scholarly journals Multiplexed temporally focused light shaping through a GRIN lens for precise in-depth optogenetic photostimulation

2019 ◽  
Author(s):  
Nicolò Accanto ◽  
I-Wen Chen ◽  
Emiliano Ronzitti ◽  
Clément Molinier ◽  
Christophe Tourain ◽  
...  
Keyword(s):  
Neural Circuits ◽  
Three Dimensions ◽  
Optical Methods ◽  
Optical Investigation ◽  
Two Photon ◽  
Grin Lens ◽  
All Optical ◽  
Living Tissues ◽  
And Control

AbstractIn the past 10 years, the use of light has become irreplaceable for the optogenetic study and control of neurons and neural circuits. Optical techniques are however limited by scattering and can only see through a depth of few hundreds µm in living tissues. GRIN lens based micro-endoscopes represent a powerful solution to reach deeper regions. In this work we demonstrate that cutting edge optical methods for the precise photostimulation of multiple neurons in three dimensions can be performed through a GRIN lens. By spatio-temporally shaping a laser beam in the two-photon regime we project several tens of targets, spatially confined to the size of a single cell, in a volume of 150×150×400 μm3. We then apply such concept to the optogenetic stimulation of multiple neurons simultaneously in vivo in mice. Our work paves the way for an all-optical investigation of neural circuits at previously unattainable depths.

scholarly journals All-optical interrogation of neural circuits in behaving mice

2021 ◽  
Author(s):  
Lloyd E. Russell ◽  
Henry W.P. Dalgleish ◽  
Rebecca Nutbrown ◽  
Oliver Gauld ◽  
Dustin Herrmann ◽  
...  
Keyword(s):  
Neural Circuits ◽  
Neural Circuit ◽  
Barrel Cortex ◽  
Brain Area ◽  
Imaging Data ◽  
Temporal Precision ◽  
Two Photon ◽  
All Optical ◽  
The Impact

Recent advances combining two-photon calcium imaging and two-photon optogenetics with digital holography now allow us to read and write neural activity in vivo at cellular resolution with millisecond temporal precision. Such 'all-optical' techniques enable experimenters to probe the impact of functionally defined neurons on neural circuit function and behavioural output with new levels of precision. This protocol describes the experimental strategy and workflow for successful completion of typical all-optical interrogation experiments in awake, behaving head-fixed mice. We describe modular procedures for the setup and calibration of an all-optical system, the preparation of an indicator and opsin-expressing and task-performing animal, the characterization of functional and photostimulation responses and the design and implementation of an all-optical experiment. We discuss optimizations for efficiently selecting and targeting neuronal ensembles for photostimulation sequences, as well as generating photostimulation response maps from the imaging data that can be used to examine the impact of photostimulation on the local circuit. We demonstrate the utility of this strategy using all-optical experiments in three different brain areas - barrel cortex, visual cortex and hippocampus - using different experimental setups. This approach can in principle be adapted to any brain area for all-optical interrogation experiments to probe functional connectivity in neural circuits and for investigating the relationship between neural circuit activity and behaviour.

scholarly journals Precise, 3-D optogenetic control of the diameter of single arterioles

2021 ◽  
Author(s):  
Philip J. O’Herron ◽  
David A. Hartmann ◽  
Kun Xie ◽  
Prakash Kara ◽  
Andy Y. Shih
Keyword(s):  
Blood Flow ◽  
Neural Activity ◽  
Light Modulator ◽  
Spatiotemporal Resolution ◽  
Two Photon ◽  
Light Pulses ◽  
All Optical ◽  
Health And Disease ◽  
And Control

AbstractModulation of brain arteriole diameter is critical for maintenance of cerebral blood pressure and control of hyperemia during regional neural activity. However, studies of hemodynamic function in health and disease have lacked a method to control and monitor blood flow with high spatiotemporal resolution. Here, we describe a new all-optical approach to precisely control and monitor arteriolar contractility in vivo using combined two-photon optogenetics and imaging. The expression of the excitatory opsin, ReaChR, in vascular smooth muscle cells enabled rapid and repeated vasoconstriction following brief light pulses. Targeted two-photon activation of ReaCHR using a spatial light modulator (SLM) produced highly localized constrictions when targeted to individual arteries within the neocortex. We demonstrate the utility of this method for examining arteriole contractile dynamics and creating transient blood flow reductions. Additionally, we show that optogenetic constriction can offset or completely block sensory stimulus evoked vasodilation, providing a valuable tool to dissociate blood flow changes from neural activity.

