Biology subjects

Holder, D.

Publications and source records attributed to Holder, D..

9 recordsLinked to original sources

bioRxiv2026

Extracellular matrix defects destabilise apical cytoarchitecture and mechanical properties during early Down syndrome neurodevelopment

Down syndrome (DS) is associated with altered brain development, especially in the cerebellum. However, how trisomy 21 (TS21) is linked to cerebellar defects remains poorly understood. Cerebellar organoids reveal that TS21 leads to extracellular matrix (ECM) alterations, perturbing downstream pathways linked to reduction of apical RAB11+ endosomes. These changes lead to impaired apical maintenance and altered progenitor composition. Significantly, ECM-enriched culture rescues apical defects in TS21 organoids, supporting a functional role for ECM in preserving apical organisation and progenitor niche integrity. In the developing human DS cerebellum and 2D neural cultures, where exogenous ECM is present, severe epithelial disorganisation is attenuated. Nevertheless, ECM organisation and mechanical properties remain altered in 2D cultures. Our findings identify TS21-driven ECM abnormalities as a mechanism impairing apical integrity, progenitor composition and mechanical proprieties in neural stem cell models of DS. Altogether, we established apical maintenance instability as an early developmental defect contributing for DS neuropathology.

Developmental Biology↗

bioRxiv2026

An Implantable Wireless Battery-Free Selective Vagus Nerve Stimulator

Objective. Vagus nerve stimulation (VNS) is an established clinical therapy for drug-resistant epilepsy and other inflammatory conditions. However, off-target stimulation can produce unwanted side effects that limit therapeutic stimulation and hinder the development of new neuromodulation therapies. Selective VNS (sVNS) offers a strategy to reduce off-target organ activation; however, this approach is not available in humans, and no implantable or portable devices exist to trial sVNS in the clinical setup. This work aimed to design, manufacture, and validate an implantable wireless, battery-free stimulator with a selectively addressable output stage for targeted current delivery to specific regions of the human vagus nerve (VN). Approach. We developed a near-field communication-controlled, wirelessly powered, battery-free, temporary implantable multichannel stimulation device, compatible with a 15-channel sVNS cuff electrode (14 selective electrode pairs and one circumferential whole-nerve channel). The device was encapsulated for short-term implantation and evaluated through benchtop characterisation, accelerated ageing, and validation in an acute porcine and a pilot human study. Main result. The sVNS device was evaluated in a porcine (n = 4) trial and a first-in-human pilot study (n = 1). Selective bradycardia of 23.28 {+/-} 12.91% was observed in pigs and 7.5% in the human participant. In the human, a clear separation of bradycardic and tachycardic effects was observed, with additional selectivity in laryngeal activity. Cardiac and laryngeal responses were separated by 231{degrees} around the circumference of the nerve. Significance. This work demonstrates the feasibility of wireless battery-free sVNS for cardiac applications using a temporary implantable device. Geometrically selective stimulation has the potential to improve therapeutic efficacy while reducing stimulation-related side effects, and may facilitate future therapies for heart failure and other autonomic disorders.

bioengineering↗

bioRxiv2025

Evaluation of Aggregate Oral Fluid Sampling for Early Detection of African Swine Fever Virus Infection

