Genetically Encoded Calcium Indicators for Multi-Color Neural Activity Imaging and Combination with Optogenetics
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Optical Inhibition of Larval Zebrafish Behaviour with Anion
Mohamed et al. BMC Biology (2017) 15:103 DOI 10.1186/s12915-017-0430-2 METHODOLOGY ARTICLE Open Access Optical inhibition of larval zebrafish behaviour with anion channelrhodopsins Gadisti Aisha Mohamed1, Ruey-Kuang Cheng1, Joses Ho2, Seetha Krishnan3, Farhan Mohammad4, Adam Claridge-Chang2,4 and Suresh Jesuthasan1,2,4* Abstract Background: Optical silencing of activity provides a way to test the necessity of neurons in behaviour. Two light-gated anion channels, GtACR1 and GtACR2, have recently been shown to potently inhibit activity in cultured mammalian neurons and in Drosophila. Here, we test the usefulness of these channels in larval zebrafish, using spontaneous coiling behaviour as the assay. Results: When the GtACRs were expressed in spinal neurons of embryonic zebrafish and actuated with blue or green light, spontaneous movement was inhibited. In GtACR1-expressing fish, only 3 μW/mm2 of light was sufficient to have an effect; GtACR2, which is poorly trafficked, required slightly stronger illumination. No inhibition was seen in non-expressing siblings. After light offset, the movement of GtACR-expressing fish increased, which suggested that termination of light- induced neural inhibition may lead to activation. Consistent with this, two-photon imaging of spinal neurons showed that blue light inhibited spontaneous activity in spinal neurons of GtACR1-expressing fish, and that the level of intracellular calcium increased following light offset. Conclusions: These results show that GtACR1 and GtACR2 can be used to optically inhibit neurons in larval zebrafish with high efficiency. The activity elicited at light offset needs to be taken into consideration in experimental design, although this property can provide insight into the effects of transiently stimulating a circuit. -
Imaging Thoughts - an Investigation of Neural Circuits Encoding Changes in Behavior in Zebrafish By
Imaging thoughts - an investigation of neural circuits encoding changes in behavior in zebrafish by Claire Oldfield A dissertation submitted in partial satisfaction of the requirement for the degree of Doctor of Philosophy in Neuroscience In the Graduate Division of the University of California, Berkeley Committee in charge: Professor Ehud Y. Isacoff, Chair Professor Dan E. Feldman Professor Marla Feller Professor Craig Miller Summer 2016 Abstract Imaging thoughts - an investigation of neural circuits encoding changes in behavior in zebrafish By Claire Oldfield Doctor of Philosophy in Neuroscience University of California, Berkeley Professor Ehud Isacoff, Chair Experience influences how we perceive the world, how we interact with our environment, and how we develop. In fact, almost all animals can modify their behavior as a result of experience. Psychologists have long distinguished between different forms of learning and memory, and later determined that they are encoded in distinct brain areas. Neuroscientists dating back to Santiago Ramón y Cajal suggested that learning and memory might be encoded as changes in synaptic connections between neurons, but it wasn’t until the second half of the 20th century that experimental evidence corroborated this idea. Specific firing patterns of neurons during learning results in strengthening or weakening of synapses, and morphological modifications that can lead to long lasting changes in neural circuits. The molecular mechanisms that drive these changes are remarkably conserved across vertebrates. However, understanding how synaptic plasticity is integrated at a network level remains a big challenge in neuroscience. Relatively few studies have focused on how neural circuits encode changes in behavior in a natural context. -
A Voltage-Dependent Fluorescent Indicator for Optogenetic Applications, Archaerhodopsin-3: Structure and Optical Properties
