Voltage Imaging in Drosophila Using a Hybrid Chemical-Genetic Rhodamine Voltage Reporter
We combine a chemically-synthesized, voltage-sensitive fluorophore with a genetically encoded, self-labeling enzyme to enable voltage imaging in Drosophila melanogaster. Previously, we showed that a rhodamine voltage reporter (RhoVR) combined with the HaloTag self-labeling enzyme could be used to mo...
| Main Authors: | , , , , , , , , , , |
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| Format: | Article |
| Language: | English |
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Frontiers Media S.A.
2021-11-01
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| Series: | Frontiers in Neuroscience |
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| Online Access: | https://www.frontiersin.org/articles/10.3389/fnins.2021.754027/full |
| _version_ | 1818750687043911680 |
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| author | Molly J. Kirk Brittany R. Benlian Yifu Han Arya Gold Ashvin Ravi Parker E. Deal Rosana S. Molina Mikhail Drobizhev Dion Dickman Kristin Scott Kristin Scott Evan W. Miller Evan W. Miller Evan W. Miller |
| author_facet | Molly J. Kirk Brittany R. Benlian Yifu Han Arya Gold Ashvin Ravi Parker E. Deal Rosana S. Molina Mikhail Drobizhev Dion Dickman Kristin Scott Kristin Scott Evan W. Miller Evan W. Miller Evan W. Miller |
| author_sort | Molly J. Kirk |
| collection | DOAJ |
| description | We combine a chemically-synthesized, voltage-sensitive fluorophore with a genetically encoded, self-labeling enzyme to enable voltage imaging in Drosophila melanogaster. Previously, we showed that a rhodamine voltage reporter (RhoVR) combined with the HaloTag self-labeling enzyme could be used to monitor membrane potential changes from mammalian neurons in culture and brain slice. Here, we apply this hybrid RhoVR-Halo approach in vivo to achieve selective neuron labeling in intact fly brains. We generate a Drosophila UAS-HaloTag reporter line in which the HaloTag enzyme is expressed on the surface of cells. We validate the voltage sensitivity of this new construct in cell culture before driving expression of HaloTag in specific brain neurons in flies. We show that selective labeling of synapses, cells, and brain regions can be achieved with RhoVR-Halo in either larval neuromuscular junction (NMJ) or in whole adult brains. Finally, we validate the voltage sensitivity of RhoVR-Halo in fly tissue via dual-electrode/imaging at the NMJ, show the efficacy of this approach for measuring synaptic excitatory post-synaptic potentials (EPSPs) in muscle cells, and perform voltage imaging of carbachol-evoked depolarization and osmolarity-evoked hyperpolarization in projection neurons and in interoceptive subesophageal zone neurons in fly brain explants following in vivo labeling. We envision the turn-on response to depolarizations, fast response kinetics, and two-photon compatibility of chemical indicators, coupled with the cellular and synaptic specificity of genetically-encoded enzymes, will make RhoVR-Halo a powerful complement to neurobiological imaging in Drosophila. |
| first_indexed | 2024-12-18T04:23:38Z |
| format | Article |
| id | doaj.art-8bb0c427fcf548cfaf73e51d8ca6a5f4 |
| institution | Directory Open Access Journal |
| issn | 1662-453X |
| language | English |
| last_indexed | 2024-12-18T04:23:38Z |
| publishDate | 2021-11-01 |
| publisher | Frontiers Media S.A. |
| record_format | Article |
| series | Frontiers in Neuroscience |
| spelling | doaj.art-8bb0c427fcf548cfaf73e51d8ca6a5f42022-12-21T21:21:09ZengFrontiers Media S.A.Frontiers in Neuroscience1662-453X2021-11-011510.3389/fnins.2021.754027754027Voltage Imaging in Drosophila Using a Hybrid Chemical-Genetic Rhodamine Voltage ReporterMolly J. Kirk0Brittany R. Benlian1Yifu Han2Arya Gold3Ashvin Ravi4Parker E. Deal5Rosana S. Molina6Mikhail Drobizhev7Dion Dickman8Kristin Scott9Kristin Scott10Evan W. Miller11Evan W. Miller12Evan W. Miller13Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA, United StatesDepartment of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA, United StatesDepartment of