scholarly journals Imaging Voltage with Microbial Rhodopsins

2021 ◽  
Vol 8 ◽  
Author(s):  
Xiao Min Zhang ◽  
Tatsushi Yokoyama ◽  
Masayuki Sakamoto
Keyword(s):  
Membrane Potential ◽  
Brain Function ◽  
Critical Parameter ◽  
Neural Circuits ◽  
Superior Performance ◽  
Neuronal Dynamics ◽  
Spatiotemporal Resolution ◽  
Invasive Approach ◽  
Electrophysiological Recordings ◽  
Genetic Specificity

Membrane potential is the critical parameter that reflects the excitability of a neuron, and it is usually measured by electrophysiological recordings with electrodes. However, this is an invasive approach that is constrained by the problems of lacking spatial resolution and genetic specificity. Recently, the development of a variety of fluorescent probes has made it possible to measure the activity of individual cells with high spatiotemporal resolution. The adaptation of this technique to image electrical activity in neurons has become an informative method to study neural circuits. Genetically encoded voltage indicators (GEVIs) can be used with superior performance to accurately target specific genetic populations and reveal neuronal dynamics on a millisecond scale. Microbial rhodopsins are commonly used as optogenetic actuators to manipulate neuronal activities and to explore the circuit mechanisms of brain function, but they also can be used as fluorescent voltage indicators. In this review, we summarize recent advances in the design and the application of rhodopsin-based GEVIs.

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 Shrinkage Classification for Overlapping Time Series: An interpretable method for mapping stimulus-differentiated evoked response

2019 ◽  
Author(s):  
Peter W. Elliott ◽  
Matthew J. Boring ◽  
Yuanning Li ◽  
R. Mark Richardson ◽  
Avniel Singh Ghuman ◽  
...  
Keyword(s):  
Time Series ◽  
Brain Function ◽  
Multivariate Time Series ◽  
Nonnegative Matrix ◽  
Brain Regions ◽  
Neural Processing ◽  
Neural Decoding ◽  
Aggregate Information ◽  
Electrophysiological Recordings ◽  
Spatio Temporal

AbstractMultivariate time series from neural electrophysiological recordings are a rich source of information about neural processing systems and require appropriate methods for proper analysis. Current methods for mapping brain function in these data using neural decoding aggregate information across space and time in limited ways, rarely incorporating spatial dependence across recording locations. We propose Shrinkage Classification for Overlapping Time Series (SCOTS), a neural decoding method that maps brain function, while accounting for spatio-temporal dependence, through interpretable dimensionality reduction and classification of multivariate neural time series. SCOTS has two components: first, overlapping clustering from sparse semi-nonnegative matrix factorization gives a data-driven aggregation of neural information across space; second, wavelet-transformed nearest shrunken centroids with sparse group lasso performs multi-class classification with selection of informative clusters and time intervals. We demonstrate use of SCOTS by applying it to human intracranial electrophysiological and MEG data collected while participants viewed visual stimuli from a range of categories. The method reveals the dynamic activation of brain regions with sensitivity to different object categories, giving insight into spatio-temporal contributions of these neural processing systems.

scholarly journals Generation of viral vectors specific to neuronal subtypes of targeted brain regions by Enhancer-Driven Gene Expression (EDGE)

2019 ◽  
Author(s):  
Rajeevkumar Raveendran Nair ◽  
Stefan Blankvoort ◽  
Maria Jose Lagartos ◽  
Cliff Kentros
Keyword(s):  
Gene Expression ◽  
Brain Function ◽  
Viral Vectors ◽  
Neural Circuits ◽  
Tissue Sample ◽  
Transgenic Animals ◽  
Regulatory Elements ◽  
Brain Regions ◽  
Transgenic Lines ◽  
The Right

