Biology subjects

Wilson, S. P.

Publications and source records attributed to Wilson, S. P..

9 recordsLinked to original sources

bioRxiv2024

Single cell RNA sequencing reveals a shift in cell function and maturation of endogenous and infiltrating cell types in response to acute intervertebral disc injury

Intervertebral disc (IVD) degeneration contributes to disabling back pain. Degeneration can be initiated by injury, and progressively leads to irreversible cell loss and loss of IVD function. Attempts to restore IVD function through cell replacement therapies have had limited success due to knowledge gaps in the critical cell populations and molecular crosstalk after injury. Here, we used single cell RNA sequencing to identify the transcriptional changes of endogenous cells of the IVD and infiltrating cell populations following IVD injury. Control and Injured coccygeal IVDs were extracted from 12 week old female C57BL/6J mice 7 days post injury and subjected to single-cell resolution transcriptomic sequencing. Clustering, gene ontology, and pseudotime trajectory analyses determined transcriptomic divergences in the cells of the Injured IVD, flow cytometry identified they types of infiltrating immune cells, and immunofluorescence was utilized to define mesenchymal stem cell (MSC) localization. Clustering analysis revealed 11 distinct cell populations that included IVD, immune, vascular cells, and MSCs. Differential gene expression analysis determined that Outer Annulus Fibrosus, Neutrophils, Saa2-High MSCs, Macrophages, and Krt18+ Nucleus Pulposus (NP) cells were the major drivers of transcriptomic differences between Control and Injured cells. Gene ontology revealed that the most upregulated biological pathways were angiogenesis and T cell-related while wound healing and ECM regulation categories were downregulated. Pseudotime trajectory analyses revealed that IVD injury directed cells towards increased differentiation in all clusters, except for Krt18+ NP cells which remained in a less mature cell state. Saa2-High and Grem1-High MSCs populations drifted towards more differentiated IVD cells profiles with injury and localized distinctly within the IVD. This study revealed novel MSC populations in a heterogeneous landscape of IVD cell populations during injury, and these cells may be leveraged for future IVD repair studies. Lay SummaryThe intervertebral disc (IVD) is a spinal joint that accumulates damage with age but has limited tissue repair capabilities. IVD injuries progress into degeneration, and IVD degeneration is a leading cause of lower back pain. Understanding the cell populations that change and respond to injury will uncover targets to restore IVD function. Mesenchymal stem cells (MSCs) are cells within the IVD that can potentially replenish the cells lost after IVD damage. To better understand how IVD cell populations are affected by tissue damage, we performed single cell RNA sequencing of IVD tissue 7 days post injury and analyzed the differences in gene regulation. We identified diverse cells populations such as IVD, immune cells, vascular cells, and MSCs. We discovered the presence of Saa2 and Grem1 expressing MSCs that become less stem cell-like and express higher levels of IVD gene markers after injury. We also determined that Saa2 and Grem1 have varying gene expression patterns in IVD tissues that becomes attenuated after injury. These MSCs could be targeted for future stem cell therapies to prevent IVD degeneration.

cell biology↗

bioRxiv2023

Prions in carnivore feces: recovery from spiking experiments and detection in free-ranging carnivores

Chronic wasting disease (CWD) is a highly contagious, fatal neurodegenerative disease caused by infectious prions (PrPCWD) affecting wild and captive cervids. Although experimental feeding studies have demonstrated prions in feces of crows (Corvus brachyrhynchos), coyotes (Canis latrans), and cougars (Puma concolor), the role of scavengers and predators in CWD epidemiology remains poorly understood. Here we applied the real-time quaking-induced conversion (RT-QuIC) assay to detect PrPCWD in feces from cervid consumers, to advance surveillance approaches, which could be used to improve disease research and adaptive management of CWD. We assessed recovery and detection of PrPCWD by experimental spiking of PrPCWD into carnivore feces from 9 species sourced from CWD-free populations or captive facilities. We then applied this technique to detect PrPCWD from feces of predators and scavengers in free-ranging populations. Our results demonstrate that spiked PrPCWD is detectable from feces of free-ranging mammalian and avian carnivores using RT-QuIC. Results show that PrPCWD acquired in natural settings is detectable in feces from free-ranging carnivores, and that PrPCWD rates of detection in carnivore feces reflect relative prevalence estimates observed in the corresponding cervid populations. This study adapts an important diagnostic tool for CWD, allowing investigation of the epidemiology of CWD at the community-level.

