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      Optical voltage imaging in neurons: moving from technology development to practical tool

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      Nature Reviews Neuroscience

      Springer Science and Business Media LLC

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          Most cited references 56

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          Reconstruction and Simulation of Neocortical Microcircuitry.

          We present a first-draft digital reconstruction of the microcircuitry of somatosensory cortex of juvenile rat. The reconstruction uses cellular and synaptic organizing principles to algorithmically reconstruct detailed anatomy and physiology from sparse experimental data. An objective anatomical method defines a neocortical volume of 0.29 ± 0.01 mm(3) containing ~31,000 neurons, and patch-clamp studies identify 55 layer-specific morphological and 207 morpho-electrical neuron subtypes. When digitally reconstructed neurons are positioned in the volume and synapse formation is restricted to biological bouton densities and numbers of synapses per connection, their overlapping arbors form ~8 million connections with ~37 million synapses. Simulations reproduce an array of in vitro and in vivo experiments without parameter tuning. Additionally, we find a spectrum of network states with a sharp transition from synchronous to asynchronous activity, modulated by physiological mechanisms. The spectrum of network states, dynamically reconfigured around this transition, supports diverse information processing strategies.
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            Transgenic mice for intersectional targeting of neural sensors and effectors with high specificity and performance.

            An increasingly powerful approach for studying brain circuits relies on targeting genetically encoded sensors and effectors to specific cell types. However, current approaches for this are still limited in functionality and specificity. Here we utilize several intersectional strategies to generate multiple transgenic mouse lines expressing high levels of novel genetic tools with high specificity. We developed driver and double reporter mouse lines and viral vectors using the Cre/Flp and Cre/Dre double recombinase systems and established a new, retargetable genomic locus, TIGRE, which allowed the generation of a large set of Cre/tTA-dependent reporter lines expressing fluorescent proteins, genetically encoded calcium, voltage, or glutamate indicators, and optogenetic effectors, all at substantially higher levels than before. High functionality was shown in example mouse lines for GCaMP6, YCX2.60, VSFP Butterfly 1.2, and Jaws. These novel transgenic lines greatly expand the ability to monitor and manipulate neuronal activities with increased specificity.
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              Improving FRET dynamic range with bright green and red fluorescent proteins

              A variety of genetically encoded reporters use changes in fluorescence (or Förster) resonance energy transfer (FRET) to report on biochemical processes in living cells. The standard genetically encoded FRET pair consists of cyan and yellow fluorescent proteins (CFP and YFP), but many CFP-YFP reporters suffer from low FRET dynamic range, phototoxicity from the CFP excitation light, and complex photokinetic events such as reversible photobleaching and photoconversion. Here, we engineered two fluorescent proteins, Clover and mRuby2, which are the brightest green and red fluorescent proteins to date, and have the highest Förster radius of any ratiometric FRET pair yet described. Replacement of CFP and YFP in reporters of kinase activity, small GTPase activity, and transmembrane voltage significantly improves photostability, FRET dynamic range, and emission ratio changes. These improvements enhance detection of transient biochemical events such as neuronal action potential firing and RhoA activation in growth cones.
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                Author and article information

                Journal
                Nature Reviews Neuroscience
                Nat Rev Neurosci
                Springer Science and Business Media LLC
                1471-003X
                1471-0048
                November 8 2019
                Article
                10.1038/s41583-019-0231-4
                30580abd-9107-4caa-b29b-087f5cb6e66f
                © 2019

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