SummaryUnderstanding brain function requires understanding neural circuits at the level of specificity at which they operate. While recent years have seen the development of a variety of remarkable molecular tools for the study of neural circuits, their utility is currently limited by the inability to deploy them in specific elements of native neural circuits, i.e. particular neuronal subtypes. One can obtain a degree of specificity with neuron-specific promoters, but native promoters are almost never sufficiently specific restricting this approach to transgenic animals. We recently showed that one can obtain transgenic mice with augmented anatomical specificity in targeted brain regions by identifyingcis-regulatory elements (i.e. enhancers) uniquely active in those brain regions and combining them with a heterologous promoter, an approach we call EDGE (Enhancer-Driven Gene Expression). Here we extend this strategy to the generation of viral (rAAV) vectors, showing that when combined with the right minimal promoter they largely recapitulate the specificity seen in the corresponding transgenic lines in wildtype animals, even of another species. Because active enhancers can be identified in any tissue sample, this approach promises to enable the kind of circuit-specific manipulations in any species. This should not only greatly enhance our understanding of brain function, but may one day even provide novel therapeutic avenues to correct the imbalances in neural circuits underlying many disorders of the brain.

scholarly journals Cavβ surface charged residues contribute to the regulation of neuronal calcium channels

2022 ◽  
Vol 15 (1) ◽  
Author(s):  
Alexandra Tran-Van-Minh ◽  
Michel De Waard ◽  
Norbert Weiss
Keyword(s):  
Calcium Channels ◽  
Brain Function ◽  
Binding Pocket ◽  
Surface Expression ◽  
Alanine Scanning Mutagenesis ◽  
Voltage Gated Calcium Channels ◽  
Electrophysiological Recordings ◽  
Charged Residues ◽  
Direct Binding ◽  
Fine Tune

AbstractVoltage-gated calcium channels are essential regulators of brain function where they support depolarization-induced calcium entry into neurons. They consist of a pore-forming subunit (Cavα1) that requires co-assembly with ancillary subunits to ensure proper functioning of the channel. Among these ancillary subunits, the Cavβ plays an essential role in regulating surface expression and gating of the channels. This regulation requires the direct binding of Cavβ onto Cavα1 and is mediated by the alpha interacting domain (AID) within the Cavα1 subunit and the α binding pocket (ABP) within the Cavβ subunit. However, additional interactions between Cavα1 and Cavβ have been proposed. In this study, we analyzed the importance of Cavβ3 surface charged residues in the regulation of Cav2.1 channels. Using alanine-scanning mutagenesis combined with electrophysiological recordings we identified several amino acids within the Cavβ3 subunit that contribute to the gating of the channel. These findings add to the notion that additional contacts besides the main AID/ABP interaction may occur to fine-tune the expression and properties of the channel.

scholarly journals One pair of motor neurons provokes early spontaneous motor behavior through periodic bursting in the chordate ascidian embryo

2021 ◽  
Author(s):  
Taichi Akahoshi ◽  
Kouhei Oonuma ◽  
Makoto Murakami ◽  
Takeo Horie ◽  
Takehiro G. Kusakabe ◽  
...  
Keyword(s):  
Membrane Potential ◽  
Single Cell ◽  
Motor Neurons ◽  
Neural Circuits ◽  
Motor Behavior ◽  
Cell Lineage ◽  
Muscle Cells ◽  
Single Pair ◽  
Motor Behaviors ◽  
Tail Muscle

AbstractSwimming aquatic animals exhibit spontaneous motor behaviors before the maturation of swimming locomotion. In this study, we demonstrated that a single pair of motor neurons, A10.64/MN2, which exhibits periodic bursting, is essential for early spontaneous motor behaviors in the invertebrate chordate Ciona intestinalis type A (Ciona robusta). The Ca2+ oscillation was first observed only in a pair of cells at mid tailbud II (St.22) with an 80-sec interval, which shortened to 25 sec at late tailbud II (St.24). A dissociation experiment revealed that the Ca2+ oscillation occurred in a single cell independently. The start of the Ca2+ oscillation coincided with the Ca2+ elevation in ipsilateral tail muscle cells at late tailbud I (St.23). Cell lineage tracking revealed that a pair of cells exhibiting Ca2+ oscillation corresponded to A10.64/MN2 motor neurons. Simultaneous imaging of Ca2+ and membrane potential demonstrated that the Ca2+ oscillation coupled with a burst firing of membrane potential. Interestingly, the number and frequency of tail muscle contractions initially coincided with those of the burst, but gradually came to coincide with those of spikes in the burst as the embryos developed toward late tailbud II (St.24). Finally, single-cell photoablation of A10.64/MN2 abolished early spontaneous motor behaviors until late tailbud II (St.24), suggesting that the early spontaneous motor behavior of Ciona is directly regulated by only a single pair of A10.64/MN2 motor neurons. These findings revealed that the chordate early spontaneous motor behavior was generated by a minimum motor circuit, consisting of a pair of motor neurons exhibiting periodic bursting.Significance StatementThe ascidian provides insights into formation of chordate motor neural circuits that generate early spontaneous motor behavior. Whole-body Ca2+ imaging revealed that a pair of motor neurons exhibit spontaneous Ca2+ oscillation coinciding with Ca2+ elevation in tail muscle cells at a later stage. The cell lineage of the neuron exhibiting Ca2+ oscillation was identified as a motor neuron, A10.64/MN2. In this study, we emphasize that only one pair of motor neurons firstly exhibit rhythmic activity and directly triggers early spontaneous motor behavior. These findings markedly improve our understanding of the development of chordate motor neural circuits involved in early spontaneous motor behavior.

