Wir danken D. Johnson und K. Doelling für die Unterstützung bei der Datenerhebung. Wir danken auch M.-H. Monfils, J. LeDoux, Y. Niv und M. Milad für Beratung über die experimentelle Protokolle. Diese Studie wurde von der James S. McDonnell Foundation und National Institutes of Health (NIH) grant R21 MH072279 (EAP), NIH Zuschüsse R37 MH038774, P50 MH058911, RO1 MH046516 und K05 MH067048 (JEL), Postdoctoral Fellowships NSERC, CIHR und AHFMR finanziert (M.-HM) und ein Fulbright und Blavatnik Auszeichnungen (DS).

<ol>
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P., Johanses, J. P., Blair, H. T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Placing+prediction+into+the+fear+circuit.">Placing prediction into the fear circuit.</a> Trends in Cognitive Neuroscience. 34, 283-292 (2011).</li><li>Herry, C., Ferraguti, F., Singewald, N., Letzkus, J. J., Ehrlich, I., Luthi, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Neural+circuits+of+fear+extinction.">Neural circuits of fear extinction.</a> European Journal of Neuroscience. 31, 599-612 (2010).</li><li>Phelps, E. A., Delgado, M. R., Nearing, K. I., LeDoux, J. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Extinction+learning+in+humans:+Role+of+the+amygdala+and+vmPFC.">Extinction learning in humans: Role of the amygdala and vmPFC.</a> Neuron. 43, 897-905 (2004).</li><li>Milad, M. R., Quirk, G. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Neurons+in+the+medial+prefrontal+cortex+signal+memory+for+fear+extinction.">Neurons in the medial prefrontal cortex signal memory for fear extinction.</a> Nature. 420, 70-74 (2002).</li><li>Schiller, D., Levy, I., Niv, Y., LeDoux, J. E., Phelps, E. 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Das Protokoll, das wir hier vorgestellten ermöglicht das Erstellen eines einfachen konditionierte Furcht Reaktion auf einen neutralen Reiz mit einem unangenehmen Ergebnis gepaart. Einen Tag später haben wir reaktiviert die Angst Speicher, um seine Rückverfestigung auslösen. Während dieser Phase induzierten Störungen in Form des Aussterbens Ausbildung. Um sicherzustellen, dass vom Aussterben Rückverfestigung betrifft verwendeten wir zwei Kontrollgruppen. Eine Gruppe unterzog Aussterben ohne Erinnerung Studie Ausl?...

extinction training during the reconsolidation window prevents recovery of fear

Angst ist maladaptive wenn speichert sie lange nach Umständen geworden sicher. Es ist daher von entscheidender Bedeutung für einen Ansatz, der anhaltend verhindert, dass die Rückkehr der Angst zu entwickeln. Pawlowschen Angst Klimaanlage Paradigmen werden üblicherweise verwendet, um eine kontrollierte, Roman Angst Vereins im Labor zu schaffen. Nach der Koppelung einen unschädlichen Stimulus (konditionierten Stimulus, CS) mit einem unangenehmen Ergebnis (unbedingter Stimulus, US) können wir eine Angstreaktion (bedingten Reaktion oder CR) die Vorlage nur den Stimulus allein 1,2 hervorzurufen. Sobald Angst erworben wird, kann es unter Verwendung vermindert Extinktion trainiert wurden, wodurch die behandelte Stimulus wiederholt ohne den aversiven Ergebnis vorgelegt, bis nicht mehr befürchten 3 ausgedrückt. Dieser hemmende Lernen schafft eine neue, sichere Darstellung für die CS, die für die Expression konkurriert mit der ursprünglichen Angst Speicher 4. Obwohl Extinktion ist wirksam bei der Hemmung Angst, ist es nicht dauerhaft. Angst kann spontanerholen sich im Laufe der Zeit. Umgang mit Stress oder die Rückkehr in den Kontext der anfänglichen Lernen kann auch dazu führen, Angst auftauchen 3,4. Unser Protokoll befasst sich mit der Vergänglichkeit des Aussterbens indem sie auf die Rückverfestigung Fenster emotionale Erinnerung in eine dauerhafte Weise zu modifizieren. Ausreichend Hinweise darauf, dass die Reaktivierung eines konsolidierten Speicher gibt sie an einen labilen Zustand, in dem der Speicher wieder störanfällig 5-9. Das Fenster der Gelegenheit wird in Kürze eröffnet nach der Reaktivierung und schließen etwa 6 Stunden später 5,11,16, obwohl dies je nach Stärke und Alter des Speichers 15. Dadurch, dass neue Informationen in die ursprüngliche Gedächtnisspur übernehmen kann dieser Speicher aktualisiert, wie es 10,11 reconsolidates werden. Studien mit nicht-menschlichen Tieren haben erfolgreich die Expression Furchtgedächtnis pharmakologischen Manipulationen durch Einführen innerhalb der re blockiertKonsolidierung Fensters, jedoch sind die meisten Wirkstoffe entweder humantoxisch oder zweideutige zeigen Wirkungen, wenn in Humanstudien 12-14 verwendet. Unser Protokoll diesen Herausforderungen, indem sie eine effektive, aber nicht-invasiv, dh Verhaltensstörungen Manipulation sicher für Mensch. Durch aufgefordert Angst Gedächtnisabruf vor dem Aussterben, wir im Wesentlichen auslösen Rückverfestigung Prozess, so dass neue Sicherheits-Informationen (dh Extinktion) eingebaut, während die Angst Speicher ist immer noch anfällig für Störungen sein. Eine kürzlich durchgeführte Studie beschäftigt dieses Verhaltens Manipulation bei Ratten erfolgreich Angst Speicher blockiert Verwendung dieser zeitliche Parameter 11. Zusätzliche Studien an Menschen haben gezeigt, dass die Einführung von neuen Informationen nach dem Abruf von konsolidierten Motor 16, episodische 17 oder 18 deklarativen Speicher führt zu Interferenz mit der ursprünglichen Gedächtnisspur 14. Wir skizzieren unter einem neuen ProProtokoll verwendet, um Angst Erholung in den Menschen zu blockieren.

