scholarly journals Three-photon fluorescence microscopy with an axially elongated Bessel focus

2017 ◽  
Author(s):  
Cristina Rodríguez ◽  
Yajie Liang ◽  
Rongwen Lu ◽  
Na Ji
Keyword(s):  
Fluorescence Microscopy ◽  
Mouse Brain ◽  
Bessel Beam ◽  
Brain Slices ◽  
Numerical Aperture ◽  
Spatiotemporal Resolution ◽  
Volumetric Imaging ◽  
Imaging Tool ◽  
Photon Fluorescence

Volumetric imaging tools that are simple to adopt, flexible, and robust, are in high demand in the field of neuroscience, where the ability to image neurons and their networks with high spatiotemporal resolution is essential. Using an axially elongated focus approximating a Bessel beam, in combination with two-photon fluorescence microscopy, has proven successful at such an endeavor. Here we demonstrate three-photon fluorescence imaging with an axially extended Bessel focus. We use an axicon-based module which allowed for the generation of Bessel foci of varying numerical aperture and axial length, and apply this volumetric imaging tool to image mouse brain slices and for in vivo imaging of the mouse brain.

scholarly journals 1700 nm optical coherence microscopy enables minimally invasive, label-free, in vivo optical biopsy deep in the mouse brain

2021 ◽  
Vol 10 (1) ◽  
Author(s):  
Jun Zhu ◽  
Hercules Rezende Freitas ◽  
Izumi Maezawa ◽  
Lee-way Jin ◽  
Vivek J. Srinivasan
Keyword(s):  
Minimally Invasive ◽  
Mouse Brain ◽  
Numerical Aperture ◽  
Optical Biopsy ◽  
Optical Coherence ◽  
Label Free ◽  
Optical Coherence Microscopy ◽  
Minimal Invasiveness ◽  
Cortical Regions

AbstractIn vivo, minimally invasive microscopy in deep cortical and sub-cortical regions of the mouse brain has been challenging. To address this challenge, we present an in vivo high numerical aperture optical coherence microscopy (OCM) approach that fully utilizes the water absorption window around 1700 nm, where ballistic attenuation in the brain is minimized. Key issues, including detector noise, excess light source noise, chromatic dispersion, and the resolution-speckle tradeoff, are analyzed and optimized. Imaging through a thinned-skull preparation that preserves intracranial space, we present volumetric imaging of cytoarchitecture and myeloarchitecture across the entire depth of the mouse neocortex, and some sub-cortical regions. In an Alzheimer’s disease model, we report that findings in superficial and deep cortical layers diverge, highlighting the importance of deep optical biopsy. Compared to other microscopic techniques, our 1700 nm OCM approach achieves a unique combination of intrinsic contrast, minimal invasiveness, and high resolution for deep brain imaging.

scholarly journals Two-Photon Microscopy Allows Imaging and Characterization of Cochlear Microvasculature In Vivo

2015 ◽  
Vol 2015 ◽  
pp. 1-8 ◽  
Author(s):  
Friedrich Ihler ◽  
Mattis Bertlich ◽  
Bernhard Weiss ◽  
Steffen Dietzel ◽  
Martin Canis
Keyword(s):  
Fluorescence Microscopy ◽  
Inner Ear ◽  
Ex Vivo ◽  
Small Scale ◽  
Published Data ◽  
Cochlear Blood Flow ◽  
Two Photon ◽  
Two Photon Fluorescence ◽  
Photon Fluorescence

Impairment of cochlear blood flow has been discussed as factor in the pathophysiology of various inner ear disorders. However, the microscopic study of cochlear microcirculation is limited due to small scale and anatomical constraints. Here, two-photon fluorescence microscopy is applied to visualize cochlear microvessels. Guinea pigs were injected with Fluorescein isothiocyanate- or Texas red-dextrane as plasma marker. Intravital microscopy was performed in four animals and explanted cochleae from four animals were studied. The vascular architecture of the cochlea was visualized up to a depth of90.0±22.7 μm. Imaging yielded a mean contrast-to-noise ratio (CNR) of3.3±1.7. Mean diameter in vivo was16.5±6.0 μm for arterioles and8.0±2.4 μm for capillaries. In explanted cochleae, the diameter of radiating arterioles and capillaries was measured with12.2±1.6 μm and6.6±1.0 μm, respectively. The difference between capillaries and arterioles was statistically significant in both experimental setups (P<0.001andP=0.022, two-way ANOVA). Measured vessel diameters in vivo and ex vivo were in agreement with published data. We conclude that two-photon fluorescence microscopy allows the investigation of cochlear microvessels and is potentially a valuable tool for inner ear research.