ecology↗

bioRxiv2023

Leptin activation of dorsal raphe neurons inhibits feeding behavior

Leptin is a homeostatic regulatory element that signals the presence of energy stores -in the form of adipocytes-which ultimately reduces food intake and increases energy expenditure. Similarly, serotonin (5-HT), a signaling molecule found in both the central and peripheral nervous systems, also regulates food intake. Here we use a combination of pharmacological manipulations, optogenetics, retrograde tracing, and in situ hybridization, combined with behavioral endpoints to physiologically and anatomically identify a novel leptin-mediated pathway between 5-HT neurons in the dorsal raphe nucleus (DRN) and hypothalamic arcuate nucleus (ARC) that controls food intake. In this study, we show that microinjecting leptin directly into the DRN reduces food intake in male Sprague-Dawley rats. This effect is mediated by leptin-receptor expressing neurons in the DRN as selective optogenetic activation of these neurons at either their ARC terminals or DRN cell bodies also reduces food intake. Anatomically, we identified a unique population of serotonergic raphe neurons expressing leptin receptors that send projections to the ARC. Finally, by utilizing in vivo microdialysis and high-performance liquid chromatography, we show that leptin administration to the DRN increases 5-HT efflux into the ARC. Overall, this study identifies a novel circuit for leptin-mediated control of food intake through a DRN-ARC pathway, utilizing 5-HT as a mechanism to control feeding behavior. Characterization of this new pathway creates opportunities for understanding how the brain controls eating behavior, as well as opens alternative routes for the treatment of eating disorders. Significance:Leptin and serotonin both play a vital role in the regulation of food intake, yet there is still uncertainty in how these two molecules interact to control appetite. The purpose of this study is to further understand the anatomical and functional connections between leptin receptor expressing neurons in the dorsal raphe nucleus, the main source of serotonin, and the arcuate nucleus of the hypothalamus, and how serotonin plays a role in this pathway to reduce food intake. Insight gained from this study will contribute to a more thorough understanding of the networks that regulate food intake, and open alternative avenues for the development of treatments for obesity and eating disorders.

neuroscience↗

bioRxiv2021

Time to stop; agent-based modelling of chemoaffinity with competition

In the classic Chemoaffinity theory, the retinotectal axon projection is thought to use pairs of orthogonal signalling gradients in the retina to specify the eventual location of synapses made on the surface of the tectum/superior colliculus. Similar orthogonal gradients in the tectum provide a coordinate system which allows the axons to match their prespecified destination with the correct location. Although the Ephrins have been shown to guide axons toward their destination, there has yet to emerge a complete account of the local interactions which halt the axonal growth cones in the correct locations to recreate the topography of the retinal cells. The model of Simpson and Goodhill (2011) provides an account of the basic topographic arrangement of cells on the tectum, as well as reproducing well known surgical and genetic manipulation experiments. However, it suffers from the absence of a local chemotactic guidance mechanism. Instead, each agent in their model is given instantaneous knowledge of the vector that would move it toward its pre specified destination. In addition to the globally supervised chemoaffinity term, Simpson and Goodhill (2011) introduced a competitive interaction for space between growth cone agents and a receptor-ligand axon-axon interaction in order to account for the full set of experimental manipulations. Here, we propose the replacement of the chemoaffinity term with a gradient following model consisting of axonal growth cone agents which carry receptor molecule expression determined by their somas location of origin on the retina. Growth cones move on the simulated tectum guided by two pairs of opposing, orthogonal signalling molecules representing the Ephrin ligands. We show that with only the chemoaffinity term and a receptor-ligand based axon-axon interaction term (meaning that all growth cone interactions are by receptor-ligand signalling), a full range of experimental manipulations to the retinotectal system can be reproduced. Furthermore, we show that the observation that competition is not and essential requirement for axons to find their way (Gosse et al., 2008) is also accounted for by the model, due to the opposing influences of signalling gradient pairs. Finally, we demonstrate that, assuming exponentially varying receptor expression in the retina, ligand expression should either be exponential if the receptor-ligand signal induces repulsion (i.e. gradient descent) or logarithmic if the signal induces attraction (gradient ascent). Thus, we find that a model analogous to the one we presented in James et al. (2020) that accounts for murine barrel patterning is also a candidate mechanism for the arrangement of the more continuous retinotectal system.

