical arousal from sleep, brainstem coma, and anesthesia.

neuroscience↗

bioRxiv2024

From micro to macro units: a mathematical framework for identifying the causal grain of a system from its intrinsic perspective

Integrated information theory (IIT) aims to account for the quality and quantity of consciousness in physical terms. According to IIT, a substrate of consciousness must be a system of units that is a maximum of intrinsic, irreducible cause-effect power, quantified by integrated information ({varphi}s). Moreover, the grain of each unit must be the one-- from micro (finer) to macro (coarser)--that maximizes the systems intrinsic irreducibility (i.e., maximizes{varphi} s). The units that maximize{varphi} s are called the intrinsic units of the system. This work extends the mathematical framework of IIT 4.0 to assess cause-effect power at different grains and thereby determine a systems intrinsic units. Using simple, simulated systems, we show that the cause-effect power of a system of macro units can be higher than the cause-effect power of the corresponding micro units. Two examples highlight specific kinds of macro units, and how each kind can increase cause-effect power. The implications of the framework are discussed in the broader context of IIT, including how it provides a foundation for tests and inferences about consciousness.

systems biology↗

bioRxiv2022

Ultrastructural effects of sleep and wake on the parallel fiber synapses of the cerebellum

Multiple evidence in rodents shows that the strength of excitatory synapses in the cerebral cortex and hippocampus is greater after wake than after sleep. The widespread synaptic weakening afforded by sleep is believed to keep the cost of synaptic activity under control, promote memory consolidation, and prevent synaptic saturation, thus preserving the brains ability to learn day after day. The cerebellum is highly plastic and the Purkinje cells, the sole output neurons of the cerebellar cortex, are endowed with a staggering number of excitatory parallel fiber synapses. However, whether these synapses are affected by sleep and wake is unknown. Here we used serial block face scanning electron microscopy to obtain the full 3D reconstruction of more than 7,000 spines and their parallel fiber synapses in the mouse posterior vermis. We find that most Purkinje cell spines carry a synapse, but some do not. The latter, which we call "naked" spines, are [~]5% of all spines after wake but grow to [~]10% of all spines after sleep. Further analysis shows that the changes in the number of naked synapses with wake and sleep can be accounted for by a change in the number of "branched" synapses, which are housed in two or more spines sharing the same neck. Thus, during sleep branched spines may lose one or more synapses or convert to single spines, while the opposite changes occur after wake. Because branched synapses almost always contact different parallel fibers, these results also suggest that during wake, coincidences of firing over parallel fibers may translate into the formation of synapses converging on the same branched spine, which may be especially effective at driving the soma of Purkinje cells. Sleep, on the other hand, may promote the pruning of branched synapses that were formed due to spurious coincidences.

neuroscience↗

bioRxiv2022

A survey of neurophysiological differentiation across mouse visual brain areas and timescales