F1000Research 2017, 6:33 Last updated: 30 JUL 2021 RESEARCH NOTE A voltage-dependent fluorescent indicator for optogenetic applications, archaerhodopsin-3: Structure and optical properties from in silico modeling [version 3; peer review: 3 approved] Dmitrii M. Nikolaev1, Anton Emelyanov1, Vitaly M. Boitsov1, Maxim S Panov2, Mikhail N. Ryazantsev 2,3 1Saint-Petersburg National Research Academic University of the Russian Academy of Science, St. Petersburg, Russian Federation 2Saint-Petersburg State University, St. Petersburg, Russian Federation 3Saint-Petersburg Scientific Center of the Russian Academy of Sciences, St. Petersburg, Russian Federation v3 First published: 11 Jan 2017, 6:33 Open Peer Review https://doi.org/10.12688/f1000research.10541.1 Second version: 17 Jan 2017, 6:33 https://doi.org/10.12688/f1000research.10541.2 Reviewer Status Latest published: 15 Nov 2017, 6:33 https://doi.org/10.12688/f1000research.10541.3 Invited Reviewers 1 2 3 Abstract It was demonstrated in recent studies that some rhodopsins can be version 3 used in optogenetics as fluorescent indicators of membrane voltage. (revision) report report One of the promising candidates for these applications is 15 Nov 2017 archaerhodopsin-3. While it has already shown encouraging results, there is still a large room for improvement. One of possible directions version 2 is increasing the intensity of the protein's fluorescent signal. Rational (revision) report report report design of mutants with an improved signal is an important task, which 17 Jan 2017 requires both experimental and theoretical studies. Herein, we used a homology-based computational approach to predict the three- version 1 dimensional structure of archaerhodopsin-3, and a Quantum 11 Jan 2017 Mechanics/Molecular Mechanics (QM/MM) hybrid approach with high- level multireference ab initio methodology (SORCI+Q/AMBER) to 1. -
Optogenetic Neuronal Silencing in Drosophila During Visual Processing Alex S
www.nature.com/scientificreports OPEN Optogenetic Neuronal Silencing in Drosophila during Visual Processing Alex S. Mauss , Christian Busch & Alexander Borst Optogenetic channels and ion pumps have become indispensable tools in neuroscience to manipulate Received: 26 April 2017 neuronal activity and thus to establish synaptic connectivity and behavioral causality. Inhibitory Accepted: 6 October 2017 channels are particularly advantageous to explore signal processing in neural circuits since they permit Published: xx xx xxxx the functional removal of selected neurons on a trial-by-trial basis. However, applying these tools to study the visual system poses a considerable challenge because the illumination required for their activation usually also stimulates photoreceptors substantially, precluding the simultaneous probing of visual responses. Here, we explore the utility of the recently discovered anion channelrhodopsins GtACR1 and GtACR2 for application in the visual system of Drosophila. We frst characterized their properties using a larval crawling assay. We further obtained whole-cell recordings from cells expressing GtACR1, which mediated strong and light-sensitive photocurrents. Finally, using physiological recordings and a behavioral readout, we demonstrate that GtACR1 enables the fast and reversible silencing of genetically targeted neurons within circuits engaged in visual processing. Genetically expressed optogenetic ion channels and pumps confer light sensitivity to neurons of interest, allow- ing to control their activity on demand1,2. Such techniques have become powerful means to establish neuronal connectivity as well as causal relationships between neuronal activity and behavior. Remote control of neuronal activity by light has many advantages: it is fast, reversible, easy to parameterize and applicable in intact behaving animals. However, it poses challenges for studies in visual systems, since here endogenous light-sensing cells, the photoreceptors, are also activated by light required for optogenetic control. -
Contribution of Apical and Basal Dendrites of L2/3 Pyramidal Neurons to Orientation Encoding in Mouse V1 Jiyoung Park1,4*†, Athanasia Papoutsi3†, Ryan T
bioRxiv preprint doi: https://doi.org/10.1101/566588; this version posted March 5, 2019. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC-ND 4.0 International license. Contribution of Apical and Basal Dendrites of L2/3 Pyramidal Neurons to Orientation Encoding in Mouse V1 Jiyoung Park1,4*†, Athanasia Papoutsi3†, Ryan T. Ash2,4, Miguel A. Marin2, Panayiota Poirazi3* & Stelios M. Smirnakis4* 1Program in Structural and Computational Biology and Molecular Biophysics, Baylor College of Medicine, Houston, Texas 2Department of Neuroscience, Baylor College of Medicine, Houston, Texas 3Institute of Molecular Biology and Biotechnology (IMBB), Foundation of Research and Technology Hellas (FORTH), Vassilika Vouton, Heraklion, Crete, Greece 4Brigham and Women’s Hospital and Jamaica Plain VA Hospital, Harvard Medical School, Boston, MA †Equal contribution. * Correspondence: [email protected], [email protected], [email protected] Abstract: Pyramidal neurons integrate synaptic inputs from basal and apical dendrites to generate stimulus-specific responses. It has been proposed that feed-forward inputs to basal dendrites drive a neuron’s stimulus preference, while feedback inputs to apical dendrites sharpen selectivity. However, how a neuron’s dendritic domains relate to its functional selectivity has not been demonstrated experimentally. We performed 2-photon dendritic micro-dissection on layer- 2/3 pyramidal neurons in mouse primary visual cortex. We found that removing the apical dendritic tuft did not alter orientation-tuning. Furthermore, orientation-tuning curves were remarkably robust to the removal of basal dendrites: ablation of 2-3 basal dendrites was needed to cause a small shift in orientation preference, without significantly altering tuning width. -
