Dual-color volumetric imaging of neural activity of cortical columns in vivo (Conference Presentation)

To capture the emergent properties of neural circuits, high-speed volumetric imaging of neural activity at cellular resolution is desirable. But while conventional two-photon calcium imaging is a powerful tool to study population activity in vivo, it is restrained to two-dimensional planes. Expanding it to 3D while maintaining high spatiotemporal resolution appears necessary. Here, we developed a two-photon microscope with dual-color laser excitation that can image neural activity in a 3D volume. We imaged the neuronal activity of primary visual cortex from awake mice, spanning from L2 to L5 with 10 planes, at a rate of 10 vol/sec, and demonstrated volumetric imaging of L1 long-range PFC projections and L2/3 somatas. Using this method, we map visually-evoked neuronal ensembles in 3D, finding a lack of columnar structure in orientation responses and revealing functional correlations between cortical layers which differ from trial to trial and are missed in sequential imaging. We also reveal functional interactions between presynaptic L1 axons and postsynaptic L2/3 neurons. Volumetric two-photon imaging appears an ideal method for functional connectomics of neural circuits.

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Deep brain fluorescence imaging with minimally invasive ultra-thin optical fibers

10.1101/116350 ◽

2017 ◽

Cited By ~ 1

Author(s):

Shay Ohayon ◽

Antonio M. Caravaca-Aguirre ◽

Rafael Piestun ◽

James J. DiCarlo

Keyword(s):

Optical Fibers ◽

Neural Activity ◽

Brain Regions ◽

Volumetric Imaging ◽

Non Invasive ◽

Cellular Resolution ◽

Calcium Indicator ◽

Acquisition Speed ◽

Resolution Imaging

AbstractA major open challenge in neuroscience is the ability to measure and perturb neural activity in vivo from well-defined neural sub-populations at cellular resolution anywhere in the brain. However, limitations posed by scattering and absorption prohibit non-invasive (surface) multiphoton approaches1,2 for deep (>2mm) structures, while Gradient Refreactive Index (GRIN) endoscopes2–4 are thick and cause significant damage upon insertion. Here, we demonstrate a novel microendoscope to image neural activity at arbitrary depths via an ultrathin multimode optical fiber (MMF) probe that is 5-10X thinner than commercially available microendoscopes. We demonstrate micron-scale resolution, multispectral and volumetric imaging. In contrast to previous approaches1,5–8 we show that this method has an improved acquisition speed that is sufficient to capture rapid neuronal dynamics in-vivo in rodents expressing a genetically encoded calcium indicator. Our results emphasize the potential of this technology in neuroscience applications and open up possibilities for cellular resolution imaging in previously unreachable brain regions.

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3D visualization of macromolecule synthesis

eLife ◽

10.7554/elife.60354 ◽

2020 ◽

Vol 9 ◽

Author(s):

Timothy J Duerr ◽

Ester Comellas ◽

Eun Kyung Jeon ◽

Johanna E Farkas ◽

Marylou Joetzjer ◽

...

Keyword(s):

3D Visualization ◽

Fluorescent Microscopy ◽

Three Dimensional ◽

Light Sheet ◽

Macromolecular Synthesis ◽

Volumetric Imaging ◽

Processing Pipeline ◽

Powerful Means

Measuring nascent macromolecular synthesis in vivo is key to understanding how cells and tissues progress through development and respond to external cues. Here we perform in vivo injection of alkyne- or azide-modified analogs of thymidine, uridine, methionine, and glucosamine to label nascent synthesis of DNA, RNA, protein, and glycosylation. Three-dimensional volumetric imaging of nascent macromolecule synthesis was performed in axolotl salamander tissue using whole-mount click chemistry-based fluorescent staining followed by light sheet fluorescent microscopy. We also developed an image processing pipeline for segmentation and classification of morphological regions of interest and individual cells, and we apply this pipeline to the regenerating humerus. We demonstrate our approach is sensitive to biological perturbations by measuring changes in DNA synthesis after limb denervation. This method provides a powerful means to quantitatively interrogate macromolecule synthesis in heterogenous tissues at the organ, cellular, and molecular levels of organization.