Watch this Scientific Journal Video about Extinction Training During the Reconsolidation Window Prevents Recovery of Fear at JoVE.com

Extinction Training During the Reconsolidation Window Prevents Recovery of Fear

Konditionierten Furcht kann durch eine inhibitorische Prozess namens Extinktion vermindert werden, kann aber wieder auftauchen unter Bedingungen wie im Verlauf der Zeit oder durch Einwirkung von Stress. Unser Protokoll stellt eine neuartige Methode zur Verhinderung Angst Rückgewinnung durch Einführen Extinktion während der Rückverfestigung Fenster (die Umlagerung Phase eines reaktivierten Memory).

Extinction Training Während der Rückverfestigung Fenster Verhindert Recovery-of Fear

Fear is maladaptive when it persists long after circumstances have become safe. It is therefore crucial to develop an approach that persistently prevents ...

extinction-training-during-reconsolidation-window-prevents-recovery

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Neuroscience

24.9K Aufrufe.  Mt. Sinai School of Medicine. Konditionierten Furcht kann durch eine inhibitorische Prozess namens Extinktion vermindert werden, kann aber wieder auftauchen unter Bedingungen wie im Verlauf der Zeit oder durch Einwirkung von Stress. Unser Protokoll stellt eine neuartige Methode zur Verhinderung Angst Rückgewinnung durch Einführen Extinktion während der Rückverfestigung Fenster (die Umlagerung Phase eines reaktivierten Memory).

Serie: Extinction Training During the Reconsolidation Window Prevents Recovery of Fear

Human Fear Conditioning Conducted in Full Immersion 3-Dimensional Virtual Reality