A Striatal Interneuron Circuit for Continuous Target Pursuit
ARTICLE https://doi.org/10.1038/s41467-019-10716-w OPEN A striatal interneuron circuit for continuous target pursuit Namsoo Kim1, Haofang E. Li1, Ryan N. Hughes1, Glenn D.R. Watson1, David Gallegos2, Anne E. West 2, Il Hwan Kim3 & Henry H. Yin1,2 Most adaptive behaviors require precise tracking of targets in space. In pursuit behavior with a moving target, mice use distance to target to guide their own movement continuously. 1234567890():,; Here, we show that in the sensorimotor striatum, parvalbumin-positive fast-spiking inter- neurons (FSIs) can represent the distance between self and target during pursuit behavior, while striatal projection neurons (SPNs), which receive FSI projections, can represent self- velocity. FSIs are shown to regulate velocity-related SPN activity during pursuit, so that movement velocity is continuously modulated by distance to target. Moreover, bidirectional manipulation of FSI activity can selectively disrupt performance by increasing or decreasing the self-target distance. Our results reveal a key role of the FSI-SPN interneuron circuit in pursuit behavior and elucidate how this circuit implements distance to velocity transforma- tion required for the critical underlying computation. 1 Department of Psychology and Neuroscience, Duke University, Durham, NC 27708, USA. 2 Department of Neurobiology, Duke University, Durham, NC 27708, USA. 3 Department of Anatomy and Neurobiology, University of Tennessee Health and Science Center, Memphis, TN 27708, USA. Correspondence and requests for materials should be addressed to H.H.Y. (email: [email protected]) NATURE COMMUNICATIONS | (2019) 10:2715 | https://doi.org/10.1038/s41467-019-10716-w | www.nature.com/naturecommunications 1 ARTICLE NATURE COMMUNICATIONS | https://doi.org/10.1038/s41467-019-10716-w hether pursuing a prey or approaching a mate, natural pursuit performance: the worse the pursuit performance, the Wbehaviors often involve continuous tracking of targets more self-velocity lags self-target distance, as expected if distance in space. -
Wide. Fast. Deep. Recent Advances in Multi-Photon Microscopy of in Vivo Neuronal Activity
TechSights Wide. Fast. Deep. Recent Advances in Multi- Photon Microscopy of in vivo Neuronal Activity https://doi.org/10.1523/JNEUROSCI.1527-18.2019 Cite as: J. Neurosci 2019; 10.1523/JNEUROSCI.1527-18.2019 Received: 2 March 2019 Revised: 27 September 2019 Accepted: 27 September 2019 This Early Release article has been peer-reviewed and accepted, but has not been through the composition and copyediting processes. The final version may differ slightly in style or formatting and will contain links to any extended data. Alerts: Sign up at www.jneurosci.org/alerts to receive customized email alerts when the fully formatted version of this article is published. Copyright © 2019 the authors 1 Wide. Fast. Deep. Recent Advances in Multi-Photon Microscopy of in vivo Neuronal Activity. 2 Abbreviated title: Recent Advances of in vivo Multi-Photon Microscopy 3 Jérôme Lecoq1, Natalia Orlova1, Benjamin F. Grewe2,3,4 4 1 Allen Institute for Brain Science, Seattle, USA 5 2 Institute of Neuroinformatics, UZH and ETH Zurich, Switzerland 6 3 Dept. of Electrical Engineering and Information Technology, ETH Zurich, Switzerland 7 4 Faculty of Sciences, University of Zurich, Switzerland 8 9 Corresponding author: Jérôme Lecoq, [email protected] 10 Number of pages: 24 11 Number of figures: 6 12 Number of tables: 1 13 Number of words for: 14 ● abstract: 196 15 ● introduction: 474 16 ● main text: 5066 17 Conflict of interest statement: The authors declare no competing financial interests. 18 Acknowledgments: We thank Kevin Takasaki and Peter Saggau (Allen Institute for Brain Science) for providing helpful 19 comments on the manuscript; we thank Bénédicte Rossi for providing scientific illustrations. -
Han-Optogenetic-Review-ACS-2012.Pdf