developmental biology↗

bioRxiv2021

Comparing cortex-wide gene expression patterns between species in a common reference frame

Advances in sequencing techniques have made comparative studies of gene expression a current focus for understanding evolutionary and developmental processes. However, insights into the spatial expression of genes have been limited by a lack of robust methodology. We therefore developed a set of algorithms for quantifying and comparing tissue-wide spatial patterns of gene expression within and across species. Here we apply these algorithms to compare cortex-wide expression of Id2 and RZR{beta} mRNA in early postnatal mice and voles. We show that neocortical patterns of Id2 expression are moderately conserved between species, but that the degree of conservation varies by cortical layer and area. By comparison, patterns of RZR{beta} expression are highly conserved in somatosensory areas, and more variable between species in visual and auditory areas. We consider if these differences reflect independent evolution in the 35 million years since the last common ancestor.

neuroscience↗

bioRxiv2021

Biological Action at a Distance: Correlated pattern formation in adjacent tessellation domains without communication

Tessellations emerge in many natural systems, and the constituent domains often contain regular patterns, raising the intriguing possibility that pattern formation within adjacent domains might be correlated by the geometry, without the direct exchange of information between parts comprising either domain. We confirm this paradoxical effect, by simulating pattern formation via reaction-diffusion in domains whose boundary shapes tessellate, and showing that correlations between adjacent patterns are strong compared to controls that self-organise in domains with equivalent sizes but unrelated shapes. The effect holds in systems with linear and non-linear diffusive terms, and for boundary shapes derived from regular and irregular tessellations. Based on the prediction that correlations between adjacent patterns should be bimodally distributed, we develop methods for testing whether a given set of domain boundaries constrained pattern formation within those domains. We then confirm such a prediction by analysing the development of subbarrel patterns, which are thought to emerge via reaction-diffusion, and whose enclosing borders form a Voronoi tessellation on the surface of the rodent somatosensory cortex. In more general terms, this result demonstrates how causal links can be established between the dynamical processes through which biological patterns emerge and the constraints that shape them.

developmental biology↗

bioRxiv2020

Infralimbic cortical glutamate output is necessary for the neural and behavioral consequences of chronic stress

Exposure to prolonged stress is a major risk-factor for psychiatric disorders such as generalized anxiety and major depressive disorder (MDD). Human imaging studies have identified structural and functional abnormalities in the prefrontal cortex of MDD patients, particularly Brodmanns area 25 (BA25). Further, deep brain stimulation of BA25 reduces symptoms of treatment-resistant depression. The rat homolog of BA25 is the infralimbic cortex (IL), which is critical for cognitive appraisal, executive function, and physiological stress reactivity. Previous studies indicate that the IL undergoes stress-induced changes in excitatory/inhibitory balance culminating in reduced activity of glutamate output neurons. However, the regulatory role of IL glutamate output in mood-related behaviors after chronic variable stress (CVS) is unknown. Here, we utilized a lentiviral-packaged small-interfering RNA to reduce translation of vesicular glutamate transporter 1 (vGluT1 siRNA), thereby constraining IL glutamate output. This viral-mediated gene transfer was used in conjunction with a quantitative anatomical analysis of cells expressing the stable immediate-early gene product {Delta}FosB, which accumulates in response to repeated neural activation. Through assessment of {Delta}FosB-expressing neurons across the frontal lobe in adult male rats, we mapped regions altered by chronic stress and determined the coordinating role of the IL in frontal cortical plasticity. Specifically, CVS-exposed rats had increased density of {Delta}FosB-expressing cells in the IL and decreased density in the anterior insula. The later effect was dependent on IL glutamate output. Next, we examined the interaction of CVS and reduced IL glutamate output in behavioral assays examining coping, anxiety-like behavior, associative learning, and nociception. IL glutamate knockdown decreased immobility during the forced swim test compared to GFP controls, both in rats exposed to CVS as well as rats without previous stress exposure. Further, vGluT1 siRNA prevented CVS-induced avoidance behaviors, while also reducing risk aversion and passive coping. Ultimately, this study identifies the necessity of IL glutamatergic output for regulating frontal cortical neural activity and behavior following chronic stress. These findings also highlight how disruption of excitatory/inhibitory balance within specific frontal cortical cell populations may impact neurobehavioral adaptation and lead to stress-related disorders.

