Large-scale characterization of the microvascular geometry in development and disease by tissue clearing and quantitative ultramicroscopy

Three-dimensional assessment of optically cleared, entire organs and organisms has recently become possible by tissue clearing and selective plane illumination microscopy (“ultramicroscopy”). Resulting datasets can be highly complex, encompass over a thousand images with millions of objects and data of several gigabytes per acquisition. This constitutes a major challenge for quantitative analysis. We have developed post-processing tools to quantify millions of microvessels and their distribution in three-dimensional datasets from ultramicroscopy and demonstrate the capabilities of our pipeline within entire mouse brains and embryos. Using our developed acquisition, segmentation, and analysis platform, we quantify physiological vascular networks in development and the healthy brain. We compare various geometric vessel parameters (e.g. vessel density, radius, tortuosity) in the embryonic spinal cord and brain as well as in different brain regions (basal ganglia, corpus callosum, cortex). White matter tract structures (corpus callosum, spinal cord) showed lower microvascular branch densities and longer vessel branch length compared to grey matter (cortex, basal ganglia). Furthermore, we assess tumor neoangiogenesis in a mouse glioma model to compare tumor core and tumor border. The developed methodology allows rapid quantification of three-dimensional datasets by semi-automated segmentation of fluorescently labeled objects with conventional computer hardware. Our approach can aid preclinical investigations and paves the way towards “quantitative ultramicroscopy”.

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Symptoms, Related Brain Regions, and Diagnosis

Dementia with Lewy Body and Parkinson's Disease Patients ◽

10.1093/oso/9780199977567.003.0008 ◽

2013 ◽

Author(s):

J. Eric Ahlskog

Keyword(s):

Spinal Cord ◽

Nervous System ◽

Basal Ganglia ◽

Brain Stem ◽

Muscle Activation ◽

Sensory Information ◽

Brain Regions ◽

Electrical Signal ◽

Brain And Spinal Cord ◽

The Brain

As a prelude to the treatment chapters that follow, we need to define and describe the types of problems and symptoms encountered in DLB and PDD. The clinical picture can be quite varied: problems encountered by one person may be quite different from those encountered by another person, and symptoms that are problematic in one individual may be minimal in another. In these disorders, the Lewy neurodegenerative process potentially affects certain nervous system regions but spares others. Affected areas include thinking and memory circuits, as well as movement (motor) function and the autonomic nervous system, which regulates primary functions such as bladder, bowel, and blood pressure control. Many other brain regions, by contrast, are spared or minimally involved, such as vision and sensation. The brain and spinal cord constitute the central nervous system. The interface between the brain and spinal cord is by way of the brain stem, as shown in Figure 4.1. Thought, memory, and reasoning are primarily organized in the thick layers of cortex overlying lower brain levels. Volitional movements, such as writing, throwing, or kicking, also emanate from the cortex and integrate with circuits just below, including those in the basal ganglia, shown in Figure 4.2. The basal ganglia includes the striatum, globus pallidus, subthalamic nucleus, and substantia nigra, as illustrated in Figure 4.2. Movement information is integrated and modulated in these basal ganglia nuclei and then transmitted down the brain stem to the spinal cord. At spinal cord levels the correct sequence of muscle activation that has been programmed is accomplished. Activated nerves from appropriate regions of the spinal cord relay the signals to the proper muscles. Sensory information from the periphery (limbs) travels in the opposite direction. How are these signals transmitted? Brain cells called neurons have long, wire-like extensions that interface with other neurons, effectively making up circuits that are slightly similar to computer circuits; this is illustrated in Figure 4.3. At the end of these wire-like extensions are tiny enlargements (terminals) that contain specific biological chemicals called neurotransmitters. Neurotransmitters are released when the electrical signal travels down that neuron to the end of that wire-like process.

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Aberrant White Matter Networks Mediate Cognitive Impairment in Patients with Silent Lacunar Infarcts in Basal Ganglia Territory

Journal of Cerebral Blood Flow & Metabolism ◽

10.1038/jcbfm.2015.67 ◽

2015 ◽

Vol 35 (9) ◽

pp. 1426-1434 ◽

Cited By ~ 9

Author(s):

Jinfu Tang ◽

Suyu Zhong ◽

Yaojing Chen ◽

Kewei Chen ◽

Junying Zhang ◽

...