Neurophysiological differentiation (ND), a metric that quantifies the number of distinct activity states that the brain or its part visits over a period of time, has been used as a correlate of meaningfulness or subjective perception of visual stimuli. ND has largely been studied in non-invasive human whole-brain recordings where spatial resolution is limited. However, it is likely that perception is supported by discrete populations of spiking neurons rather than the whole brain. Therefore, in this study, we use Neuropixels recordings from the mouse brain to characterize the ND metric within neural populations recorded at single-cell resolution in localized regions. Using the spiking activity of thousands of simultaneously recorded neurons spanning 6 visual cortical areas as well as the visual thalamus, we show that the ND of stimulus-evoked activity of the entire visual cortex is higher for naturalistic stimuli relative to artificial ones. This finding holds in most individual areas throughout the visual hierarchy as well. For animals performing an image change detection task, ND of the entire visual cortex (though not individual areas) is higher for successful detection compared to failed trials, consistent with the assumed perception of the stimulus. Analysis of spiking activity allows us to characterize the ND metric across a wide range of timescales from 10s of milliseconds to a few seconds. This analysis reveals that although ND of activity of single neurons is often maximized at an optimal timescale around 100 ms, the optimum shifts to under 5 ms for ND of neuronal ensembles. Finally, we find that the ND of activations in convolutional neural networks (CNNs) trained on an image classification task shows distinct trends relative to the mouse visual system: ND is often higher for less naturalistic stimuli and varies by orders of magnitude across the hierarchy, compared to modest variation in the mouse brain. Together, these results suggest that ND computed on cellular-level neural recordings can be a useful tool highlighting cell populations that may be involved in subjective perception. SummaryAdvances in our understanding on neural coding has revealed that information about visual stimuli is represented across several brain regions. However, availability of information does not imply that it is necessarily utilized by the brain, much less that it is subjectively perceived. Since percepts originate in neural activity, distinct percepts must be associated with distinct states of neural activity, at least within the brain region that supports the percepts. Thus, one approach developed in this direction is to quantify the number of distinct states that the activity of the brain goes through, called neurophysiological differentiation (ND). ND of the entire brain has been shown to reflect subjective reports of visual stimulus meaningfulness. But what specific subpopulations within the brain could be supporting conscious perception, and what is the correct timescale on which states should be quantified? In this study, we analyze ND of spiking neural activity in the mouse visual cortex recorded using Neuropixels probes, allowing us to characterize the ND metric across a wide range of timescales all the way down from 5 ms to a few seconds. It also allows us to understand the ND of neural activity of different ensembles of neurons, from individual thalamic or cortical ensembles to those spanning across multiple visual areas in the mouse brain.

neuroscience↗

bioRxiv2021

Dynamic configuration of large-scale cortical networks during an inhibitory task accounts for heterogeneity in attention-deficit/hyperactivity disorder traits

The heterogeneity of attention-deficit/hyperactivity disorder (ADHD) traits (inattention vs. hyperactivity/impulsivity) complicates diagnosis and intervention. Identifying how the configuration of large-scale functional brain networks during cognitive processing correlate with this heterogeneity could help us understand the neural mechanisms altered across ADHD presentations. Here, we recorded high-density EEG while 62 non-clinical participants (ages 18-24; 32 male) underwent an inhibitory control task (Go/No-Go). Functional EEG networks were created using sensors as nodes and across-trial phase-lag index values as edges. Using cross-validated LASSO regression, we examined whether graph-theory metrics applied to both static networks (averaged across time-windows: -500-0ms, 0-500ms) and dynamic networks (temporally layered with 2ms intervals), were associated with hyperactive/impulsive and inattentive traits. Network configuration during response execution/inhibition was associated with hyperactive/impulsive (mean R2 across test sets = .20, SE = .02), but not inattentive traits. Post-stimulus results at higher frequencies (Beta, 14-29Hz; Gamma, 30-90Hz) showed the strongest association with hyperactive/impulsive traits, and predominantly reflected less burst-like integration between modules in oscillatory beta networks during execution, and increased integration/small-worldness in oscillatory gamma networks during inhibition. We interpret the beta network results as reflecting weaker integration between specialized pre-frontal and motor systems during motor response preparation, and the gamma results as reflecting a compensatory mechanism used to integrate processing between less functionally specialized networks. This research demonstrates that the neural network mechanisms underlying response execution/inhibition might be associated with hyperactive/impulsive traits, and that dynamic, task-related changes in EEG functional networks may be useful in disentangling ADHD heterogeneity.

neuroscience↗

bioRxiv2020

Measuring stimulus-evoked neurophysiological differentiation in distinct populations of neurons in mouse visual cortex

Despite significant progress in understanding neural coding, it remains unclear how the coordinated activity of large populations of neurons relates to what an observer actually perceives. Since neurophysiological differences must underlie differences among percepts, differentiation analysis--quantifying distinct patterns of neurophysiological activity--is an "inside out" approach that addresses this question. We used two-photon calcium imaging in mice to systematically survey stimulus-evoked neurophysiological differentiation in excitatory populations across 3 cortical layers (L2/3, L4, and L5) in each of 5 visual cortical areas (primary, lateral, anterolateral, posteromedial, and anteromedial) in response to naturalistic and phase-scrambled movie stimuli. We find that unscrambled stimuli evoke greater neurophysiological differentiation than scrambled stimuli specifically in L2/3 of the anterolateral and anteromedial areas, and that this effect is modulated by arousal state and locomotion. Contrariwise, decoding performance was far above chance and did not vary substantially across areas and layers. Differentiation also differed within the unscrambled stimulus set, suggesting that differentiation analysis may be used to probe the ethological relevance of individual stimuli.

