A Light-Gated, Potassium-Selective Glutamate Receptor for the Optical Inhibition of Neuronal Firing

Department of Molecular and Cell Biology and Helen Wills Neuroscience Institute, University of California, Berkeley, California, USA.
Nature Neuroscience (Impact Factor: 14.98). 08/2010; 13(8):1027-32. DOI: 10.1038/nn.2589
Source: PubMed

ABSTRACT Genetically targeted light-activated ion channels and pumps make it possible to determine the role of specific neurons in neuronal circuits, information processing and behavior. We developed a K+-selective ionotropic glutamate receptor that reversibly inhibits neuronal activity in response to light in dissociated neurons and brain slice and also reversibly suppresses behavior in zebrafish. The receptor is a chimera of the pore region of a K+-selective bacterial glutamate receptor and the ligand-binding domain of a light-gated mammalian kainate receptor. This hyperpolarizing light-gated channel, HyLighter, is turned on by a brief light pulse at one wavelength and turned off by a pulse at a second wavelength. The control is obtained at moderate intensity. After optical activation, the photocurrent and optical silencing of activity persists in the dark for extended periods. The low light requirement and bi-stability of HyLighter represent advantages for the dissection of neural circuitry.

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    • "For example, a new kind of light controllable rapidly reversible TrpA1 ligand, optovin, was recently discovered in just this way (Kokel et al., 2013a). Several novel light activated molecules have been developed using zebrafish behavioral readouts (Szobota et al., 2007; Janovjak et al., 2010; Levitz et al., 2013). This suggests that truly novel compounds are waiting to be found, if only we use the right methods to look for them. "
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    ABSTRACT: Most neuroactive drugs were discovered through unexpected behavioral observations. Systematic behavioral screening is inefficient in most model organisms. But, automated technologies are enabling a new phase of discovery-based research in central nervous system (CNS) pharmacology. Researchers are using large-scale behavior-based chemical screens in zebrafish to discover compounds with new structures, targets, and functions. These compounds are powerful tools for understanding CNS signaling pathways. Substantial differences between human and zebrafish biology will make it difficult to translate these discoveries to clinical medicine. However, given the molecular genetic similarities between humans and zebrafish, it is likely that some of these compounds will have translational utility. We predict that the greatest new successes in CNS drug discovery will leverage many model systems, including in vitro assays, cells, rodents, and zebrafish.
    Frontiers in Pharmacology 07/2014; 5:153. DOI:10.3389/fphar.2014.00153 · 3.80 Impact Factor
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    • "VI. COMBINING OPTOGENETICS WITH TWO-PHOTON MICROSCOPY As discussed earlier, methods for guiding spatial delivery of single-photon light excitation have been used to improve the precision of optogenetic modulation [69], [131]–[138]. Light localization with higher precision can be achieved using multiphoton excitation microscopy. "
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    ABSTRACT: The brain is a large network of interconnected neurons where each cell functions as a nonlinear processing element. Unraveling the mysteries of information processing in the complex networks of the brain requires versatile neurostimulation and imaging techniques. Optogenetics is a new stimulation method which allows the activity of neurons to be modulated by light. For this purpose, the cell-types of interest are genetically targeted to produce light-sensitive proteins. Once these proteins are expressed, neural activity can be controlled by exposing the cells to light of appropriate wavelengths. Optogenetics provides a unique combination of features, including multimodal control over neural function and genetic targeting of specific cell-types. Together, these versatile features combine to a powerful experimental approach, suitable for the study of the circuitry of psychiatric and neurological disorders. The advent of optogenetics was followed by extensive research aimed to produce new lines of light-sensitive proteins and to develop new technologies: for example, to control the distribution of light inside the brain tissue or to combine optogenetics with other modalities including electrophysiology, electrocorticography, nonlinear microscopy, and functional magnetic resonance imaging. In this paper, the authors review some of the recent advances in the field of optogenetics and related technologies and provide their vision for the future of the field.
    01/2014; 7:3-30. DOI:10.1109/RBME.2013.2294796
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    • "Further, Tol2-mediated bacterial artificial chromosome (BAC) transgenesis has been applied to the generation of transgenic fish expressing Gal4 in specific neuronal types (Suster et al., 2009a,b; Bussmann and Schulte-Merker, 2011). With all of these technological advancements and optimizations, the Gal4-UAS system has been applied to a detailed visualization of neuronal morphology with a variety of fluorophores (Sagasti et al., 2005; Aramaki and Hatta, 2006; Hatta et al., 2006; Scott et al., 2007; Arrenberg et al., 2009), monitoring neural activity with fluorescent probes (Dreosti et al., 2009; Del Bene et al., 2010; Muto et al., 2011; Marvin et al., 2013) and manipulating neuronal function with neurotoxin, cytotoxin, and optogenetic probes (Szobota et al., 2007; Asakawa et al., 2008; Douglass et al., 2008; Arrenberg et al., 2009; Del Bene et al., 2010; Janovjak et al., 2010; Yokogawa et al., 2012). "
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    ABSTRACT: Bacterial artificial chromosome (BAC) transgenesis and gene/enhancer trapping are effective approaches for identification of genetically defined neuronal populations in the central nervous system (CNS). Here, we applied these techniques to zebrafish (Danio rerio) in order to obtain insights into the cellular architecture of the axial motor column in vertebrates. First, by using the BAC for the Mnx class homeodomain protein gene mnr2b/mnx2b, we established the mnGFF7 transgenic line expressing the Gal4FF transcriptional activator in a large part of the motor column. Single cell labeling of Gal4FF-expressing cells in the mnGFF7 line enabled a detailed investigation of the morphological characteristics of individual spinal motoneurons, as well as the overall organization of the motor column in a spinal segment. Secondly, from a large-scale gene trap screen, we identified transgenic lines that marked discrete subpopulations of spinal motoneurons with Gal4FF. Molecular characterization of these lines led to the identification of the ADAMTS3 gene, which encodes an evolutionarily conserved ADAMTS family of peptidases and is dynamically expressed in the ventral spinal cord. The transgenic fish established here, along with the identified gene, should facilitate an understanding of the cellular and molecular architecture of the spinal cord motor column and its connection to muscles in vertebrates.
    Frontiers in Neural Circuits 05/2013; 7:100. DOI:10.3389/fncir.2013.00100 · 2.95 Impact Factor
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