scholarly journals Optogenetic strategies for high-efficiency all-optical interrogation using blue light-sensitive opsins

eLife ◽  
2021 ◽  
Vol 10 ◽  
Author(s):  
Angelo Forli ◽  
Matteo Pisoni ◽  
Yoav Printz ◽  
Ofer Yizhar ◽  
Tommaso Fellin
Keyword(s):  
Blue Light ◽  
Large Scale ◽  
High Efficiency ◽  
Average Power ◽  
Optical Methods ◽  
Neuronal Ensembles ◽  
Two Photon ◽  
All Optical ◽  
Stimulation Of

All-optical methods for imaging and manipulating brain networks with high spatial resolution are fundamental to study how neuronal ensembles drive behavior. Stimulation of neuronal ensembles using holographic techniques requires high-sensitivity actuators to avoid photodamage and heating. Moreover, two-photon-excitable opsins should be insensitive to light at wavelengths used for imaging. To achieve this goal, we developed a novel soma-targeted variant of the large-conductance blue light-sensitive opsin CoChR (stCoChR). In the mouse cortex in vivo, we combined holographic two-photon stimulation of stCoChR with an amplified laser tuned at the opsin absorption peak and imaging of the red-shifted indicator jRCaMP1a. Compared to previously characterized blue light-sensitive soma-targeted opsins in vivo, stCoChR allowed neuronal stimulation with more than 10-fold lower average power and no spectral crosstalk. The combination of stCoChR, tuned amplified laser stimulation, and red-shifted functional indicators promises to be a powerful tool for large-scale interrogation of neural networks in the intact brain.

scholarly journals Single cell in vivo brain optogenetic stimulation by two-photon excitation fluorescence transfer

2020 ◽  
Author(s):  
Lei Tong ◽  
Peng Yuan ◽  
Minggang Chen ◽  
Fuyi Chen ◽  
Joerg Bewersdorf ◽  
...  
Keyword(s):  
Single Cell ◽  
Cross Sections ◽  
Single Photon ◽  
Photon Absorption ◽  
Two Photon ◽  
Photon Excitation ◽  
All Optical ◽  
Stimulation Efficiency ◽  
And Control

AbstractOptogenetics at single-cell resolution can be achieved by two-photon stimulation; however, this requires intense or holographic illumination. We markedly improve stimulation efficiency by positioning fluorophores with high two-photon cross-sections adjacent to opsins. The two-photon-excited fluorescence matches the opsin absorbance and can stimulate opsins in a highly localized manner through efficient single-photon absorption. This indirect fluorescence transfer illumination allows experiments difficult to implement in the live brain such as all-optical neural interrogation and control of regional cerebral blood flow.

scholarly journals Simultaneous two-photon imaging and two-photon optogenetics of cortical circuits in three dimensions

eLife ◽  
2018 ◽  
Vol 7 ◽  
Author(s):  
Weijian Yang ◽  
Luis Carrillo-Reid ◽  
Yuki Bando ◽  
Darcy S Peterka ◽  
Rafael Yuste
Keyword(s):  
Neural Activity ◽  
Pyramidal Neurons ◽  
Three Dimensions ◽  
Cortical Circuits ◽  
Two Photon ◽  
Simultaneous Imaging ◽  
All Optical ◽  
Holographic Approach ◽  
Mouse Visual Cortex

The simultaneous imaging and manipulating of neural activity could enable the functional dissection of neural circuits. Here we have combined two-photon optogenetics with simultaneous volumetric two-photon calcium imaging to measure and manipulate neural activity in mouse neocortex in vivo in three-dimensions (3D) with cellular resolution. Using a hybrid holographic approach, we simultaneously photostimulate more than 80 neurons over 150 μm in depth in layer 2/3 of the mouse visual cortex, while simultaneously imaging the activity of the surrounding neurons. We validate the usefulness of the method by photoactivating in 3D selected groups of interneurons, suppressing the response of nearby pyramidal neurons to visual stimuli in awake animals. Our all-optical approach could be used as a general platform to read and write neuronal activity.

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 Motor Learning Drives Dynamic Patterns of Intermittent Myelination on Behaviorally-Activated Axons

2021 ◽  
Author(s):  
Clara M. Bacmeister ◽  
Rongchen Huang ◽  
Michael A. Thornton ◽  
Lauren Conant ◽  
Anthony R. Chavez ◽  
...  
Keyword(s):  
Motor Learning ◽  
Computational Modeling ◽  
Learning And Memory ◽  
Neuronal Activity ◽  
Neural Circuits ◽  
Cortical Layers ◽  
Nodes Of Ranvier ◽  
Two Photon ◽  
Myelin Sheaths

