FGCaMP7, an Improved Version of Fungi-Based Ratiometric Calcium Indicator for In Vivo Visualization of Neuronal Activity
Genetically encoded calcium indicators (GECIs) have become a widespread tool for the visualization of neuronal activity. As compared to popular GCaMP GECIs, the FGCaMP indicator benefits from calmodulin and M13-peptide from the fungi <i>Aspergillus niger</i> and <i>Aspergillus fumi...
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2020-04-01
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| Series: | International Journal of Molecular Sciences |
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| Online Access: | https://www.mdpi.com/1422-0067/21/8/3012 |
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| author | Natalia V. Barykina Vladimir P. Sotskov Anna M. Gruzdeva You Kure Wu Ruben Portugues Oksana M. Subach Elizaveta S. Chefanova Viktor V. Plusnin Olga I. Ivashkina Konstantin V. Anokhin Anna V. Vlaskina Dmitry A. Korzhenevskiy Alena Y. Nikolaeva Konstantin M. Boyko Tatiana V. Rakitina Anna M. Varizhuk Galina E. Pozmogova Fedor V. Subach |
| author_facet | Natalia V. Barykina Vladimir P. Sotskov Anna M. Gruzdeva You Kure Wu Ruben Portugues Oksana M. Subach Elizaveta S. Chefanova Viktor V. Plusnin Olga I. Ivashkina Konstantin V. Anokhin Anna V. Vlaskina Dmitry A. Korzhenevskiy Alena Y. Nikolaeva Konstantin M. Boyko Tatiana V. Rakitina Anna M. Varizhuk Galina E. Pozmogova Fedor V. Subach |
| author_sort | Natalia V. Barykina |
| collection | DOAJ |
| description | Genetically encoded calcium indicators (GECIs) have become a widespread tool for the visualization of neuronal activity. As compared to popular GCaMP GECIs, the FGCaMP indicator benefits from calmodulin and M13-peptide from the fungi <i>Aspergillus niger</i> and <i>Aspergillus fumigatus</i>, which prevent its interaction with the intracellular environment. However, FGCaMP exhibits a two-phase fluorescence behavior with the variation of calcium ion concentration, has moderate sensitivity in neurons (as compared to the GCaMP6s indicator), and has not been fully characterized in vitro and in vivo. To address these limitations, we developed an enhanced version of FGCaMP, called FGCaMP7. FGCaMP7 preserves the ratiometric phenotype of FGCaMP, with a 3.1-fold larger ratiometric dynamic range in vitro. FGCaMP7 demonstrates 2.7- and 8.7-fold greater photostability compared to mEGFP and mTagBFP2 fluorescent proteins in vitro, respectively. The ratiometric response of FGCaMP7 is 1.6- and 1.4-fold higher, compared to the intensiometric response of GCaMP6s, in non-stimulated and stimulated neuronal cultures, respectively. We reveal the inertness of FGCaMP7 to the intracellular environment of HeLa cells using its truncated version with a deleted M13-like peptide; in contrast to the similarly truncated variant of GCaMP6s. We characterize the crystal structure of the parental FGCaMP indicator. Finally, we test the in vivo performance of FGCaMP7 in mouse brain using a two-photon microscope and an NVista miniscope; and in zebrafish using two-color ratiometric confocal imaging. |
| first_indexed | 2024-03-10T20:15:53Z |
| format | Article |
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| institution | Directory Open Access Journal |
| issn | 1661-6596 1422-0067 |
| language | English |
| last_indexed | 2024-03-10T20:15:53Z |
| publishDate | 2020-04-01 |
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| series | International Journal of Molecular Sciences |
| spelling | doaj.art-dc22dc0e72834a9db05d0ed698c67f8c2023-11-19T22:36:10ZengMDPI AGInternational Journal of Molecular Sciences1661-65961422-00672020-04-01218301210.3390/ijms21083012FGCaMP7, an Improved Version of Fungi-Based Ratiometric Calcium Indicator for In Vivo Visualization of Neuronal ActivityNatalia V. Barykina0Vladimir P. Sotskov1Anna M. Gruzdeva2You Kure Wu3Ruben Portugues4Oksana M. Subach5Elizaveta S. Chefanova6Viktor V. Plusnin7Olga I. Ivashkina8Konstantin V. Anokhin9Anna V. Vlaskina10Dmitry A. Korzhenevskiy11Alena Y. Nikolaeva12Konstantin M. Boyko13Tatiana V. Rakitina14Anna M. Varizhuk15Galina E. Pozmogova16Fedor V. Subach17Laboratory for Neurobiology of Memory, P.K. Anokhin Research Institute of Normal Physiology, 125315 Moscow, RussiaInstitute for Advanced Brain Studies, M.V. Lomonosov Moscow State University, 119991 Moscow, RussiaInstitute for Advanced Brain Studies, M.V. Lomonosov Moscow State University, 119991 Moscow, RussiaSensorimotor Control Research Group, Max Planck Institute of Neurobiology, 82152 Martinsried, GermanySensorimotor Control Research Group, Max Planck Institute of Neurobiology, 82152 Martinsried, GermanyComplex of NBICS Technologies, National Research Center “Kurchatov Institute”, 123182 