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2 Publications

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    02/26/18 | A robotic multidimensional directed evolution approach applied to fluorescent voltage reporters.
    Piatkevich KD, Jung EE, Straub C, Linghu C, Park D, Suk H, Hochbaum DR, Goodwin D, Pnevmatikakis E, Pak N, Kawashima T, Yang C, Rhoades JL, Shemesh O, Asano S, Yoon Y, Freifeld L, Saulnier JL, Riegler C, Engert F, Hughes T, Drobizhev M, Szabo B, Ahrens MB, Flavell SW, Sabatini BL, Boyden ES
    Nature Chemical Biology. 2018 Feb 26:. doi: 10.1038/s41589-018-0004-9

    We developed a new way to engineer complex proteins toward multidimensional specifications using a simple, yet scalable, directed evolution strategy. By robotically picking mammalian cells that were identified, under a microscope, as expressing proteins that simultaneously exhibit several specific properties, we can screen hundreds of thousands of proteins in a library in just a few hours, evaluating each along multiple performance axes. To demonstrate the power of this approach, we created a genetically encoded fluorescent voltage indicator, simultaneously optimizing its brightness and membrane localization using our microscopy-guided cell-picking strategy. We produced the high-performance opsin-based fluorescent voltage reporter Archon1 and demonstrated its utility by imaging spiking and millivolt-scale subthreshold and synaptic activity in acute mouse brain slices and in larval zebrafish in vivo. We also measured postsynaptic responses downstream of optogenetically controlled neurons in C. elegans.

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    02/24/18 | Integrative whole-brain neuroscience in larval zebrafish.
    Vanwalleghem GC, Ahrens MB, Scott EK
    Current Opinion in Neurobiology. 2018 Feb 24;50:136-145. doi: 10.1016/j.conb.2018.02.004

    Due to their small size and transparency, zebrafish larvae are amenable to a range of fluorescence microscopy techniques. With the development of sensitive genetically encoded calcium indicators, this has extended to the whole-brain imaging of neural activity with cellular resolution. This technique has been used to study brain-wide population dynamics accompanying sensory processing and sensorimotor transformations, and has spurred the development of innovative closed-loop behavioral paradigms in which stimulus-response relationships can be studied. More recently, microscopes have been developed that allow whole-brain calcium imaging in freely swimming and behaving larvae. In this review, we highlight the technologies underlying whole-brain functional imaging in zebrafish, provide examples of the sensory and motor processes that have been studied with this technique, and discuss the need to merge data from whole-brain functional imaging studies with neurochemical and anatomical information to develop holistic models of functional neural circuits.

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