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Large field-of-view incoherent volumetric imaging in living human retina by confocal oblique scanning laser ophthalmoscopy

10.1101/2021.08.05.455286 ◽

2021 ◽

Author(s):

Wenjun Shao ◽

Ji Yi

Keyword(s):

Clinical Care ◽

Retinal Imaging ◽

Field Of View ◽

Scanning Laser ◽

Large Field ◽

Scanning Laser Ophthalmoscopy ◽

Human Retina ◽

Label Free ◽

Volumetric Imaging

Three-dimensional (3D) volumetric imaging of the human retina is instrumental to monitor and diagnose blinding conditions. Although coherent retinal imaging is well established by optical coherence tomography, it is still a large void for incoherent volumetric imaging in the human retina. Here, we report confocal oblique scanning laser ophthalmoscopy (CoSLO), to fill that void and harness incoherent optical contrast in 3D. CoSLO uses oblique scanning laser and remote focusing to acquire depth signal in parallel, avoid the lengthy z-stacking, and image a large field of view (FOV). In addition, confocal gating is introduced by a linear sensor array to improve the contrast and resolution. For the first time, we achieved incoherent 3D human retinal imaging with >20° viewing angle within only 5 seconds. The depth resolution is ~45 microns in vivo. We demonstrated label-free incoherent contrast by CoSLO, revealing unique features in the retina. CoSLO will be an important technique for clinical care of retinal conditions and fundamental vision science, by offering unique volumetric incoherent contrasts.

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Ex Vivo Brain Preparation to Analyze Vocal Pathways of Xenopus Frogs

Cold Spring Harbor Protocols ◽

10.1101/pdb.prot106872 ◽

2021 ◽

Vol 2021 (9) ◽

pp. pdb.prot106872

Author(s):

Ayako Yamaguchi

Keyword(s):

Neural Activity ◽

Ex Vivo ◽

Neural Circuitry ◽

Neural Basis ◽

Vocal Production ◽

Basic Principles ◽

Electrophysiological Recordings ◽

The Brain

Understanding the neural basis of behavior is a challenging task for technical reasons. Most methods of recording neural activity require animals to be immobilized, but neural activity associated with most behavior cannot be recorded from an anesthetized, immobilized animal. Using amphibians, however, there has been some success in developing in vitro brain preparations that can be used for electrophysiological and anatomical studies. Here, we describe an ex vivo frog brain preparation from which fictive vocalizations (the neural activity that would have produced vocalizations had the brain been attached to the muscle) can be elicited repeatedly. When serotonin is applied to the isolated brains of male and female African clawed frogs, Xenopus laevis, laryngeal nerve activity that is a facsimile of those that underlie sex-specific vocalizations in vivo can be readily recorded. Recently, this preparation was successfully used in other species within the genus including Xenopus tropicalis and Xenopus victorianus. This preparation allows a variety of techniques to be applied including extracellular and intracellular electrophysiological recordings and calcium imaging during vocal production, surgical and pharmacological manipulation of neurons to evaluate their impact on motor output, and tract tracing of the neural circuitry. Thus, the preparation is a powerful tool with which to understand the basic principles that govern the production of coherent and robust motor programs in vertebrates.

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Artifact‐Free 2D Mapping of Neural Activity In Vivo through Transparent Gold Nanonetwork Array

Advanced Functional Materials ◽

10.1002/adfm.202000896 ◽

2020 ◽

Vol 30 (34) ◽

pp. 2000896

Author(s):

Ji‐Won Seo ◽

Kiup Kim ◽

Ki‐Won Seo ◽

Mi Kyung Kim ◽

Sohyeon Jeong ◽

...

Keyword(s):

Neural Activity

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Volumetric imaging of myelin in vivo using 3D inversion recovery‐prepared ultrashort echo time cones magnetic resonance imaging

NMR in Biomedicine ◽

10.1002/nbm.4326 ◽

2020 ◽

Vol 33 (10) ◽

Author(s):

Ya‐Jun Ma ◽

Adam C. Searleman ◽

Hyungseok Jang ◽

Shu‐Juan Fan ◽

Jonathan Wong ◽

...