Moscow, RussiaDepartment of NBIC-technologies, Moscow Institute of Physics and Technology, 123182 Moscow, RussiaComplex of NBICS Technologies, National Research Center “Kurchatov Institute”, 123182 Moscow, RussiaLaboratory for Neurobiology of Memory, P.K. Anokhin Research Institute of Normal Physiology, 125315 Moscow, RussiaLaboratory for Neurobiology of Memory, P.K. Anokhin Research Institute of Normal Physiology, 125315 Moscow, RussiaComplex of NBICS Technologies, National Research Center “Kurchatov Institute”, 123182 Moscow, RussiaComplex of NBICS Technologies, National Research Center “Kurchatov Institute”, 123182 Moscow, RussiaComplex of NBICS Technologies, National Research Center “Kurchatov Institute”, 123182 Moscow, RussiaBach Institute of Biochemistry, Research Center of Biotechnology of the Russian Academy of Sciences, 119071 Moscow, RussiaComplex of NBICS Technologies, National Research Center “Kurchatov Institute”, 123182 Moscow, RussiaDepartment of Biophysics, Research and Clinical Center of Physical-Chemical Medicine of Federal Medical Biological Agency, 119435 Moscow, RussiaDepartment of Biophysics, Research and Clinical Center of Physical-Chemical Medicine of Federal Medical Biological Agency, 119435 Moscow, RussiaComplex of NBICS Technologies, National Research Center “Kurchatov Institute”, 123182 Moscow, RussiaGenetically encoded calcium indicators (GECIs) have become a widespread tool for the visualization of neuronal activity. As compared to popular GCaMP GECIs, the FGCaMP indicator benefits from calmodulin and M13-peptide from the fungi <i>Aspergillus niger</i> and <i>Aspergillus fumigatus</i>, which prevent its interaction with the intracellular environment. However, FGCaMP exhibits a two-phase fluorescence behavior with the variation of calcium ion concentration, has moderate sensitivity in neurons (as compared to the GCaMP6s indicator), and has not been fully characterized in vitro and in vivo. To address these limitations, we developed an enhanced version of FGCaMP, called FGCaMP7. FGCaMP7 preserves the ratiometric phenotype of FGCaMP, with a 3.1-fold larger ratiometric dynamic range in vitro. FGCaMP7 demonstrates 2.7- and 8.7-fold greater photostability compared to mEGFP and mTagBFP2 fluorescent proteins in vitro, respectively. The ratiometric response of FGCaMP7 is 1.6- and 1.4-fold higher, compared to the intensiometric response of GCaMP6s, in non-stimulated and stimulated neuronal cultures, respectively. We reveal the inertness of FGCaMP7 to the intracellular environment of HeLa cells using its truncated version with a deleted M13-like peptide; in contrast to the similarly truncated variant of GCaMP6s. We characterize the crystal structure of the parental FGCaMP indicator. Finally, we test the in vivo performance of FGCaMP7 in mouse brain using a two-photon microscope and an NVista miniscope; and in zebrafish using two-color ratiometric confocal imaging.https://www.mdpi.com/1422-0067/21/8/3012calcium imaginggenetically encoded calcium indicatorprotein engineeringcrystal structureFGCaMP7FGCaMP |
| spellingShingle | Natalia V. Barykina Vladimir P. Sotskov Anna M. Gruzdeva You Kure Wu Ruben Portugues Oksana M. Subach Elizaveta S. Chefanova Viktor V. Plusnin Olga I. Ivashkina Konstantin V. Anokhin Anna V. Vlaskina Dmitry A. Korzhenevskiy Alena Y. Nikolaeva Konstantin M. Boyko Tatiana V. Rakitina Anna M. Varizhuk Galina E. Pozmogova Fedor V. Subach FGCaMP7, an Improved Version of Fungi-Based Ratiometric Calcium Indicator for In Vivo Visualization of Neuronal Activity International Journal of Molecular Sciences calcium imaging genetically encoded calcium indicator protein engineering crystal structure FGCaMP7 FGCaMP |
| title | FGCaMP7, an Improved Version of Fungi-Based Ratiometric Calcium Indicator for In Vivo Visualization of Neuronal Activity |
| title_full | FGCaMP7, an Improved Version of Fungi-Based Ratiometric Calcium Indicator for In Vivo Visualization of Neuronal Activity |
| title_fullStr | FGCaMP7, an Improved Version of Fungi-Based Ratiometric Calcium Indicator for In Vivo Visualization of Neuronal Activity |
| title_full_unstemmed | FGCaMP7, an Improved Version of Fungi-Based Ratiometric Calcium Indicator for In Vivo Visualization of Neuronal Activity |
| title_short | FGCaMP7, an Improved Version of Fungi-Based Ratiometric Calcium Indicator for In Vivo Visualization of Neuronal Activity |
| title_sort | fgcamp7 an improved version of fungi based ratiometric calcium indicator for in vivo visualization of neuronal activity |
| topic | calcium imaging genetically encoded calcium indicator protein engineering crystal structure FGCaMP7 FGCaMP |
| url | https://www.mdpi.com/1422-0067/21/8/3012 |
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