neuroscience↗

bioRxiv2019

Modelling the emergence of whisker barrels

Brain development relies on an interplay between genetic specification and self-organization. Striking examples of this relationship can be found in the somatosensory brainstem, thalamus, and cortex of rats and mice, where the arrangement of the facial whiskers is preserved in the arrangement of cell aggregates to form precise somatotopic maps. We show in simulation how realistic whisker maps can self-organize, by assuming that information is exchanged between adjacent cells only, under the guidance of gene expression gradients. The resulting model provides a simple account of how patterns of gene expression can constrain spontaneous pattern formation to faithfully reproduce functional maps in subsequent brain structures.

developmental biology↗

bioRxiv2019

Prefrontal Cortex Regulates Chronic Stress-Induced Cardiovascular Susceptibility

The medial prefrontal cortex (mPFC) is necessary for appropriate appraisal of stressful information, as well as coordinating visceral and behavioral processes. However, prolonged stress impairs mPFC function and prefrontal-dependent behaviors. Additionally, chronic stress induces sympathetic predominance, contributing to health detriments associated with autonomic imbalance. Previous studies identified a subregion of rodent prefrontal cortex, infralimbic cortex (IL), as a key regulator of neuroendocrine-autonomic integration after chronic stress, suggesting that IL output may prevent chronic stress-induced autonomic imbalance. In the current study, we tested the hypothesis that the IL regulates hemodynamic, vascular, and cardiac responses to chronic stress. To address this hypothesis, a viral-packaged siRNA construct was used to knockdown vesicular glutamate transporter 1 (vGluT1) and reduce glutamate packaging and release from IL projection neurons. Male rats were injected with a vGluT1 siRNA-expressing construct or GFP control into the IL and then remained as unstressed controls or were exposed to chronic variable stress (CVS). IL vGluT1 knockdown increased heart rate and mean arterial pressure (MAP) reactivity, while CVS increased chronic MAP only in siRNA-treated rats. In a separate cohort, CVS and vGluT1 knockdown interacted to impair both endothelial-dependent and endothelial-independent vasoreactivity ex vivo. Furthermore, vGluT1 knockdown and CVS increased histological markers of fibrosis and hypertrophy. Thus, knockdown of glutamate release from IL projection neurons indicates that these cells are necessary to prevent the enhanced physiological responses to stress that promote susceptibility to cardiovascular pathophysiology. Ultimately, these findings provide evidence for a neurobiological mechanism mediating the relationship between stress and poor cardiovascular health outcomes.\n\nClinical perspectiveWhat is new? O_LIKnockdown of glutamate release from infralimbic cortex increases heart rate and arterial pressure reactivity\nC_LIO_LIDecreased infralimbic glutamate output leads to vascular dysfunction after chronic stress\nC_LIO_LIThese functional changes associate with histological indicators of cardiac hypertrophy, as well as vascular hypertrophy and fibrosis\nC_LI\n\nWhat are the clinical implications? O_LIThese studies provide a neurobiological mechanism that may account for the link between long-term stress and increased cardiovascular disease risk\nC_LI

neuroscience↗