Review pubs.acs.org/chemneuro In Vivo Application of Optogenetics for Neural Circuit Analysis Xue Han Biomedical Engineering Department, Boston University, Boston, Massachusetts, United States ABSTRACT: Optogenetics combines optical and genetic methods to rapidly and reversibly control neural activities or other cellular functions. Using genetic methods, specific cells or anatomical pathways can be sensitized to light through exogenous expression of microbial light activated opsin proteins. Using optical methods, opsin expressing cells can be rapidly and reversibly controlled by pulses of light of specific wavelength. With the high spatial temporal precision, optogenetic tools have enabled new ways to probe the causal role of specific cells in neural computation and behavior. Here, we overview the current state of the technology, and provide a brief introduction to the practical considerations in applying optogenetics in vivo to analyze neural circuit functions. KEYWORDS: Channelrhodopsin, archaerhodopsin, halorhodopsin, cell type specificity ptogenetics is a new field being rapidly established upon algae), and others that mediate a variety of cellular functions O the first demonstration of precise activation of neurons (for reviews on opsin structure and function, see refs 9 and 10). expressing a light-activated microbial opsin, channelrhodopsin- Most sensory rhodopsins function through recruiting intra- 2, with pulses of blue light in 2005.1 Microbial (type I) opsins cellular signaling molecules without direct ion transport are classes of monolithic light activated proteins, encoded by function. However, channelrhodopsins can mediate phototaxis small genes of under a kilobase long. Three major classes of as a light-gated cation channel at high light intensity and as a microbial opsins have been adapted to optogenetically control calcium channel at low light intensity.11 cellular functions, channelrhodopsins, halorhodopsins, and Microbial opsins share sequence homology and are archaerhodopsins (Figure 1). -
Improved Calcium Sensor Gcamp-X Overcomes the Calcium Channel Perturbations Induced by the Calmodulin in Gcamp
ARTICLE DOI: 10.1038/s41467-018-03719-6 OPEN Improved calcium sensor GCaMP-X overcomes the calcium channel perturbations induced by the calmodulin in GCaMP Yaxiong Yang 1,2,3,4, Nan Liu1,7, Yuanyuan He1, Yuxia Liu1, Lin Ge1, Linzhi Zou5, Sen Song1,4, Wei Xiong4,5 & Xiaodong Liu 1,2,3,4,5,6 2+ 1234567890():,; GCaMP, one popular type of genetically-encoded Ca indicator, has been associated with various side-effects. Here we unveil the intrinsic problem prevailing over different versions and applications, showing that GCaMP containing CaM (calmodulin) interferes with both gating and signaling of L-type calcium channels (CaV1). GCaMP acts as an impaired apoCaM 2+ 2+ and Ca /CaM, both critical to CaV1, which disrupts Ca dynamics and gene expression. We then design and implement GCaMP-X, by incorporating an extra apoCaM-binding motif, effectively protecting CaV1-dependent excitation–transcription coupling from perturbations. GCaMP-X resolves the problems of detrimental nuclear accumulation, acute and chronic Ca2+ dysregulation, and aberrant transcription signaling and cell morphogenesis, while still demonstrating excellent Ca2+-sensing characteristics partly inherited from GCaMP. In summary, CaM/CaV1 gating and signaling mechanisms are elucidated for GCaMP side- effects, while allowing the development of GCaMP-X to appropriately monitor cytosolic, submembrane or nuclear Ca2+, which is also expected to guide the future design of CaM- based molecular tools. 1 Department of Biomedical Engineering, School of Medicine, X-Lab for Transmembrane Signaling Research, Tsinghua University, Beijing 100084, China. 2 School of Biological Science and Medical Engineering, Beihang University, Beijing 100083, China. 3 Beijing Advanced Innovation Center for Biomedical Engineering, Beihang University, Beijing 102402, China. -
Optical Inhibition of Zebrafish Behavior with Anion Channelrhodopsins
bioRxiv preprint doi: https://doi.org/10.1101/158899; this version posted July 4, 2017. The copyright holder for this preprint (which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. Optical inhibition of zebrafish behavior with anion channelrhodopsins Gadisti Aisha Mohamed1, Ruey-Kuang Cheng1, Joses Ho2, Seetha Krishnan3, Farhan Mohammad4, Adam Claridge-Chang2, 4 and Suresh Jesuthasan1, 2 1. Lee Kong Chian School of Medicine, Nanyang Technological University, Singapore. 2. Institute of Molecular and Cell Biology, Singapore. 3. Graduate School for Integrative Sciences and Engineering, National University of Singapore. 