Fear conditioning is a widely used paradigm in non-human animal research to investigate the neural mechanisms underlying fear and anxiety. A major challenge in conducting conditioning studies in humans is the ability to strongly manipulate or simulate the environmental contexts that are associated with conditioned emotional behaviors. In this regard, virtual reality (VR) technology is a promising tool. Yet, adapting this technology to meet experimental constraints requires special accommodations. Here we address the methodological issues involved when conducting fear conditioning in a fully immersive 6-sided VR environment and present fear conditioning data.
In the real world, traumatic events occur in complex environments that are made up of many cues, engaging all of our sensory modalities. For example, cues that form the environmental configuration include not only visual elements, but aural, olfactory, and even tactile. In rodent studies of fear conditioning animals are fully immersed in a context that is rich with novel visual, tactile and olfactory cues. However, standard laboratory tests of fear conditioning in humans are typically conducted in a nondescript room in front of a flat or 2D computer screen and do not replicate the complexity of real world experiences. On the other hand, a major limitation of clinical studies aimed at reducing (extinguishing) fear and preventing relapse in anxiety disorders is that treatment occurs after participants have acquired a fear in an uncontrolled and largely unknown context. Thus the experimenters are left without information about the duration of exposure, the true nature of the stimulus, and associated background cues in the environment1. In the absence of this information it can be difficult to truly extinguish a fear that is both cue and context-dependent. Virtual reality environments address these issues by providing the complexity of the real world, and at the same time allowing experimenters to constrain fear conditioning and extinction parameters to yield empirical data that can suggest better treatment options and/or analyze mechanistic hypotheses.
In order to test the hypothesis that fear conditioning may be richly encoded and context specific when conducted in a fully immersive environment, we developed distinct virtual reality 3-D contexts in which participants experienced fear conditioning to virtual snakes or spiders. Auditory cues co-occurred with the CS in order to further evoke orienting responses and a feeling of "presence" in subjects 2 . Skin conductance response served as the dependent measure of fear acquisition, memory retention and extinction.

Classical fear conditioning paradigm was adapted for human participants in a fully immersive virtual reality setting. Using a discrimination paradigm, conditioned fear, cue and context memory retention, and extinction was measured with skin conductance response to dynamic virtual snakes and spiders (the conditioned stimuli) in two distinct virtual contexts.

Menschliche Angstkonditionierung in Full Immersion 3-Dimensional Virtual Reality Geleitete

Fear conditioning is a widely used paradigm in non-human animal research to investigate the neural mechanisms underlying fear and anxiety. A major ...

Forschung

Neurowissenschaften

Intravital Microscopy of the Mouse Brain Microcirculation using a Closed Cranial Window

This experimental model was designed to assess the mouse pial microcirculation during acute and chronic, physiological and pathophysiological hemodynamic, inflammatory and metabolic conditions, using in vivo fluorescence microscopy. A closed cranial window is placed over the left parieto-occipital cortex of the mice. Local microcirculation is recorded in real time through the window using epi and fluorescence illumination, and measurements of vessels diameters and red blood cell (RBC) velocities are performed. RBC velocity is measured using real-time cross-correlation and/or fluorescent-labeled erythrocytes. Leukocyte and platelet adherence to pial vessels and assessment of perfusion and vascular leakage are made with the help of fluorescence-labeled markers such as Albumin-FITC and anti-CD45-TxR antibodies. Microcirculation can be repeatedly video-recorded over several days. We used for the first time the close window brain intravital microscopy to study the pial microcirculation to follow dynamic changes during the course of Plasmodium berghei ANKA infection in mice and show that expression of CM is associated with microcirculatory dysfunctions characterized by vasoconstriction, profound decrease in blood flow and eventually vascular collapse.

Intravital microscopy to follow temporal and spatial hemodynamic and inflammatory events in the pial microcirculation.

Intravitalmikroskopie von Gehirn der Maus Mikrozirkulation mit einem Closed Cranial Fenster

This experimental model was designed to assess the mouse pial microcirculation during acute and chronic, physiological and pathophysiological hemodynamic, ...

Investigating the Neural Mechanisms of Aware and Unaware Fear Memory with fMRI

Pavlovian fear conditioning is often used in combination with functional magnetic resonance imaging (fMRI) in humans to investigate the neural substrates of associative learning 1-5. In these studies, it is important to provide behavioral evidence of conditioning to verify that differences in brain activity are learning-related and correlated with human behavior. 
Fear conditioning studies often monitor autonomic responses (e.g. skin conductance response; SCR) as an index of learning and memory 6-8. In addition, other behavioral measures can provide valuable information about the learning process and/or other cognitive functions that influence conditioning. For example, the impact unconditioned stimulus (UCS) expectancies have on the expression of the conditioned response (CR) and unconditioned response (UCR) has been a topic of interest in several recent studies 9-14. SCR and UCS expectancy measures have recently been used in conjunction with fMRI to investigate the neural substrates of aware and unaware fear learning and memory processes 15. Although these cognitive processes can be evaluated to some degree following the conditioning session, post-conditioning assessments cannot measure expectations on a trial-to-trial basis and are susceptible to interference and forgetting, as well as other factors that may distort results 16,17 .
Monitoring autonomic and behavioral responses simultaneously with fMRI provides a mechanism by which the neural substrates that mediate complex relationships between cognitive processes and behavioral/autonomic responses can be assessed. However, monitoring autonomic and behavioral responses in the MRI environment poses a number of practical problems. Specifically, 1) standard behavioral and physiological monitoring equipment is constructed of ferrous material that cannot be safely used near the MRI scanner, 2) when this equipment is placed outside of the MRI scanning chamber, the cables projecting to the subject can carry RF noise that produces artifacts in brain images, 3) artifacts can be produced within the skin conductance signal by switching gradients during scanning, 4) the fMRI signal produced by the motor demands of behavioral responses may need to be distinguished from activity related to the cognitive processes of interest. Each of these issues can be resolved with modifications to the setup of physiological monitoring equipment and additional data analysis procedures. Here we present a methodology to simultaneously monitor autonomic and behavioral responses during fMRI, and demonstrate the use of these methods to investigate aware and unaware memory processes during fear conditioning.