scholarly journals Dual-color volumetric imaging of neural activity of cortical columns

2018 ◽  
Author(s):  
Shuting Han ◽  
Weijian Yang ◽  
Rafael Yuste
Keyword(s):  
Neural Activity ◽  
High Speed ◽  
Neural Circuits ◽  
Emergent Properties ◽  
Spatiotemporal Resolution ◽  
Population Activity ◽  
Volumetric Imaging ◽  
Two Photon ◽  
Dual Color

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.

scholarly journals In vivo volumetric imaging of calcium and glutamate activity at synapses with high spatiotemporal resolution

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Wei Chen ◽  
Ryan G. Natan ◽  
Yuhan Yang ◽  
Shih-Wei Chou ◽  
Qinrong Zhang ◽  
...  
Keyword(s):  
Adaptive Optics ◽  
Temporal Resolution ◽  
Simultaneous Recording ◽  
High Temporal Resolution ◽  
Spatiotemporal Resolution ◽  
Volumetric Imaging ◽  
High Spatiotemporal Resolution ◽  
Mouse Cortex ◽  
Two Photon Fluorescence

AbstractStudying neuronal activity at synapses requires high spatiotemporal resolution. For high spatial resolution in vivo imaging at depth, adaptive optics (AO) is required to correct sample-induced aberrations. To improve temporal resolution, Bessel focus has been combined with two-photon fluorescence microscopy (2PFM) for fast volumetric imaging at subcellular lateral resolution. To achieve both high-spatial and high-temporal resolution at depth, we develop an efficient AO method that corrects the distorted wavefront of Bessel focus at the objective focal plane and recovers diffraction-limited imaging performance. Applying AO Bessel focus scanning 2PFM to volumetric imaging of zebrafish larval and mouse brains down to 500 µm depth, we demonstrate substantial improvements in the sensitivity and resolution of structural and functional measurements of synapses in vivo. This enables volumetric measurements of synaptic calcium and glutamate activity at high accuracy, including the simultaneous recording of glutamate activity of apical and basal dendritic spines in the mouse cortex.

scholarly journals Quantitative secretome analysis establishes the cell type-resolved mouse brain secretome

2020 ◽  
Author(s):  
Johanna Tüshaus ◽  
Stephan A. Müller ◽  
Evans Sioma Kataka ◽  
Jan Zaucha ◽  
Laura Sebastian Monasor ◽  
...  
Keyword(s):  
Mouse Brain ◽  
High Performance ◽  
Brain Slices ◽  
Large Cell ◽  
Cell Type ◽  
Cellular Origin ◽  
Secretome Analysis ◽  
Csf Proteins

AbstractTo understand how cells communicate in the nervous system, it is essential to define their secretome, which is challenging for primary cells because of large cell numbers being required. Here, we miniaturized secretome analysis by developing the high-performance secretome-protein-enrichment-with-click-sugars method (hiSPECS). To demonstrate its broad utility, hiSPECS was used to identify the secretory response of brain slices upon LPS-induced neuroinflammation and to establish the cell type-resolved mouse brain secretome resource using primary astrocytes, microglia, neurons and oligodendrocytes. This resource allowed mapping the cellular origin of CSF proteins and revealed that an unexpectedly high number of secreted proteins in vitro and in vivo are proteolytically-cleaved membrane protein ectodomains. Two examples are neuronally secreted ADAM22 and CD200, which we identified as substrates of the Alzheimer-linked protease BACE1. hiSPECS and the brain secretome resource can be widely exploited to systematically study protein secretion, brain function and to identify cell type-specific biomarkers for CNS diseases.

Nonlinear adaptive optics: aberration correction in three photon fluorescence microscopy for mouse brain imaging

2017 ◽  
Author(s):  
David Sinefeld ◽  
Hari P. Paudel ◽  
Tianyu Wang ◽  
Mengran Wang ◽  
Dimitre G. Ouzounov ◽  
...  
Keyword(s):  
Fluorescence Microscopy ◽  
Brain Imaging ◽  
Adaptive Optics ◽  
Mouse Brain ◽  
Aberration Correction ◽  
Photon Fluorescence

Fast, Volumetric Imaging of In Vivo Mouse Brain with Swept Confocally Aligned Planar Excitation (SCAPE) Microscopy

2015 ◽  
Author(s):  
Venkatakaushik Voleti ◽  
Matthew B. Bouchard ◽  
Clay Lacefield ◽  
Randy Bruno ◽  
Elizabeth M. C. Hillman
Keyword(s):  
Mouse Brain ◽  
Volumetric Imaging
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