4. Duke-NUS Medical School, Singapore Abstract In behavioral analysis, optical electrical silencing provides a way to test neu- ronal necessity. Two light-gated anion channels, GtACR1 and GtACR2, have recently been shown—in neuronal culture and in Drosophila—to inhibit neu- rons potently. Here, we test the usefulness of these channels in zebrafish. When the GtACRs were expressed in motor neurons and actuated with blue or green light, fish spontaneous movement was inhibited. In GtACR1-expressing fish, only 3 µW/mm2 of light was sufficient to have an effect; GtACR2, which is poorly trafficked, required stronger illumination. After light offset, GtACR- expressing fish movement increased; this suggested that termination of light- induced neural inhibition may lead to depolarization. Consistent with this, two-photon imaging of spinal neurons showed that intracellular calcium also increased following light offset. The activity elicited at light offset needs to be taken into consideration in experimental design, although this property may help provide insight into the effects of stimulating a circuit transiently. -
High-Performance Genetically Targetable Optical Neural Silencing by Light-Driven Proton Pumps
Vol 463 | 7 January 2010 | doi:10.1038/nature08652 LETTERS High-performance genetically targetable optical neural silencing by light-driven proton pumps Brian Y. Chow1,2*, Xue Han1,2*, Allison S. Dobry1,2, Xiaofeng Qian1,2, Amy S. Chuong1,2, Mingjie Li1,2, Michael A. Henninger1,2, Gabriel M. Belfort2, Yingxi Lin2, Patrick E. Monahan1,2 & Edward S. Boyden1,2 The ability to silence the activity of genetically specified neurons in screen, as did two other proton pumps, the Leptosphaeria maculans a temporally precise fashion would provide the opportunity to opsin (Mac/LR/Ops)4 and cruxrhodopsin-1 (ref. 10) (albeit less than investigate the causal role of specific cell classes in neural compu- that of Arch; Fig. 1a). All light-driven chloride pumps assessed had tations, behaviours and pathologies. Here we show that members lower screen photocurrents than these light-driven proton pumps. of the class of light-driven outward proton pumps can mediate Arch is a yellow–green light-sensitive (Fig. 1b) opsin that seems to powerful, safe, multiple-colour silencing of neural activity. The express well on the neural plasma membrane (Fig. 1c; see Sup- gene archaerhodopsin-3 (Arch)1 from Halorubrum sodomense plementary Notes on Arch expression levels and enhancing Arch enables near-100% silencing of neurons in the awake brain when membrane trafficking). Arch-mediated currents exhibited excellent virally expressed in the mouse cortex and illuminated with yellow kinetics of light-activation and post-light recovery. After illumination, light. Arch mediates currents of several hundred picoamps at low Arch currents rose with a 15–85% onset time of 8.8 6 1.8 ms light powers, and supports neural silencing currents approaching (mean 6 standard error (s.e.) reported throughout, unless otherwise 900 pA at light powers easily achievable in vivo. -
GPCR-Based Dopamine Sensors—A Detailed Guide to Inform Sensor Choice for in Vivo Imaging
International Journal of Molecular Sciences Review GPCR-Based Dopamine Sensors—A Detailed Guide to Inform Sensor Choice for In Vivo Imaging 1,2, 3,4, 4,5, Marie A. Labouesse y, Reto B. Cola y and Tommaso Patriarchi * 1 Department of Psychiatry, College of Physicians and Surgeons, Columbia University, New York, NY 10032, USA; [email protected] 2 Division of Molecular Therapeutics, New York State Psychiatric Institute, New York, NY 10032, USA 3 Anatomy and Program in Neuroscience, University of Fribourg, 1700 Fribourg, Switzerland; [email protected] 4 Institute of Pharmacology and Toxicology, University of Zurich, 8057 Zurich, Switzerland 5 Neuroscience Center Zurich, University and ETH Zurich, 8057 Zurich, Switzerland * Correspondence: [email protected]; Tel.: +41-044-635-59-21 These authors share equal co-first contribution. y Received: 10 September 2020; Accepted: 26 September 2020; Published: 28 October 2020 Abstract: Understanding how dopamine (DA) encodes behavior depends on technologies that can reliably monitor DA release in freely-behaving animals. Recently, red and green genetically encoded sensors for DA (dLight, GRAB-DA) were developed and now provide the ability to track release dynamics at a subsecond resolution, with submicromolar affinity and high molecular specificity. Combined with rapid developments in in vivo imaging, these sensors have the potential to transform the field of DA sensing and DA-based drug discovery. When implementing these tools in the laboratory, it is important to consider there is not a ‘one-size-fits-all’ sensor. Sensor properties, most importantly their affinity and dynamic range, must be carefully chosen to match local DA levels.