Filosa Alessandro

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Filosa
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Alessandro
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Optimization of a GCaMP calcium indicator for neural activity imaging

2012-10-03 , Akerboom, Jasper , Chen, Tsai-Wen , Wardill, Trevor J. , Tian, Lin , Marvin, Jonathan S. , Mutlu, Sevinc , Calderon, Nicole Carreras , Esposti, Federico , Borghuis, Bart G. , Sun, Xiaonan Richard , Gordus, Andrew , Orger, Michael B. , Portugues, Ruben , Engert, Florian , Macklin, John J. , Filosa, Alessandro , Aggarwal, Aman , Kerr, Rex A. , Takagi, Ryousuke , Kracun, Sebastian , Shigetomi, Eiji , Khakh, Baljit S. , Baier, Herwig , Lagnado, Leon , Wang, Samuel S.-H. , Bargmann, Cornelia I. , Kimmel, Bruce E. , Jayaraman, Vivek , Svoboda, Karel , Kim, Douglas S. , Schreiter, Eric R. , Looger, Loren L.

Genetically encoded calcium indicators (GECIs) are powerful tools for systems neuroscience. Recent efforts in protein engineering have significantly increased the performance of GECIs. The state-of-the art single-wavelength GECI, GCaMP3, has been deployed in a number of model organisms and can reliably detect three or more action potentials in short bursts in several systems in vivo. Through protein structure determination, targeted mutagenesis, high-throughput screening, and a battery of in vitro assays, we have increased the dynamic range of GCaMP3 by severalfold, creating a family of “GCaMP5” sensors. We tested GCaMP5s in several systems: cultured neurons and astrocytes, mouse retina, and in vivo in Caenorhabditis chemosensory neurons, Drosophila larval neuromuscular junction and adult antennal lobe, zebrafish retina and tectum, and mouse visual cortex. Signal-to-noise ratio was improved by at least 2- to 3-fold. In the visual cortex, two GCaMP5 variants detected twice as many visual stimulus-responsive cells as GCaMP3. By combining in vivo imaging with electrophysiology we show that GCaMP5 fluorescence provides a more reliable measure of neuronal activity than its predecessor GCaMP3. GCaMP5 allows more sensitive detection of neural activity in vivo and may find widespread applications for cellular imaging in general.

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