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All-Optical Interrogation of Neural Circuits
The Journal of neuroscience : the official journal of the Society for Neuroscience, 2015
There have been two recent revolutionary advances in neuroscience: First, genetically encoded activity sensors have brought the goal of optical detection of single action potentials in vivo within reach. Second, optogenetic actuators now allow the activity of neurons to be controlled with millisecond precision. These revolutions have now been combined, together with advanced microscopies, to allow "all-optical" readout and manipulation of activity in neural circuits with single-spike and single-neuron precision. This is a transformational advance that will open new frontiers in neuroscience research. Harnessing the power of light in the all-optical approach requires coexpression of genetically encoded activity sensors and optogenetic probes in the same neurons, as well as the ability to simultaneously target and record the light from the selected neurons. It has recently become possible to combine sensors and optical strategies that are sufficiently sensitive and cross tal...
Temporally precise single-cell-resolution optogenetics
Nature neuroscience, 2017
Optogenetic control of individual neurons with high temporal precision within intact mammalian brain circuitry would enable powerful explorations of how neural circuits operate. Two-photon computer-generated holography enables precise sculpting of light and could in principle enable simultaneous illumination of many neurons in a network, with the requisite temporal precision to simulate accurate neural codes. We designed a high-efficacy soma-targeted opsin, finding that fusing the N-terminal 150 residues of kainate receptor subunit 2 (KA2) to the recently discovered high-photocurrent channelrhodopsin CoChR restricted expression of this opsin primarily to the cell body of mammalian cortical neurons. In combination with two-photon holographic stimulation, we found that this somatic CoChR (soCoChR) enabled photostimulation of individual cells in mouse cortical brain slices with single-cell resolution and <1-ms temporal precision. We used soCoChR to perform connectivity mapping on in...
Optogenetic investigation of neural circuits in vivo
Trends in Molecular Medicine, 2011
The recent development of light-activated optogenetic probes allows for the identification and manipulation of specific neural populations and their connections in awake animals with unprecedented spatial and temporal precision. This review describes the use of optogenetic tools to investigate neurons and neural circuits in vivo. We describe the current panel of optogenetic probes, methods of targeting these probes to specific cell types in the nervous system, and strategies of photostimulating cells in awake, behaving animals. Finally, we survey the application of optogenetic tools to studying functional neuroanatomy, behavior, and the etiology and treatment of various neurological disorders. Optogenetics The mammalian brain is composed of billions of neurons interconnected into circuits by trillions of synapses [1]. Some neural systems have been difficult to describe anatomically, such as the exquisitely complicated wiring of the cerebral cortex, while other neural systems are relatively well-characterized, such as the circuits that mediate vision, motor movements, breathing/respiration, and sleep/wake architecture. However, even these systems require functional dissection so that the relative contributions of individual cell types and their synaptic connections can be discerned. Perturbing one element in a neural circuit is especially difficult in vivo, where the complex environment of the brain in an awake, behaving animal imposes obstacles to the stimulation, inhibition, or manipulation of biochemical signaling events in specific cell types. Traditionally, cells and synapses have been manipulated using electrical, physical, pharmacological, and genetic methods (Figure 1) [2]. Although much progress has been made using these classical techniques, considerable drawbacks prevent their use in the study of neural circuits with fine spatial and temporal precision in vivo. Electrical and physical techniques are not spatially precise and can cause stimulation, inhibition, or inactivation of surrounding cells and processes. Pharmacological and genetic methods exhibit improved spatial selectivity but lack temporal resolution at the scale of single action potentials. To overcome these limitations, a new set of tools collectively referred to as "optogenetics" [3] has been developed to precisely stimulate [4-10], inhibit [11-16], or alter biochemical activity [17, 18] in specific cells or their processes with high temporal precision and rapid
Frontiers in Systems Neuroscience, 2011
Optical manipulation of neuronal activity has rapidly developed into the most powerful and widely used approach to study mechanisms related to neuronal connectivity over a range of scales. Since the early use of single site uncaging to map network connectivity, rapid technological development of light modulation techniques has added important new options, such as fast scanning photostimulation, massively parallel control of light stimuli, holographic uncaging, and two-photon stimulation techniques. Exciting new developments in optogenetics complement neurotransmitter uncaging techniques by providing cell-type specificity and in vivo usability, providing optical access to the neural substrates of behavior. Here we review the rapid evolution of methods for the optical manipulation of neuronal activity, emphasizing crucial recent developments. The age of enlightenment: evolving opportunities in brain research through optical manipulation of neuronal activity. Front. Syst. Neurosci. 5:95.
Simultaneous two-photon imaging and two-photon optogenetics of cortical circuits in three dimensions
eLife
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.
Toward the Second Generation of Optogenetic Tools
Journal of Neuroscience, 2010
This mini-symposium aims to provide an integrated perspective on recent developments in optogenetics. Research in this emerging field combines optical methods with targeted expression of genetically encoded, protein-based probes to achieve experimental manipulation and measurement of neural systems with superior temporal and spatial resolution. The essential components of the optogenetic toolbox consist of two kinds of molecular devices: actuators and reporters, which respectively enable light-mediated control or monitoring of molecular processes. The first generation of genetically encoded calcium reporters, fluorescent proteins, and neural activators has already had a great impact on neuroscience. Now, a second generation of voltage reporters, neural silencers, and functionally extended fluorescent proteins hold great promise for continuing this revolution. In this review, we will evaluate and highlight the limitations of presently available optogenic tools and discuss where these technologies and their applications are headed in the future.
A system for optically controlling neural circuits with very high spatial and temporal resolution
IEEE 13th International Conference on Bioinformatics and Bioengineering (BIBE), 2013
Optogenetics offers a powerful new approach for controlling neural circuits. It has a vast array of applications in both basic and clinical science. For basic science, it opens the door to unraveling circuit operations, since one can perturb specific circuit components with high spatial (single cell) and high temporal (millisecond) resolution. For clinical applications, it allows new kinds of selective treatments, because it provides a method to inactivate or activate specific components in a malfunctioning circuit and bring it back into a normal operating range [1-3]. To harness the power of optogenetics, though, one needs stimulating tools that work with the same high spatial and temporal resolution as the molecules themselves, the channelrhodopsins. To date, most stimulating tools require a tradeoff between spatial and temporal precision and are prohibitively expensive to integrate into a stimulating/recording setup in a laboratory or a device in a clinical setting [4, 5]. Here we describe a Digital Light Processing (DLP)-based system capable of extremely high temporal resolution (sub-millisecond), without sacrificing spatial resolution. Furthermore, it is constructed using off-theshelf components, making it feasible for a broad range of biology and bioengineering labs. Using transgenic mice that express channelrhodopsin-2 (ChR2), we demonstrate the system’s capability for stimulating channelrhodopsinexpressing neurons in tissue with single cell and submillisecond precision.