African swine fever (ASF) is a highly infectious viral disease that poses significant threat to the United States and global pig industries. Given the lack of effective vaccines, control and prevention of the spread of African swine fever virus (ASFV) is dependent on enhanced surveillance and early disease detection. Commercial swine operations in the US are characterized by comparatively large number of pigs, and sampling individual pigs, which represents the main strategy for current ASF surveillance, is both costly and labor intensive. The major objective of this study was to estimate the diagnostic sensitivity of pen-based aggregate oral fluid testing for ASFV in infected pigs in a pen of 30 animals and evaluate its utility as a tool to support surveillance of ASF in the United States. The study was performed in three phases: (i) Virus (Ghana ASFV24) amplification in a target host species to generate the challenge inoculum, (ii) Titration of the inoculum (10% spleen homogenate) in target host species to determine the minimum dose inducing acute ASF in pigs with survival up to 5 - 6 days post-inoculation (dpi), and (iii) The main study involving 186 pigs consisting of 6 replicates of 30 pigs per pen and one seeder pig inoculated with the Ghana ASFV24 per pen. Daily sampling of aggregate oral fluids, uncoagulated blood, oropharyngeal swabs, fecal and water nipple swabs, and recording of rectal temperatures and clinical observations, was carried out. The seeder pigs were each inoculated intramuscularly with 0.5 ml of the 10% spleen homogenate which induced the desired clinical course of ASF in the pigs with survival of up to 6 dpi. ASFV DNA could be detected in the seeder pigs as early as 1 dpi and 2 dpi in the blood and oropharyngeal swabs, respectively. Transmission of ASFV from the seeder pigs to the contact pig population was detected via positive amplification of ASFV DNA in aggregate oral fluid samples at 3 days post-contact (dpc) in 4 out of 6 pens, and in all 6 pens at 4 dpc. Testing of oropharyngeal swabs and blood samples from individual pigs revealed variable number of ASFV positive pigs between 3 and 5 dpc, with detection of 100% positivity between 6 and 18 dpc, the study endpoint. These findings demonstrate the potential utility of aggregate oral fluid sampling for sensitive and early detection of ASFV incursion into naive swine herds. It also demonstrates that testing of environmental samples from the premises could further enhance overall ASF early detection and surveillance strategy. Author summaryEarly detection of ASFV in swine farms requires robust passive surveillance using sample types and sampling methods that allow sensitive and timely detection. Commercial swine operations in the US or North America are characterized by comparatively large number of pigs, and sampling individual pigs, which represents the current strategy for ASF surveillance, is both costly and labor-intensive. Oral fluid has been shown to be an acceptable sample type for detection of ASFV in individual infected pigs. For the first time on such a scale, we conducted a study enrolling 186 pigs, with daily sampling, in six experimental replicates at 3.2% pen prevalence using a highly virulent ASFV (Ghana ASFV24) to evaluate the utility of aggregate oral fluids for early detection of ASFV. Whole genome sequencing and characterization confirmed grouping of the virus with those in the p72 genotype II cluster. We demonstrate that intramuscular inoculation of the seeder pigs induces acute ASF and transmission to the contact pigs which is detectable in aggregate oral fluids as early as 3 - 4 dpc. We have shown that ASFV DNA detections in aggregate oral fluids correlate with the oropharyngeal swabs of individual pigs. In comparison, ASFV DNA in individual blood samples is detected 1-3 days later. The study demonstrates the potential utility of aggregate oral fluid sampling for enhanced surveillance of ASFV in large commercial swine operations.

microbiology↗

bioRxiv2024

Functional architecture of intracellular oscillations in hippocampal dendrites

Fast electrical signaling in dendrites is central to neural computations that support adaptive behaviors. Conventional techniques lack temporal and spatial resolution and the ability to track underlying membrane potential dynamics present across the complex three-dimensional dendritic arbor in vivo. Here, we perform fast two-photon imaging of dendritic and somatic membrane potential dynamics in single pyramidal cells in the CA1 region of the mouse hippocampus during awake behavior. We study the dynamics of subthreshold membrane potential and suprathreshold dendritic events throughout the dendritic arbor in vivo by combining voltage imaging with simultaneous local field potential recording, post hoc morphological reconstruction, and a spatial navigation task. We systematically quantify the modulation of local event rates by locomotion in distinct dendritic regions and report an advancing gradient of dendritic theta phase along the basal-tuft axis, then describe a pre-dominant hyperpolarization of the dendritic arbor during sharp-wave ripples. Finally, we find spatial tuning of dendritic representations dynamically reorganizes following place field formation. Our data reveal how the organization of electrical signaling in dendrites maps onto the anatomy of the dendritic tree across behavior, oscillatory network, and functional cell states.

neuroscience↗

bioRxiv2024

Anatomical and functional organization of cardiac fibers in the porcine cervical vagus nerve allows spatially selective efferent neuromodulation

