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2013-03-04Zeitschriftenartikel DOI: 10.3389/fnmol.2013.00002
Genetically encoded calcium indicators for multi-color neural activity imaging and combination with optogenetics
dc.contributor.authorAkerboom, Jasper
dc.contributor.authorCarreras Calderón, Nicole
dc.contributor.authorTian, Lin
dc.contributor.authorWabnig, Sebastian
dc.contributor.authorPrigge, Matthias
dc.contributor.authorTolö, Johan
dc.contributor.authorGordus, Andrew
dc.contributor.authorOrger, Michael B.
dc.contributor.authorSeveri, Kristen E.
dc.contributor.authorMacklin, John J.
dc.contributor.authorPatel, Ronak
dc.contributor.authorPulver, Stefan R.
dc.contributor.authorWardill, Trevor J.
dc.contributor.authorFischer, Elisabeth
dc.contributor.authorSchüler, Christina
dc.contributor.authorChen, Tsai-Wen
dc.contributor.authorSarkisyan, Karen S.
dc.contributor.authorMarvin, Jonathan S.
dc.contributor.authorBargmann, Cornelia I.
dc.contributor.authorKim, Douglas S.
dc.contributor.authorKügler, Sebastian
dc.contributor.authorLagnado, Leon
dc.contributor.authorHegemann, Peter
dc.contributor.authorGottschalk, Alexander
dc.contributor.authorSchreiter, Eric R.
dc.contributor.authorLooger, Loren L.
dc.date.accessioned2019-12-02T11:57:05Z
dc.date.available2019-12-02T11:57:05Z
dc.date.issued2013-03-04none
dc.date.updated2019-10-02T14:51:30Z
dc.identifier.urihttp://edoc.hu-berlin.de/18452/21600
dc.description.abstractGenetically encoded calcium indicators (GECIs) are powerful tools for systems neuroscience. Here we describe red, single-wavelength GECIs, “RCaMPs,” engineered from circular permutation of the thermostable red fluorescent protein mRuby. High-resolution crystal structures of mRuby, the red sensor RCaMP, and the recently published red GECI R-GECO1 give insight into the chromophore environments of the Ca2+-bound state of the sensors and the engineered protein domain interfaces of the different indicators. We characterized the biophysical properties and performance of RCaMP sensors in vitro and in vivo in Caenorhabditis elegans, Drosophila larvae, and larval zebrafish. Further, we demonstrate 2-color calcium imaging both within the same cell (registering mitochondrial and somatic [Ca2+]) and between two populations of cells: neurons and astrocytes. Finally, we perform integrated optogenetics experiments, wherein neural activation via channelrhodopsin-2 (ChR2) or a red-shifted variant, and activity imaging via RCaMP or GCaMP, are conducted simultaneously, with the ChR2/RCaMP pair providing independently addressable spectral channels. Using this paradigm, we measure calcium responses of naturalistic and ChR2-evoked muscle contractions in vivo in crawling C. elegans. We systematically compare the RCaMP sensors to R-GECO1, in terms of action potential-evoked fluorescence increases in neurons, photobleaching, and photoswitching. R-GECO1 displays higher Ca2+ affinity and larger dynamic range than RCaMP, but exhibits significant photoactivation with blue and green light, suggesting that integrated channelrhodopsin-based optogenetics using R-GECO1 may be subject to artifact. Finally, we create and test blue, cyan, and yellow variants engineered from GCaMP by rational design. This engineered set of chromatic variants facilitates new experiments in functional imaging and optogenetics.eng
dc.language.isoengnone
dc.publisherHumboldt-Universität zu Berlin
dc.rights(CC BY 3.0) Attribution 3.0 Unportedger
dc.rights.urihttp://creativecommons.org/licenses/by/3.0/
dc.subjectcalcium imagingeng
dc.subjectgenetically encoded calcium indicatoreng
dc.subjectmulti-color imagingeng
dc.subjectprotein engineeringeng
dc.subjectoptogeneticseng
dc.subject.ddc610 Medizin und Gesundheitnone
dc.titleGenetically encoded calcium indicators for multi-color neural activity imaging and combination with optogeneticsnone
dc.typearticle
dc.identifier.urnurn:nbn:de:kobv:11-110-18452/21600-9
dc.identifier.doi10.3389/fnmol.2013.00002none
dc.identifier.doihttp://dx.doi.org/10.18452/20868
dc.type.versionpublishedVersionnone
local.edoc.container-titleFrontiers in Molecular Neurosciencenone
local.edoc.pages29none
local.edoc.type-nameZeitschriftenartikel
local.edoc.institutionLebenswissenschaftliche Fakultätnone
local.edoc.container-typeperiodical
local.edoc.container-type-nameZeitschrift
local.edoc.container-publisher-nameFrontiers Media S.A.none
local.edoc.container-publisher-placeLausannenone
local.edoc.container-volume6none
dc.description.versionPeer Reviewednone
local.edoc.container-articlenumber2none
dc.identifier.eissn1662-5099

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