scholarly journals Neuroimaging in Psychiatry and Neurodevelopment: why the emperor has no clothes

2019 ◽  
Vol 92 (1101) ◽  
pp. 20180910 ◽  
Author(s):  
Ashley N. Anderson ◽  
Jace B. King ◽  
Jeffrey S Anderson
Keyword(s):  
Neurodevelopmental Disorders ◽  
Brain Function ◽  
Imaging Techniques ◽  
Data Sets ◽  
Imaging Data ◽  
Spatiotemporal Resolution ◽  
New Techniques ◽  
Analysis Strategies ◽  
Neuroimaging Biomarkers ◽  
Near Future

Neuroimaging has been a dominant force in guiding research into psychiatric and neurodevelopmental disorders for decades, yet researchers have been unable to formulate sensitive or specific imaging tests for these conditions. The search for neuroimaging biomarkers has been constrained by limited reproducibility of imaging techniques, limited tools for evaluating neurochemistry, heterogeneity of patient populations not defined by brain-based phenotypes, limited exploration of temporal components of brain function, and relatively few studies evaluating developmental and longitudinal trajectories of brain function. Opportunities for development of clinically impactful imaging metrics include longer duration functional imaging data sets, new engineering approaches to mitigate suboptimal spatiotemporal resolution, improvements in image post-processing and analysis strategies, big data approaches combined with data sharing of multisite imaging samples, and new techniques that allow dynamical exploration of brain function across multiple timescales. Despite narrow clinical impact of neuroimaging methods, there is reason for optimism that imaging will contribute to diagnosis, prognosis, and treatment monitoring for psychiatric and neurodevelopmental disorders in the near future.

scholarly journals An Enhanced Deep Convolutional Model for Spatiotemporal Image Fusion

2019 ◽  
Vol 11 (24) ◽  
pp. 2898 ◽  
Author(s):  
Zhenyu Tan ◽  
Liping Di ◽  
Mingda Zhang ◽  
Liying Guo ◽  
Meiling Gao
Keyword(s):  
Image Fusion ◽  
Network Architecture ◽  
Satellite Observation ◽  
Superior Performance ◽  
Dynamic Monitoring ◽  
Future Research ◽  
Observation Data ◽  
Time Data ◽  
Spatiotemporal Resolution ◽  
Advantages And Disadvantages

Earth observation data with high spatiotemporal resolution are critical for dynamic monitoring and prediction in geoscience applications, however, due to some technique and budget limitations, it is not easy to acquire satellite images with both high spatial and high temporal resolutions. Spatiotemporal image fusion techniques provide a feasible and economical solution for generating dense-time data with high spatial resolution, pushing the limits of current satellite observation systems. Among existing various fusion algorithms, deeplearningbased models reveal a promising prospect with higher accuracy and robustness. This paper refined and improved the existing deep convolutional spatiotemporal fusion network (DCSTFN) to further boost model prediction accuracy and enhance image quality. The contributions of this paper are twofold. First, the fusion result is improved considerably with brand-new network architecture and a novel compound loss function. Experiments conducted in two different areas demonstrate these improvements by comparing them with existing algorithms. The enhanced DCSTFN model shows superior performance with higher accuracy, vision quality, and robustness. Second, the advantages and disadvantages of existing deeplearningbased spatiotemporal fusion models are comparatively discussed and a network design guide for spatiotemporal fusion is provided as a reference for future research. Those comparisons and guidelines are summarized based on numbers of actual experiments and have promising potentials to be applied for other image sources with customized spatiotemporal fusion networks.