Cardiac disease progression reflects the dynamic interaction between adversely remodeled neurohumoral control systems and an abnormal cardiac substrate. Vagal nerve stimulation (VNS) is an attractive neuromodulatory option to dampen this dynamic interaction; however, it is limited by off-target effects. Spatially-selective VNS (sVNS) offers a promising solution to induce cardioprotection while mitigating off-target effects by specifically targeting pre-ganglionic parasympathetic efferent cardiac fibers. This approach also has the potential to enhance therapeutic outcomes by eliminating time-consuming titration required for optimal VNS. Recent studies have demonstrated the independent modulation of breathing rate, heart rate, and laryngeal contraction through sVNS. However, the spatial organization of afferent and efferent cardiac-related fibers within the vagus nerve remains unexplored. By using trial-and-error sVNS in vivo in combination with ex vivo micro-computed tomography fascicle tracing, we show the significant spatial separation of cardiac afferent and efferent fibers (179{+/-}55{degrees} SD microCT, p<0.05 and 200{+/-}137{degrees} SD, p<0.05 sVNS - degrees of separation across a cross-section of nerve) at the mid-cervical level. We also show that cardiac afferent fibers are located in proximity to pulmonary fibers consistent with recent findings of cardiopulmonary convergent neurons and circuits. We demonstrate the ability of sVNS to selectively elicit desired scalable heart rate decrease without stimulating afferent-related reflexes. By elucidating the spatial organization of cardiac-related fibers within the vagus nerve, our findings pave the way for more targeted neuromodulation, thereby reducing off-target effects and eliminating the need for titration. This, in turn, will enhance the precision and efficacy of VNS therapy in treating cardiac pathology, allowing for improved therapeutic efficacy. Condensed AbstractSpatially-selective vagus nerve stimulation (sVNS) presents a promising approach for addressing chronic heart disease with enhanced precision. Our study reveals significant spatial separation between cardiac afferent and efferent fibers in the vagus nerve, particularly at the mid-cervical level. Utilizing trial-and-error sVNS in vivo and micro-computed tomography fascicle tracing, we demonstrate the potential for targeted neuromodulation, achieving therapeutic effects like scalable heart rate decrease without stimulating afferent-related reflexes. This spatial understanding opens avenues for more effective VNS therapy, minimizing off-target effects and eliminating the need for titration, thereby expediting therapeutic outcomes in myocardial infarction and related conditions. TweetWith functional and structural imaging, we found organization of vagal efferent & afferent cardiac regions. We can selectively activate only cardiac efferents to achieve bradycardia; desired to reduce the effects of sympathetic overactivation associated with heart disease #VNS #Cardiac #VagusNerve Key PointsO_LISpatially-selective vagus nerve stimulation (sVNS) presents a promising approach for addressing chronic heart disease with enhanced precision. C_LIO_LIOur study reveals significant spatial separation between cardiac afferent and efferent fibers in the vagus nerve, particularly at the mid-cervical level. C_LIO_LIUtilizing trial-and-error sVNS in vivo and micro-computed tomography fascicle tracing, we demonstrate the potential for targeted neuromodulation, achieving therapeutic effects like scalable heart rate decrease without stimulating afferent-related reflexes. C_LIO_LIThis spatial understanding opens avenues for more effective VNS therapy, minimizing off-target effects and eliminating the need for titration, thereby expediting therapeutic outcomes in myocardial infarction and related conditions. C_LI

neuroscience↗

bioRxiv2023

Human vagus nerve fascicular anatomy: a microCT segmentation and histological study

BackgroundPrevious research has revealed the logical mapping of fascicles in both human somatic and pig vagus nerves, but the organization of fascicles within the human vagus nerve remains largely unknown. Understanding its fascicular arrangement would significantly advance our knowledge of the autonomic nervous system and facilitate studies and application of selective vagus nerve stimulation to avoid off-target effects. The purpose of this study was to trace the thoracic branches of human vagus nerves, investigate their fascicular organization, and analyze the nerves histologically and morphologically. MethodsBoth left and right vagus nerves were dissected from human cadavers, preserving the cardiac, recurrent laryngeal, and pulmonary branches. The nerves were prepared, scanned using microCT, and the fascicles segmented and traced from their branching points. Histology and immunohistochemistry were performed for morphological analysis and validation of the microCT segmentation. The data was then analyzed and compared between nerves. ResultsThe organization of the cardiac, pulmonary, and recurrent laryngeal fascicles was observed for a short distance from their entry point into the nerves. Initially, left vagus nerves showed merging of cardiac and pulmonary fascicles, while the recurrent laryngeal fascicles remained separate. In right vagus nerves, the cardiac fascicles merged with both pulmonary and recurrent laryngeal fascicles. MicroCT imaging limitations prevented visualization and tracing of fiber organization within merged fascicles. Immunohistochemistry and morphological analysis revealed that right vagus nerves were larger and had more fascicles than the left and fascicle counts varied along the nerve, indicating anastomoses. The superior cardiac branch was separate from other fascicles near the VNS cuff placement. ConclusionsIt is possible that organ-specific fibers may still retain some spatial organization despite most fascicles being merged at cervical level. However, fiber tracing and in vivo studies could provide valuable information beyond microCT to resolve this further. The separate superior cardiac fascicles offer potential for targeted neuromodulation of the heart, benefiting conditions like myocardial infarction, heart failure, and atrial fibrillation. Overall, the study provides insights into the morphology and anatomy of human vagus nerves. Our findings thereby contribute to the development of selective vagus nerve stimulation strategies for more precise autonomic regulation.

