Biology subjects

Turschak, E.

Publications and source records attributed to Turschak, E..

6 recordsLinked to original sources

bioRxiv2025

Evaluating Microglial Contributions to the Neurovascular Unit in Health and Neurodegeneration Using Human In Vitro Models

BackgroundMicroglia are emerging as critical regulators of neurovascular function in health and Alzheimers disease (AD), yet their interactions with the human neurovascular unit (NVU), particularly brain endothelial cells, remain incompletely understood. Current in vitro NVU platforms typically exclude microglia and lack perfusable vascular networks with physiologically relevant architecture. Here, we established complementary two-dimensional (2D) and three-dimensional (3D) NVU models to investigate microglia-endothelial and microglia-neurovascular interactions. MethodsHuman induced pluripotent stem cell derived-neurons (iNs), astrocytes (iAs), and microglia-like cells (iMGLs) were incorporated into a soft-lithography based engineered microvessel system to establish a multicellular neuroimmune-vascular model. To specifically evaluate iMGL-endothelial cell (EC) interactions, iMGL were co-cultured with primary human brain microvascular endothelial cells (HBMECs) and junctional protein localization was evaluated using immunofluorescence. The barrier integrity of engineered microvessels containing iMGL was evaluated using dextran permeability. Our 2D and 3D systems were stimulated with tumor necrosis factor- (TNF) to evaluate whether iMGL would promote or attenuate EC inflammation and barrier breakdown. ResultsIncorporation of iNs, iAs, and iMGLs into a perfusable vascular model enabled a more complete representation of NVU cellular diversity and promoted neuronal health. In monolayer co-culture with iMGL, HBMECs enhanced the junctional localization of tight and adherens junction proteins through both contact-dependent and paracrine mechanisms. Following an inflammatory challenge, iMGLs reduced endothelial inflammatory activation, suggesting a protective role in response to AD-relevant inflammatory conditions. Finally, when embedded in 3D collagen matrices surrounding perfusable endothelialized lumen networks, iMGLs reduced dextran permeability and preserved endothelial barrier integrity following TNF challenge. ConclusionsTogether, these findings establish a 3D perfusable neuroimmune-vascular model that enables the dissection of microglial contributions to neurovascular function in health and disease.

cell biology↗

bioRxiv2024

A scalable and modular computational pipeline for axonal connectomics: automated tracing and assembly of axons across serial sections

Progress in histological methods and in microscope technology has enabled dense staining and imaging of axons over large brain volumes, but tracing axons over such volumes requires new computational tools for 3D reconstruction of data acquired from serial sections. We have developed a computational pipeline for automated tracing and volume assembly of densely stained axons imaged over serial sections, which leverages machine learning-based segmentation to enable stitching and alignment with the axon traces themselves. We validated this segmentation-driven approach to volume assembly and alignment of individual axons over centimeter-scale serial sections and show the application of the output traces for analysis of local orientation and for proofreading over aligned volumes. The pipeline is scalable, and combined with recent advances in experimental approaches, should enable new studies of mesoscale connectivity and function over the whole human brain.

neuroscience↗

bioRxiv2023

Microscale visualization of cellular features in adult macaque visual cortex

Expansion microscopy and light sheet imaging enable fine-scale resolution of intracellular features that comprise neural circuits. Most current techniques visualize sparsely distributed features across whole brains or densely distributed features within individual brain regions. Here, we visualize dense distributions of immunolabeled proteins across early visual cortical areas in adult macaque monkeys. This process may be combined with multiphoton or magnetic resonance imaging to produce multimodal atlases in large, gyrencephalic brains.

neuroscience↗

bioRxiv2023

Expansion-assisted selective plane illumination microscopy for nanoscale imaging ofcentimeter-scale tissues

Recent advances in tissue processing, labeling, and fluorescence microscopy are providing unprecedented views of the structure of cells and tissues at sub-diffraction resolutions and near single molecule sensitivity, driving discoveries in diverse fields of biology, including neuroscience. Biological tissue is organized over scales of nanometers to centimeters. Harnessing molecular imaging across intact, three-dimensional samples on this scale requires new types of microscopes with larger fields of view and working distance, as well as higher throughput. We present a new expansion-assisted selective plane illumination microscope (ExA-SPIM) with aberration-free 1.5 {micro}mx1.5 {micro}mx3 {micro}m optical resolution over a large field of view (10.6x8.0 mm2) and working distance (35 mm) at speeds up to 946 megavoxels/sec. Combined with new tissue clearing and expansion methods, the microscope allows imaging centimeter-scale samples with 375 nm lateral and 750 nm axial resolution (4x expansion), including entire mouse brains, with high contrast and without sectioning. We illustrate ExA-SPIM by reconstructing individual neurons across the mouse brain, imaging cortico-spinal neurons in the macaque motor cortex, and visualizing axons in human white matter.

neuroscience↗

bioRxiv2021

Motion/direction-sensitive thalamic neurons project extensively to the middle layers of primary visual cortex

The motion/direction-sensitive and location-sensitive neurons are two major functional types in mouse visual thalamus that project to the primary visual cortex (V1). It has been proposed that the motion/direction-sensitive neurons mainly target the superficial layers in V1, in contrast to the location-sensitive neurons which mainly target the middle layers. Here, by imaging calcium activities of motion/direction-sensitive and location-sensitive axons in V1, we find no evidence for these cell-type specific laminar biases at population level. Furthermore, using a novel approach to reconstruct single-axon structures with identified in vivo response types, we show that, at single-axon level, the motion/direction-sensitive axons have middle layer preferences and project more densely to the middle layers than the location-sensitive axons. Overall, our results demonstrate that Motion/direction-sensitive thalamic neurons project extensively to the middle layers of V1, challenging the current view of the thalamocortical organizations in the mouse visual system.

neuroscience↗

bioRxiv2018

Activation of neuromodulatory axon projections in primary visual cortex during periods of locomotion and pupil dilation

Neuromodulators such as acetylcholine, noradrenaline (norepinephrine), and serotonin are released into the cortex by axons ascending from subcortical nuclei. These neuromodulators have been hypothesized to influence cortical function during behavioral periods such as arousal, locomotion, exploration, and attention. To determine when these neuromodulatory projections were active, we expressed the genetically-encoded calcium sensor GCaMP6 in neuromodulatory axons which project to the mouse primary visual cortex and performed two-photon microscopy to monitor their activity in vivo. We observed that the fluorescence of both cholinergic and noradrenergic axons increased during periods of pupil dilation, with the fluorescence of the axons rising less than one second before eye pupil dilation. We also observed increases in cholinergic and noradrenergic axon fluorescence periods of locomotion, which was accompanied by pupil dilation and nasal (forward) movement of both pupils. Locomotion was preceded by a rise in axonal fluorescence with a timing and amplitude that matched the subsequent pupil dilation, but axon fluorescence was more sustained than expected from the pupil dilation, suggesting that there is an additional physiological factor that affects cholinergic and noradrenergic axon activity in primary visual cortex during locomotion.

neuroscience↗