Myelin plasticity occurs when newly-formed and pre-existing oligodendrocytes remodel existing myelination. Recent studies show these processes occur in response to changes in neuronal activity and are required for learning and memory. However, the link between behaviorally-relevant neuronal activity and circuit-specific changes in myelination remains unknown. Using longitudinal, in vivo two-photon imaging and targeted labeling of behaviorally-activated neurons, we explore how the pattern of intermittent myelination is altered on individual cortical axons during learning of a dexterous reach task. We show that learning-induced plasticity is targeted to behaviorally-activated axons and occurs in a staged response across cortical layers. During learning, myelin sheaths retract, lengthening nodes of Ranvier. Following learning, addition of new sheaths increases the number of continuous stretches of myelination. Computational modeling suggests these changes initially slow and subsequently increase conduction speed. Thus, behaviorally-activated, circuit-specific changes to myelination may fundamentally alter how information is transferred in neural circuits during learning.

In vivo holographic photo-stimulation and two photon GCaMP6 imaging of vagus nerve axons using a GRIN lens integrated nerve cuff

2019 ◽  
Author(s):  
Gregory L. Futia ◽  
Arjun Fontaine ◽  
Samuel Littich ◽  
Connor McCullough ◽  
Diego Restrepo ◽  
...  
Keyword(s):  
Vagus Nerve ◽  
Two Photon ◽  
Grin Lens ◽  
Nerve Cuff

scholarly journals Adaptive optics two-photon endomicroscopy enables deep-brain imaging at synaptic resolution over large volumes

2020 ◽  
Vol 6 (40) ◽  
pp. eabc6521 ◽  
Author(s):  
Zhongya Qin ◽  
Congping Chen ◽  
Sicong He ◽  
Ye Wang ◽  
Kam Fai Tam ◽  
...  
Keyword(s):  
Brain Imaging ◽  
Adaptive Optics ◽  
Critical Role ◽  
Random Access ◽  
Brain Structures ◽  
Invasive Approach ◽  
Two Photon ◽  
Grin Lens ◽  
Deep Brain

Optical deep-brain imaging in vivo at high resolution has remained a great challenge over the decades. Two-photon endomicroscopy provides a minimally invasive approach to image buried brain structures, once it is integrated with a gradient refractive index (GRIN) lens embedded in the brain. However, its imaging resolution and field of view are compromised by the intrinsic aberrations of the GRIN lens. Here, we develop a two-photon endomicroscopy by adding adaptive optics based on direct wavefront sensing, which enables recovery of diffraction-limited resolution in deep-brain imaging. A new precompensation strategy plays a critical role to correct aberrations over large volumes and achieve rapid random-access multiplane imaging. We investigate the neuronal plasticity in the hippocampus, a critical deep brain structure, and reveal the relationship between the somatic and dendritic activity of pyramidal neurons.

α-[11C] methyl-L tryptophan-PET as a surrogate for interictal cerebral serotonin synthesis in migraine without aura

Cephalalgia ◽  
2013 ◽  
Vol 34 (3) ◽  
pp. 165-173 ◽  
Author(s):  
Y Sakai ◽  
M Nishikawa ◽  
M Diksic ◽  
M Aubé
Keyword(s):  
Migraine Without Aura ◽  
Surrogate Marker ◽  
Three Dimensions ◽  
Control Subjects ◽  
Positron Emission ◽  
Before And After ◽  
Regulation Mechanisms ◽  
Migraine Group ◽  
And Control

Background Alteration in central serotonin biology has been implicated in migraine, and serotonin (5-HT) agonists have been available for more than a decade in the treatment of that condition. Objectives To test this hypothesis, we studied in vivo using positron-emission tomography (PET) and α-[11C] methyl-L-tryptophan (α-[11C]MTrp) as a surrogate marker of cerebral 5-HT synthetic rate before and after administration of eletriptan in migraine and control subjects. Methods Six nonmenopausal female migraine subjects with migraine without aura (MoA) and six nonmenopausal age-matched female control subjects were scanned at baseline and after oral administration of 40 mg of eletriptan. Migraine subjects at the time of PET had to have been headache free for a minimum of three days. Images of (α-[11C]MTrp) brain trapping were colocalized with individual MRI images in three dimensions and analyzed. Results There was no difference in baseline cerebral global 5-HT synthesis between migraine and control subjects. After administration of eletriptan, there was a striking global reduction in cerebral 5-HT synthesis (K*) in the migraine group and in 22 regions of interest (ROIs). In control subjects, no significant changes were found in global cerebral 5-HT synthesis (K*) or in any of the ROIs. Conclusions These findings suggest in migraine an interictal alteration in the regulation mechanisms of cerebral 5-HT synthesis.

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