neuroscience↗

bioRxiv2022

Organotopic organisation of the cervical vagus nerve

Despite detailed characterization of fascicular organization of somatic nerves, the functional anatomy of fascicles evident in human and large mammal cervical vagus nerve is unknown. The vagus nerve is a prime target for intervention in the field of electroceuticals due to its extensive distribution to the heart, larynx, lungs, and abdominal viscera. However, current practice of the approved vagus nerve stimulation (VNS) technique is to stimulate the entire nerve. This produces indiscriminate stimulation of non-targeted effectors and undesired side effects. Selective neuromodulation is now a possibility with a spatially-selective vagal nerve cuff. However, this requires the knowledge of the fascicular organization at the level of cuff placement to inform selectivity of only the desired target organ or function. We imaged function over milliseconds with fast neural electrical impedance tomography and selective stimulation, and found consistent spatially separated regions within the nerve correlating with the three fascicular groups of interest, suggesting organotopy. This was independently verified with structural imaging by tracing anatomical connections from the end organ with microCT and the development of an anatomical map of the vagus nerve. This confirmed organotopic organization. Here we show, for the first time, localized fascicles in the porcine cervical vagus nerve which map to cardiac, pulmonary and recurrent laryngeal function (N=4). These findings pave the way for improved outcomes in VNS as unwanted side effects could be reduced by targeted selective stimulation of identified organ-specific fascicles and the extension of this technique clinically beyond the currently approved disorders to treat heart failure, chronic inflammatory disorders and more.

neuroscience↗

bioRxiv2020

Imaging fascicular organization of peripheral nerves with fast neural Electrical Impedance Tomography (EIT)

Imaging of the compound action potential (CAP) in fascicles in peripheral nerves could help avoid side effects in neuromodulation by selective stimulation of identified fascicles. Existing methods have low resolution, limited imaging depth, or are invasive. We propose fast neural electrical impedance tomography (EIT), which allows fascicular CAP imaging with a high resolution of [~]200 m, <1 ms. This uses a non-penetrating flexible cuff electrode array with 14 circumferential electrodes. This has been validated in rat sciatic nerve by comparison to micro-computed tomography (microCT) and histology with fluorescent dextran tracers (n=5). With EIT, there were reproducible localized changes in tissue impedance in response to stimulation of individual fascicles (tibial, peroneal and sural). The reconstructed EIT images corresponded to microCT scans and neural tracer histology, with significant separation between the fascicles (p<0.01), and no significant difference between techniques. The standard deviation from the mean fascicle position for EIT was 86 m (6% of nerve diameter). This suggests fast neural EIT can reliably image the functional fascicular anatomy of the nerves and so aid selective neuromodulation.

bioengineering↗

bioRxiv2019

MicroCT optimisation for imaging fascicular anatomy in peripheral nerves

Vagus nerve stimulation (VNS) is a promising therapy for treatment of various conditions resistant to standard therapeutics. However, due to the lack of understanding of the fascicular organisation of the vagus nerve, VNS leads to unwanted off-target effects. Micro-computed tomography (microCT) can be used to trace fascicles from periphery and image fascicular anatomy. In this work we optimised the microCT protocol of the rat sciatic and subsequent pig vagus nerves.\n\nAfter differential staining, the optimal staining time was selected and scanning parameters were altered in subsequent scans. Scans were reconstructed, visualised in ImageJ and fascicles segmented with a custom algorithm in Matlab to determine ultimate parameters for tracking of the nerve. Successful segmentation for tracking of individual fascicles was achieved after 24 hours and 120 hours of staining with Lugols solution (1% total iodine) for rat sciatic and pig vagus nerves, respectively, and the following scanning parameters: 4 {micro}m voxel size, 35 kVp energy, 114 {micro}A current, 4 W power, 0.25 fps in 4 s exposure time, 3176 projections and a molybdenum target.\n\nThe optimised microCT protocol allows for segmentation and tracking of the fascicles within the nerve. This will be used to scan the full length of the pig, and possibly, the human vagus nerves. The resulting segmentation map of the functional anatomical organisation of the vagus nerve will enable selective VNS ultimately allowing for the avoidance of the off-target effects and improving its therapeutic efficacy.

neuroscience↗