A methodology to investigate the neural mechanisms that support aware and unaware memory processes during fear conditioning is described. This method monitors blood oxygen level dependent (BOLD) functional magnetic resonance imaging, skin conductance response, and unconditioned stimulus expectancy during Pavlovian fear conditioning to assess the neural correlates of distinct memory processes.

Die Erforschung der neuronalen Mechanismen der Aware und Unkenntnis Angst Speicher mit fMRI

Pavlovian fear conditioning is often used in combination with functional magnetic resonance imaging (fMRI) in humans to investigate the neural substrates ...

A Polished and Reinforced Thinned-skull Window for Long-term Imaging of the Mouse Brain

In vivo imaging of cortical function requires optical access to the brain without disruption of the intracranial environment. We present a method to form a polished and reinforced thinned skull (PoRTS) window in the mouse skull that spans several millimeters in diameter and is stable for months. The skull is thinned to 10 to 15 &mu;m in thickness with a hand held drill to achieve optical clarity, and is then overlaid with cyanoacrylate glue and a cover glass to: 1) provide rigidity, 2) inhibit bone regrowth and 3) reduce light scattering from irregularities on the bone surface. Since the skull is not breached, any inflammation that could affect the process being studied is greatly reduced. Imaging depths of up to 250 &mu;m below the cortical surface can be achieved using two-photon laser scanning microscopy. This window is well suited to study cerebral blood flow and cellular function in both anesthetized and awake preparations. It further offers the opportunity to manipulate cell activity using optogenetics or to disrupt blood flow in targeted vessels by irradiation of circulating photosensitizers.

We present a method to form an imaging window in the mouse skull that spans millimeters and is stable for months without inflammation of the brain. This method is well suited for longitudinal studies of blood flow, cellular dynamics, and cell/vascular structure using two-photon microscopy.

Eine polierte und verstärkte ausgedünnten Schädel Fenster für Langzeit-Imaging im Gehirn der Maus

In vivo imaging of cortical function requires optical access to the brain without disruption of the intracranial environment. We present a method to form ...

Surgical Method for Virally Mediated Gene Delivery to the Mouse Inner Ear through the Round Window Membrane

Gene therapy, used to achieve functional recovery from sensorineural deafness, promises to grant better understanding of the underlying molecular and genetic mechanisms that contribute to hearing loss. Introduction of vectors into the inner ear must be done in a way that widely distributes the agent throughout the cochlea while minimizing injury to the existing structures. This manuscript describes a post-auricular surgical approach that can be used for mouse cochlear therapy using molecular, pharmacologic, and viral delivery to mice postnatal day 10 and older via the round window membrane (RWM). This surgical approach enables rapid and direct delivery into the scala tympani while minimizing blood loss and avoiding animal mortality. This technique involves negligible or no damage to essential structures of the inner and middle ear as well as neck muscles while wholly preserving hearing. To demonstrate the efficacy of this surgical technique, the vesicular glutamate transporter 3 knockout (VGLUT3 KO) mice will be used as an example of a mouse model of congenital deafness that recovers hearing after delivery of VGLUT3 to the inner ear using an adeno-associated virus (AAV-1).

The described post-auricular surgical approach allows rapid and direct delivery into the mouse cochlear scala tympani while minimizing blood loss and animal mortality. This method can be used for cochlear therapy using molecular, pharmacologic and viral delivery to postnatal mice through the round window membrane.