scholarly journals Experimental Study on Blue Light Interaction with Human Keloid-Derived Fibroblasts

Biomedicines ◽  
2020 ◽  
Vol 8 (12) ◽  
pp. 573 ◽  
Author(s):  
Giada Magni ◽  
Martina Banchelli ◽  
Federica Cherchi ◽  
Elisabetta Coppi ◽  
Marco Fraccalvieri ◽  
...  
Keyword(s):  
Cell Metabolism ◽  
Human Fibroblasts ◽  
Specific Staining ◽  
Invasive Approach ◽  
Blue Led ◽  
Led Light ◽  
Electrophysiological Recordings ◽  
Outward Currents ◽  
Voltage Dependent ◽  
Physical Treatments

Keloids are an exuberant response to wound healing, characterized by an exaggerated synthesis of collagen, probably due to the increase of fibroblasts activity and to the reduction of their apoptosis rate: currently no standard treatments or pharmacological therapies are able to prevent keloid recurrence. To reach this goal, in recent years some physical treatments have been proposed, and among them the PhotoBioModulation therapy (PBM). This work analyses the effects of a blue LED light irradiation (410–430 nm, 0.69 W/cm2 power density) on human fibroblasts, isolated from both keloids and perilesional tissues. Different light doses (3.43–6.87–13.7–20.6–30.9 and 41.2 J/cm2) were tested. Biochemical assays and specific staining were used to assess cell metabolism, proliferation and viability. Micro-Raman spectroscopy was used to explore direct effects of the blue LED light on the Cytochrome C (Cyt C) oxidase. We also investigated the effects of the irradiation on ionic membrane currents by patch-clamp recordings. Our results showed that the blue LED light can modulate cell metabolism and proliferation, with a dose-dependent behavior and that these effects persist at least till 48 h after treatment. Furthermore, we demonstrated that the highest fluence value can reduce cell viability 24 h after irradiation in keloid-derived fibroblasts, while the same effect is observed 48 h after treatment in perilesional fibroblasts. Electrophysiological recordings showed that the medium dose (20.6 J/cm2) of blue LED light induces an enhancement of voltage-dependent outward currents elicited by a depolarizing ramp protocol. Overall, these data demonstrate the potentials that PBM shows as an innovative and minimally-invasive approach in the management of hypertrophic scars and keloids, in association with current treatments.

Confocal ratiometric voltage imaging of cultured human keratinocytes reveals layer-specific responses to ATP

2003 ◽  
Vol 284 (4) ◽  
pp. C944-C952 ◽  
Author(s):  
Ralf Burgstahler ◽  
Heidi Koegel ◽  
Franz Rucker ◽  
David Tracey ◽  
Peter Grafe ◽  
...  
Keyword(s):  
Membrane Potential ◽  
Resonance Energy Transfer ◽  
Resonance Energy ◽  
Human Keratinocytes ◽  
Confocal Imaging ◽  
Extracellular Signaling ◽  
Electrophysiological Recordings ◽  
Tissue Systems ◽  
Complex Culture ◽  
Voltage Sensitive Dyes

Recent evidence suggests that changes in membrane potential influence the proliferation and differentiation of keratinocytes. To further elucidate the role of changes in membrane potential for their biological fate, the electrical behavior of keratinocytes needs to be studied under complex conditions such as multilayered cultures. However, electrophysiological recordings from cells in the various layers of a complex culture would be extremely difficult. Given the high spatial resolution of confocal imaging and the availability of novel voltage-sensitive dyes, we combined these methods in an attempt to develop a viable alternative for recording membrane potentials in more complex tissue systems. As a first step, we used confocal ratiometric imaging of fluorescence resonance energy transfer (FRET)-based voltage-sensitive dyes. We then validated this approach by comparing the optically recorded voltage signals in HaCaT keratinocytes with the electrophysiological signals obtained by whole cell recordings of the same preparation. We demonstrate 1) that optical recordings allow precise multisite measurements of voltage changes evoked by the extracellular signaling molecules ATP and bradykinin and 2) that responsiveness to ATP differs in various layers of cultured keratinocytes.

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