Keyword(s):

White Matter ◽

Basal Ganglia ◽

Cognitive Deficits ◽

Large Scale ◽

Diffusion Tensor ◽

Brain Regions ◽

Lacunar Infarcts ◽

Healthy Elderly ◽

Brain White Matter ◽

Diffusion Tensor Imaging Tractography

Silent lacunar infarcts, which are present in over 20% of healthy elderly individuals, are associated with subtle deficits in cognitive functions. However, it remains largely unclear how these silent brain infarcts lead to cognitive deficits and even dementia. Here, we used diffusion tensor imaging tractography and graph theory to examine the topological organization of white matter networks in 27 patients with silent lacunar infarcts in the basal ganglia territory and 30 healthy controls. A whole-brain white matter network was constructed for each subject, where the graph nodes represented brain regions and the edges represented interregional white matter tracts. Compared with the controls, the patients exhibited a significant reduction in local efficiency and global efficiency. In addition, a total of eighteen brain regions showed significantly reduced nodal efficiency in patients. Intriguingly, nodal efficiency–behavior associations were significantly different between the two groups. The present findings provide new aspects into our understanding of silent infarcts that even small lesions in subcortical brain regions may affect large-scale cortical white matter network, as such may be the link between subcortical silent infarcts and the associated cognitive impairments. Our findings highlight the need for network-level neuroimaging assessment and more medical care for individuals with silent subcortical infarcts.

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Optical imaging and modulation of neurovascular responses

Journal of Cerebral Blood Flow & Metabolism ◽

10.1177/0271678x18803372 ◽

2018 ◽

Vol 38 (12) ◽

pp. 2057-2072 ◽

Cited By ~ 5

Author(s):

Kazuto Masamoto ◽

Alberto Vazquez

Keyword(s):

Optical Imaging ◽

Vascular Function ◽

Imaging Techniques ◽

Three Dimensional ◽

Brain Regions ◽

Transgenic Animal ◽

Optical Methods ◽

Vascular Networks ◽

Temporal Properties ◽

Dimensional Reconstruction

The cerebral microvasculature consists of pial vascular networks, parenchymal descending arterioles, ascending venules and parenchymal capillaries. This vascular compartmentalization is vital to precisely deliver blood to balance continuously varying neural demands in multiple brain regions. Optical imaging techniques have facilitated the investigation of dynamic spatial and temporal properties of microvascular functions in real time. Their combination with transgenic animal models encoding specific genetic targets have further strengthened the importance of optical methods for neurovascular research by allowing for the modulation and monitoring of neuro vascular function. Image analysis methods with three-dimensional reconstruction are also helping to understand the complexity of microscopic observations. Here, we review the compartmentalized cerebral microvascular responses to global perturbations as well as regional changes in response to neural activity to highlight the differences in vascular action sites. In addition, microvascular responses elicited by optical modulation of different cell-type targets are summarized with emphasis on variable spatiotemporal dynamics of microvascular responses. Finally, long-term changes in microvascular compartmentalization are discussed to help understand potential relationships between CBF disturbances and the development of neurodegenerative diseases and cognitive decline.

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Cellular maps of gastrointestinal organs: getting the most from tissue clearing

AJP Gastrointestinal and Liver Physiology ◽

10.1152/ajpgi.00075.2020 ◽

2020 ◽

Vol 319 (1) ◽

pp. G1-G10

Author(s):

Cambrian Y. Liu ◽

D. Brent Polk

Keyword(s):

Large Scale ◽

Alimentary Tract ◽

Three Dimensional ◽

Short Review ◽

Biological Tissues ◽

Tissue Clearing ◽

Relative Paucity ◽

Whole Mount ◽

Chemical Concepts ◽

The Impact

The development of modern methods to induce optical transparency (“clearing”) in biological tissues has enabled the three-dimensional (3D) reconstruction of intact organs at cellular resolution. New capabilities in visualization of rare cellular events, long-range interactions, and irregular structures will facilitate novel studies in the alimentary tract and gastrointestinal systems. The tubular geometry of the alimentary tract facilitates large-scale cellular reconstruction of cleared tissue without specialized microscopy setups. However, with the rapid pace of development of clearing agents and current relative paucity of research groups in the gastrointestinal field using these techniques, it can be daunting to incorporate tissue clearing into experimental workflows. Here, we give some advice and describe our own experience bringing tissue clearing and whole mount reconstruction into our laboratory’s investigations. We present a brief overview of the chemical concepts that underpin tissue clearing, what sorts of questions whole mount imaging can answer, how to choose a clearing agent, an example of how to clear and image alimentary tissue, and what to do after obtaining the image. This short review will encourage other gastrointestinal researchers to consider how utilizing tissue clearing and creating 3D “maps” of tissue might deepen the impact of their studies.