Keyword(s):

Magnetic Resonance Imaging ◽

Magnetic Resonance ◽

Inversion Recovery ◽

Resonance Imaging ◽

3D Inversion ◽

Ultrashort Echo Time ◽

Volumetric Imaging ◽

Echo Time

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Forebrain NgR1 Overexpression Impairs DA Release Suggesting Synergy of Local and Global Synaptic Plasticity Mechanisms

Frontiers in Synaptic Neuroscience ◽

10.3389/fnsyn.2020.545854 ◽

2020 ◽

Vol 12 ◽

Author(s):

Emma Arvidsson ◽

Sarolta Gabulya ◽

Alvin Tore Brodin ◽

Tobias Erik Karlsson ◽

Lars Olson

Keyword(s):

Synaptic Plasticity ◽

Potassium Chloride ◽

Neural Activity ◽

Dorsal Striatum ◽

Inverse Correlation ◽

Long Term Memory ◽

The Status ◽

Global Systems ◽

Da Release

Structural synaptic reorganizations needed to permanently embed novel memories in the brain involve complex plasticity-enhancing and plasticity-inhibiting systems. Increased neural activity is linked to rapid downregulation of Nogo receptor 1 (NgR1), needed to allow local structural synaptic plasticity. This local regulation of plasticity is thought to be moderated by global systems, such as the ascending cholinergic and monoaminergic systems, adding significance to locally increased neural activity. Here we address the reverse possibility that the global systems may also be influenced by the status of local plasticity. Using NgR1-overexpressing mice, with impaired plasticity and long-term memory, we measured the ability to release dopamine (DA), implicated in regulating plasticity and memory. In vivo chronoamperometric recording with high temporal and spatial resolution revealed severe impairment of potassium chloride (KCl)-induced increase of extracellular DA in the dorsal striatum of mice overexpressing NgR1 in forebrain neurons. A similar, but lesser, impairment of DA release was seen following amphetamine delivery. In contrast, potassium chloride-evoked DA release in NgR1 knockout (KO) mice led to increased levels of extracellular DA. That NgR1 can impair DA signaling, thereby further dampening synaptic plasticity, suggests a new role for NgR1 signaling, acting in synergy with DA signaling to control synaptic plasticity.Significance Statement:The inverse correlation between local NgR1 levels and magnitude of KCl-inducible amounts of DA release in the striatum reinforces the rule of NgR1 as a regulator of structural synaptic plasticity and suggests synergy between local and global plasticity regulating systems.

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A high-speed, bright, red fluorescent voltage sensor to detect neural activity

Scientific Reports ◽

10.1038/s41598-019-52370-8 ◽

2019 ◽

Vol 9 (1) ◽

Cited By ~ 5

Author(s):

Connor Beck ◽

Yiyang Gong

Keyword(s):

Blue Light ◽

Neural Activity ◽

High Speed ◽

Fluorescent Protein ◽

Resonance Energy ◽

Fluorescent Sensors ◽

Fast Kinetics ◽

Spectral Separation ◽

Green Fluorescent

Abstract Genetically encoded voltage indicators (GEVIs) have emerged as a technology to optically record neural activity with genetic specificity and millisecond-scale temporal resolution using fluorescence microscopy. GEVIs have demonstrated ultra-fast kinetics and high spike detection fidelity in vivo, but existing red-fluorescent voltage indicators fall short of the response and brightness achieved by green fluorescent protein-based sensors. Furthermore, red-fluorescent GEVIs suffer from incomplete spectral separation from green sensors and blue-light-activated optogenetic actuators. We have developed Ace-mScarlet, a red fluorescent GEVI that fuses Ace2N, a voltage-sensitive inhibitory rhodopsin, with mScarlet, a bright red fluorescent protein (FP). Through fluorescence resonance energy transfer (FRET), our sensor detects changes in membrane voltage with high sensitivity and brightness and has kinetics comparable to the fastest green fluorescent sensors. Ace-mScarlet’s red-shifted absorption and emission spectra facilitate virtually complete spectral separation when used in combination with green-fluorescent sensors or with blue-light-sensitive sensors and rhodopsins. This spectral separation enables both simultaneous imaging in two separate wavelength channels and high-fidelity voltage recordings during simultaneous optogenetic perturbation.

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