neuroscience↗

bioRxiv2019

Multi-scale spatial heterogeneity enhances particle clearance in airway ciliary arrays

Mucus clearance constitutes the primary defence of the respiratory system against viruses, bacteria and environmental insults [1]. This transport across the entire airway emerges from the integrated activity of thousands of multiciliated cells, each containing hundreds of cilia, which together must coordinate their spatial arrangement, alignment and motility [2, 3]. The mechanisms of fluid transport have been studied extensively at the level of an individual cilium [4, 5], collectively moving metachronal waves [6-10], and more generally the hydrodynamics of active matter [11, 12]. However, the connection between local cilia architecture and the topology of the flows they generate remains largely unexplored. Here, we image the mouse airway from the sub-cellular (nm) to the organ scales (mm), characterising quantitatively its ciliary arrangement and the generated flows. Locally we measure heterogeneity in both cilia organisation and flow structure, but across the trachea fluid transport is coherent. To examine this result, a hydrodynamic model was developed for a systematic exploration of different tissue architectures. Surprisingly, we find that disorder enhances particle clearance, whether it originates from fluctuations, heterogeneity in multiciliated cell arrangement or ciliary misalignment. This resembles elements of stochastic resonance [13-15] in a self-assembled biological system. Taken together, our results shed light on how the microstructure of an active carpet [16, 17] determines its emergent dynamics. Furthermore, this work is also directly applicable to human airway pathologies [1], which are the third leading cause of deaths worldwide [18].

biophysics↗

bioRxiv2018

Electron Cryo-Tomography Provides Insight into Procentriole Architecture and Assembly Mechanism

Centriole is an essential structure with multiple functions in cellular processes. Centriole biogenesis and homeostasis is tightly regulated. Using electron cryo-tomography (cryoET) we present the structure of procentrioles from Chlamydomonas reinhardtii. We identified a set of non-tubulin components attached to the triplet microtubule (MT), many are at the junctions of tubules likely to reinforce the triplet. We describe structure of the A-C linker that bridges neighboring triplets. We find that POC1 is an integral component of the A-C linker. Its conserved WD40 {beta}-propeller domain provides sites for attachment to other A-C linker components. The twist of A-C linker results in an iris diaphragm-like motion of the triplets in the longitudinal direction of procentriole. Finally, we identified two assembly intermediates at the growing ends of procentriole allowing us to propose a model for the procentriole assembly. Our results provide a comprehensive structural framework for understanding the molecular mechanisms underpinning procentriole biogenesis and assembly.

cell biology↗

bioRxiv2017

Role of somatostatin-positive cortical interneurons in the generation of sleep slow waves

Cortical slow waves - the hallmark of NREM sleep - reflect near-synchronous OFF periods in cortical neurons. However, the mechanisms triggering such OFF periods are unclear, as there is little evidence for somatic inhibition. We studied cortical inhibitory interneurons that express somatostatin (SOM), because [~]70% of them are Martinotti cells that target diffusely layer 1 and can block excitatory transmission presynaptically, at glutamatergic terminals, and postsynaptically, at apical dendrites, without inhibiting the soma. In freely moving mice, we show that SOM+ cells can fire immediately before slow waves and their optogenetic stimulation triggers neuronal OFF periods during sleep. Next, we show that chemogenetic activation of SOM+ cells increases slow wave activity (SWA), the slope of individual slow waves, and the duration of NREM sleep; whereas their chemogenetic inhibition decreases SWA and slow wave incidence without changing time spent asleep. By contrast, activation of parvalbumin+ (PV+) cells, the most numerous population of cortical inhibitory neurons, greatly decreases SWA and cortical firing. These results indicate that SOM+ cells, but not PV+ cells, are involved in the generation of sleep slow waves. Whether Martinotti cells are solely responsible for this effect, or are complemented by other classes of inhibitory neurons, remains to be investigated.

neuroscience↗