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Perspective: Extending the Utility of Three-Dimensional Organoids by Tissue Clearing Technologies

Frontiers in Cell and Developmental Biology ◽

10.3389/fcell.2021.679226 ◽

2021 ◽

Vol 9 ◽

Author(s):

Etsuo A. Susaki ◽

Minoru Takasato

Keyword(s):

Large Scale ◽

3D Imaging ◽

Imaging Techniques ◽

3D Structure ◽

Disease Onset ◽

Three Dimensional ◽

Cell Labeling ◽

Tissue Clearing ◽

Cellular Components ◽

3D Imaging Techniques

An organoid, a self-organizing organ-like tissue developed from stem cells, can exhibit a miniaturized three-dimensional (3D) structure and part of the physiological functions of the original organ. Due to the reproducibility of tissue complexity and ease of handling, organoids have replaced real organs and animals for a variety of uses, such as investigations of the mechanisms of organogenesis and disease onset, and screening of drug effects and/or toxicity. The recent advent of tissue clearing and 3D imaging techniques have great potential contributions to organoid studies by allowing the collection and analysis of 3D images of whole organoids with a reasonable throughput and thus can expand the means of examining the 3D architecture, cellular components, and variability among organoids. Genetic and histological cell-labeling methods, together with organoid clearing, also allow visualization of critical structures and cellular components within organoids. The collected 3D data may enable image analysis to quantitatively assess structures within organoids and sensitively/effectively detect abnormalities caused by perturbations. These capabilities of tissue/organoid clearing and 3D imaging techniques not only extend the utility of organoids in basic biology but can also be applied for quality control of clinical organoid production and large-scale drug screening.

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A Probabilistic, Distributed, Recursive Mechanism for Decision-making in the Brain

10.1101/036277 ◽

2016 ◽

Author(s):

Javier A. Caballero ◽

Mark D. Humphries ◽

Kevin N. Gurney

Keyword(s):

Decision Making ◽

Basal Ganglia ◽

Neural Activity ◽

Large Scale ◽

Single Equation ◽

Brain Regions ◽

Compulsive Gambling ◽

Choice Behaviour ◽

Usable Information ◽

The Brain

AbstractDecision formation recruits many brain regions, but the procedure they jointly execute is unknown. Here we characterize its essential composition, using as a framework a novel recursive Bayesian algorithm that makes decisions based on spike-trains with the statistics of those in sensory cortex (MT). Using it to simulate the random-dot-motion task, we demonstrate it quantitatively replicates the choice behaviour of monkeys, whilst predicting losses of otherwise usable information from MT. Its architecture maps to the recurrent cortico-basal-ganglia-thalamo-cortical loops, whose components are all implicated in decision-making. We show that the dynamics of its mapped computations match those of neural activity in the sensorimotor cortex and striatum during decisions, and forecast those of basal ganglia output and thalamus. This also predicts which aspects of neural dynamics are and are not part of inference. Our single-equation algorithm is probabilistic, distributed, recursive, and parallel. Its success at capturing anatomy, behaviour, and electrophysiology suggests that the mechanism implemented by the brain has these same characteristics.Author SummaryDecision-making is central to cognition. Abnormally-formed decisions characterize disorders like over-eating, Parkinson’s and Huntington’s diseases, OCD, addiction, and compulsive gambling. Yet, a unified account of decisionmaking has, hitherto, remained elusive. Here we show the essential composition of the brain’s decision mechanism by matching experimental data from monkeys making decisions, to the knowable function of a novel statistical inference algorithm. Our algorithm maps onto the large-scale architecture of decision circuits in the primate brain, replicating the monkeys’ choice behaviour and the dynamics of the neural activity that accompany it. Validated in this way, our algorithm establishes a basic framework for understanding the mechanistic ingredients of decisionmaking in the brain, and thereby, a basic platform for understanding how pathologies arise from abnormal function.

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High-Resolution Three-Dimensional Extracellular Recording of Neuronal Activity With Microfabricated Electrode Arrays

Journal of Neurophysiology ◽

10.1152/jn.90992.2008 ◽

2009 ◽

Vol 101 (3) ◽

pp. 1671-1678 ◽

Cited By ~ 47

Author(s):

Jiangang Du ◽

Ingmar H. Riedel-Kruse ◽

Janna C. Nawroth ◽

Michael L. Roukes ◽

Gilles Laurent ◽

...

Keyword(s):

Neuronal Activity ◽

Large Scale ◽

Microelectrode Array ◽

3D Structure ◽

Three Dimensional ◽

Electrode Array ◽

Extracellular Recording ◽

Brain Regions ◽

Electrode Arrays ◽

The Brain

Microelectrode array recordings of neuronal activity present significant opportunities for studying the brain with single-cell and spike-time precision. However, challenges in device manufacturing constrain dense multisite recordings to two spatial dimensions, whereas access to the three-dimensional (3D) structure of many brain regions appears to remain a challenge. To overcome this limitation, we present two novel recording modalities of silicon-based devices aimed at establishing 3D functionality. First, we fabricated a dual-side electrode array by patterning recording sites on both the front and back of an implantable microstructure. We found that the majority of single-unit spikes could not be simultaneously detected from both sides, suggesting that in addition to providing higher spatial resolution measurements than that of single-side devices, dual-side arrays also lead to increased recording yield. Second, we obtained recordings along three principal directions with a multilayer array and demonstrated 3D spike source localization within the enclosed measurement space. The large-scale integration of such dual-side and multilayer arrays is expected to provide massively parallel recording capabilities in the brain.

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