Chirurgische Verfahren zum viral vermittelte Gentransfer in die Maus-Innenohr über die runde Fenstermembran

Gene therapy, used to achieve functional recovery from sensorineural deafness, promises to grant better understanding of the underlying molecular and ...

Ex Vivo Optogenetic Dissection of Fear Circuits in Brain Slices

Optogenetic approaches are now widely used to study the function of neural populations and circuits by combining targeted expression of light-activated proteins and subsequent manipulation of neural activity by light. Channelrhodopsins (ChRs) are light-gated cation-channels and when fused to a fluorescent protein their expression allows for visualization and concurrent activation of specific cell types and their axonal projections in defined areas of the brain. Via stereotactic injection of viral vectors, ChR fusion proteins can be constitutively or conditionally expressed in specific cells of a defined brain region, and their axonal projections can subsequently be studied anatomically and functionally via ex vivo optogenetic activation in brain slices. This is of particular importance when aiming to understand synaptic properties of connections that could not be addressed with conventional electrical stimulation approaches, or in identifying novel afferent and efferent connectivity that was previously poorly understood. Here, a few examples illustrate how this technique can be applied to investigate these questions to elucidating fear-related circuits in the amygdala. The amygdala is a key region for acquisition and expression of fear, and storage of fear and emotional memories. Many lines of evidence suggest that the medial prefrontal cortex (mPFC) participates in different aspects of fear acquisition and extinction, but its precise connectivity with the amygdala is just starting to be understood. First, it is shown how ex vivo optogenetic activation can be used to study aspects of synaptic communication between mPFC afferents and target cells in the basolateral amygdala (BLA). Furthermore, it is illustrated how this ex vivo optogenetic approach can be applied to assess novel connectivity patterns using a group of GABAergic neurons in the amygdala, the paracapsular intercalated cell cluster (mpITC), as an example.

Ex Vivo Optogenetic Dissection of Fear Circuits in Brain Slices

Optogenetic approaches are widely used to manipulate neural activity and assess the consequences for brain function. Here, a technique is outlined that&#160;upon in vivo expression of the optical activator Channelrhodopsin, allows for ex vivo analysis of synaptic properties of specific long range and local neural connections in fear-related circuits.

Ex Vivo optogenetische Dissection of Fear Schaltkreise in Hirnschnitten

Optogenetic approaches are now widely used to study the function of neural populations and circuits by combining targeted expression of light-activated ...

Preparation of Acute Brain Slices Using an Optimized N-Methyl-D-glucamine Protective Recovery Method

This protocol is a practical guide to the N-methyl-D-glucamine (NMDG) protective recovery method of brain slice preparation. Numerous recent studies have validated the utility of this method for enhancing neuronal preservation and overall brain slice viability. The implementation of this technique by early adopters has facilitated detailed investigations into brain function using diverse experimental applications and spanning a wide range of animal ages, brain regions, and cell types. Steps are outlined for carrying out the protective recovery brain slice technique using an optimized NMDG artificial cerebrospinal fluid (aCSF) media formulation and enhanced procedure to reliably obtain healthy brain slices for patch clamp electrophysiology. With this updated approach, a substantial improvement is observed in the speed and reliability of gigaohm seal formation during targeted patch clamp recording experiments while maintaining excellent neuronal preservation, thereby facilitating challenging experimental applications. Representative results are provided from multi-neuron patch clamp recording experiments to assay synaptic connectivity in neocortical brain slices prepared from young adult transgenic mice and mature adult human neurosurgical specimens. Furthermore, the optimized NMDG protective recovery method of brain slicing is compatible with both juvenile and adult animals, thus resolving a limitation of the original methodology. In summary, a single media formulation and brain slicing procedure can be implemented across various species and ages to achieve excellent viability and tissue preservation.

Preparation of Acute Brain Slices Using an Optimized N-Methyl-D-glucamine Protective Recovery Method

This protocol demonstrates the implementation of an optimized N-methyl-D-glucamine (NMDG) protective recovery method of brain slice preparation. A single media formulation is used to reliably obtain healthy brain slices from animals of any age and for diverse experimental applications.

Vorbereitung der akuten Gehirnscheiben mit einem optimierten N-Methyl-D-Glucamine-schützende Recovery-Methode

This protocol is a practical guide to the N-methyl-D-glucamine (NMDG) protective recovery method of brain slice preparation. Numerous recent studies have ...