Neurobiology, University of Southern California, Los Angeles, CA, United StatesDepartment of Chemistry, University of California, Berkeley, Berkeley, CA, United StatesDepartment of Chemistry, University of California, Berkeley, Berkeley, CA, United StatesDepartment of Chemistry, University of California, Berkeley, Berkeley, CA, United StatesDepartment of Microbiology and Cell Biology, Montana State University, Bozeman, MT, United StatesDepartment of Microbiology and Cell Biology, Montana State University, Bozeman, MT, United StatesDepartment of Neurobiology, University of Southern California, Los Angeles, CA, United StatesDepartment of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA, United StatesHelen Wills Neuroscience Institute, University of California, Berkeley, Berkeley, CA, United StatesDepartment of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA, United StatesDepartment of Chemistry, University of California, Berkeley, Berkeley, CA, United StatesHelen Wills Neuroscience Institute, University of California, Berkeley, Berkeley, CA, United StatesWe combine a chemically-synthesized, voltage-sensitive fluorophore with a genetically encoded, self-labeling enzyme to enable voltage imaging in Drosophila melanogaster. Previously, we showed that a rhodamine voltage reporter (RhoVR) combined with the HaloTag self-labeling enzyme could be used to monitor membrane potential changes from mammalian neurons in culture and brain slice. Here, we apply this hybrid RhoVR-Halo approach in vivo to achieve selective neuron labeling in intact fly brains. We generate a Drosophila UAS-HaloTag reporter line in which the HaloTag enzyme is expressed on the surface of cells. We validate the voltage sensitivity of this new construct in cell culture before driving expression of HaloTag in specific brain neurons in flies. We show that selective labeling of synapses, cells, and brain regions can be achieved with RhoVR-Halo in either larval neuromuscular junction (NMJ) or in whole adult brains. Finally, we validate the voltage sensitivity of RhoVR-Halo in fly tissue via dual-electrode/imaging at the NMJ, show the efficacy of this approach for measuring synaptic excitatory post-synaptic potentials (EPSPs) in muscle cells, and perform voltage imaging of carbachol-evoked depolarization and osmolarity-evoked hyperpolarization in projection neurons and in interoceptive subesophageal zone neurons in fly brain explants following in vivo labeling. We envision the turn-on response to depolarizations, fast response kinetics, and two-photon compatibility of chemical indicators, coupled with the cellular and synaptic specificity of genetically-encoded enzymes, will make RhoVR-Halo a powerful complement to neurobiological imaging in Drosophila.https://www.frontiersin.org/articles/10.3389/fnins.2021.754027/fullimagingfluorescencevoltageDrosophilaneuromuscular junction (NMJ) |
| spellingShingle | Molly J. Kirk Brittany R. Benlian Yifu Han Arya Gold Ashvin Ravi Parker E. Deal Rosana S. Molina Mikhail Drobizhev Dion Dickman Kristin Scott Kristin Scott Evan W. Miller Evan W. Miller Evan W. Miller Voltage Imaging in Drosophila Using a Hybrid Chemical-Genetic Rhodamine Voltage Reporter Frontiers in Neuroscience imaging fluorescence voltage Drosophila neuromuscular junction (NMJ) |
| title | Voltage Imaging in Drosophila Using a Hybrid Chemical-Genetic Rhodamine Voltage Reporter |
| title_full | Voltage Imaging in Drosophila Using a Hybrid Chemical-Genetic Rhodamine Voltage Reporter |
| title_fullStr | Voltage Imaging in Drosophila Using a Hybrid Chemical-Genetic Rhodamine Voltage Reporter |
| title_full_unstemmed | Voltage Imaging in Drosophila Using a Hybrid Chemical-Genetic Rhodamine Voltage Reporter |
| title_short | Voltage Imaging in Drosophila Using a Hybrid Chemical-Genetic Rhodamine Voltage Reporter |
| title_sort | voltage imaging in drosophila using a hybrid chemical genetic rhodamine voltage reporter |
| topic | imaging fluorescence voltage Drosophila neuromuscular junction (NMJ) |
| url | https://www.frontiersin.org/articles/10.3389/fnins.2021.754027/full |
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