A Protocol for the Administration of Real-Time fMRI Neurofeedback Training

Neurologic disorders are characterized by abnormal cellular-, molecular-, and circuit-level functions in the brain. New methods to induce and control neuroplastic processes and correct abnormal function, or even shift functions from damaged tissue to physiologically healthy brain regions, hold the potential to dramatically improve overall health. Of the current neuroplastic interventions in development, neurofeedback training (NFT) from functional Magnetic Resonance Imaging (fMRI) has the advantages of being completely non-invasive, non-pharmacologic, and spatially localized to target brain regions, as well as having no known side effects. Furthermore, NFT techniques, initially developed using fMRI, can often be translated to exercises that can be performed outside of the scanner without the aid of medical professionals or sophisticated medical equipment. In fMRI NFT, the fMRI signal is measured from specific regions of the brain, processed, and presented to the participant in real-time. Through training, self-directed mental processing techniques, that regulate this signal and its underlying neurophysiologic correlates, are developed. FMRI NFT has been used to train volitional control over a wide range of brain regions with implications for several different cognitive, behavioral, and motor systems. Additionally, fMRI NFT has shown promise in a broad range of applications such as the treatment of neurologic disorders and the augmentation of baseline human performance. In this article, we present an fMRI NFT protocol developed at our institution for modulation of both healthy and abnormal brain function, as well as examples of using the method to target both cognitive and auditory regions of the brain.

The ability to induce and/or control neural plasticity may be critical in future treatments for neurologic disorders and the recovery from brain injury. In this paper, we present a protocol on the use of neurofeedback training with functional magnetic resonance imaging to modulate human brain function.

Ein Protokoll für die Verwaltung von Real-Time-fMRT Neurofeedback Training

Neurologic disorders are characterized by abnormal cellular-, molecular-, and circuit-level functions in the brain. New methods to induce and control ...

A Microbiomechanical System for Studying Varicosity Formation and Recovery in Central Neuron Axons

Axonal varicosities are enlarged structures along the shafts of axons with a high degree of heterogeneity. They are present not only in brains with neurodegenerative diseases or injuries, but also in the normal brain. Here, we describe a newly-established micromechanical system to rapidly, reliably, and reversibly induce axonal varicosities, allowing us to understand the mechanisms governing varicosity formation and heterogeneous protein composition. This system represents a novel means to evaluate the effects of compression and shear stress on different subcellular compartments of neurons, different from other in vitro systems that mainly focus on the effect of stretching. Importantly, owing to the unique features of our system, we recently made a novel discovery showing that the application of pressurized fluid can rapidly and reversibly induce axonal varicosities through the activation of a transient receptor potential channel. Our biomechanical system can be utilized conveniently in combination with drug perfusion, live cell imaging, calcium imaging, and patch clamp recording. Therefore, this method can be adopted for studying mechanosensitive ion channels, axonal transport regulation, axonal cytoskeleton dynamics, calcium signaling, and morphological changes related to traumatic brain injury.

This protocol describes a physiologically relevant, pressurized fluid approach for rapid and reversible induction of varicosities in neurons.

Ein Microbiomechanical-System für das Studium Varizen-Bildung und Erholung in zentralen Neuronen Axone

Axonal varicosities are enlarged structures along the shafts of axons with a high degree of heterogeneity. They are present not only in brains with ...

Laminectomy and Spinal Cord Window Implantation in the Mouse

This protocol describes a method for spinal cord laminectomy and glass window implantation for in vivo imaging of the mouse spinal cord. An integrated digital vaporizer is utilized to achieve a stable plane of anesthesia at a low-flow rate of isoflurane. A single vertebral spine is removed, and a commercially available cover-glass is overlaid on a thin agarose bed. A 3D-printed plastic backplate is then affixed to the adjacent vertebral spines using tissue adhesive and dental cement. A stabilization platform is used to reduce motion artifact from respiration and heartbeat. This rapid and clamp-free method is well-suited for acute multi-photon fluorescence microscopy. Representative data are included for an application of this technique to two-photon microscopy of the spinal cord vasculature in transgenic mice expressing eGFP:Claudin-5 &#8212; a tight junction protein.

This protocol describes implantation of a glass window onto the spinal cord of a mouse to facilitate visualization by intravital microscopy.

Laminektomie und Rückenmarksfenster-Implantation in der Maus

This protocol describes a method for spinal cord laminectomy and glass window implantation for in vivo imaging of the mouse spinal cord. An integrated ...