Whole-brain in situ mapping of neuronal activation in Drosophila during social behaviors and optogenetic stimulation
Monitoring neuronal activity at single-cell resolution in freely moving Drosophila engaged in social behaviors is challenging because of their small size and lack of transparency. Extant methods, such as Flyception, are highly invasive. Whole-brain calcium imaging in head-fixed, walking flies is fea...
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eLife Sciences Publications Ltd
2024-11-01
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| Online Access: | https://elifesciences.org/articles/92380 |
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| author | Kiichi Watanabe Hui Chiu David J Anderson |
| author_facet | Kiichi Watanabe Hui Chiu David J Anderson |
| author_sort | Kiichi Watanabe |
| collection | DOAJ |
| description | Monitoring neuronal activity at single-cell resolution in freely moving Drosophila engaged in social behaviors is challenging because of their small size and lack of transparency. Extant methods, such as Flyception, are highly invasive. Whole-brain calcium imaging in head-fixed, walking flies is feasible but the animals cannot perform the consummatory phases of social behaviors like aggression or mating under these conditions. This has left open the fundamental question of whether neurons identified as functionally important for such behaviors using loss- or gain-of-function screens are actually active during the natural performance of such behaviors, and if so during which phase(s). Here, we perform brain-wide mapping of active cells expressing the Immediate Early Gene hr38 using a high-sensitivity/low background fluorescence in situ hybridization (FISH) amplification method called HCR-3.0. Using double-labeling for hr38 mRNA and for GFP, we describe the activity of several classes of aggression-promoting neurons during courtship and aggression, including P1a cells, an intensively studied population of male-specific interneurons. Using HI-FISH in combination with optogenetic activation of aggression-promoting neurons (opto-HI-FISH), we identify candidate downstream functional targets of these cells in a brain-wide, unbiased manner. Finally, we compare the activity of P1a neurons during sequential performance of courtship and aggression, using intronic vs. exonic hr38 probes to differentiate newly synthesized nuclear transcripts from cytoplasmic transcripts synthesized at an earlier time. These data provide evidence suggesting that different subsets of P1a neurons may be active during courtship vs. aggression. HI-FISH and associated methods may help to fill an important lacuna in the armamentarium of tools for neural circuit analysis in Drosophila. |
| format | Article |
| id | doaj-art-dbb98fedca7d40399ad6b824be190de6 |
| institution | Kabale University |
| issn | 2050-084X |
| language | English |
| publishDate | 2024-11-01 |
| publisher | eLife Sciences Publications Ltd |
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| spelling | doaj-art-dbb98fedca7d40399ad6b824be190de62024-11-28T16:21:02ZengeLife Sciences Publications LtdeLife2050-084X2024-11-011210.7554/eLife.92380Whole-brain in situ mapping of neuronal activation in Drosophila during social behaviors and optogenetic stimulationKiichi Watanabe0https://orcid.org/0000-0001-6505-8535Hui Chiu1https://orcid.org/0000-0002-1820-8411David J Anderson2https://orcid.org/0000-0001-6175-3872Division of Biology and Biological Engineering, Tianqiao and Chrissy Chen Institute for Neuroscience, California Institute of Technology, Pasadena, United StatesDivision of Biology and Biological Engineering, Tianqiao and Chrissy Chen Institute for Neuroscience, California Institute of Technology, Pasadena, United StatesDivision of Biology and Biological Engineering, Tianqiao and Chrissy Chen Institute for Neuroscience, California Institute of Technology, Pasadena, United States; Howard Hughes Medical Institute, Chevy Chase, United StatesMonitoring neuronal activity at single-cell resolution in freely moving Drosophila engaged in social behaviors is challenging because of their small size and lack of transparency. Extant methods, such as Flyception, are highly invasive. Whole-brain calcium imaging in head-fixed, walking flies is feasible but the animals cannot perform the consummatory phases of social behaviors like aggression or mating under these conditions. This has left open the fundamental question of whether neurons identified as functionally important for such behaviors using loss- or gain-of-function screens are actually active during the natural performance of such behaviors, and if so during which phase(s). Here, we perform brain-wide mapping of active cells expressing the Immediate Early Gene hr38 using a high-sensitivity/low background fluorescence in situ hybridization (FISH) amplification method called HCR-3.0. Using double-labeling for hr38 mRNA and for GFP, we describe the activity of several classes of aggression-promoting neurons during courtship and aggression, including P1a cells, an intensively studied population of male-specific interneurons. Using HI-FISH in combination with optogenetic activation of aggression-promoting neurons (opto-HI-FISH), we identify candidate downstream functional targets of these cells in a brain-wide, unbiased manner. Finally, we compare the activity of P1a neurons during sequential performance of courtship and aggression, using intronic vs. exonic hr38 probes to differentiate newly synthesized nuclear transcripts from cytoplasmic transcripts synthesized at an earlier time. These data provide evidence suggesting that different subsets of P1a neurons may be active during courtship vs. aggression. HI-FISH and associated methods may help to fill an important lacuna in the armamentarium of tools for neural circuit analysis in Drosophila.https://elifesciences.org/articles/92380immediate early geneHr38activity-dependent mappingsocial behaviorhybridization chain reactionHCR |
| spellingShingle | Kiichi Watanabe Hui Chiu David J Anderson Whole-brain in situ mapping of neuronal activation in Drosophila during social behaviors and optogenetic stimulation eLife immediate early gene Hr38 activity-dependent mapping social behavior hybridization chain reaction HCR |
| title | Whole-brain in situ mapping of neuronal activation in Drosophila during social behaviors and optogenetic stimulation |
| title_full | Whole-brain in situ mapping of neuronal activation in Drosophila during social behaviors and optogenetic stimulation |
| title_fullStr | Whole-brain in situ mapping of neuronal activation in Drosophila during social behaviors and optogenetic stimulation |
| title_full_unstemmed | Whole-brain in situ mapping of neuronal activation in Drosophila during social behaviors and optogenetic stimulation |
| title_short | Whole-brain in situ mapping of neuronal activation in Drosophila during social behaviors and optogenetic stimulation |
| title_sort | whole brain in situ mapping of neuronal activation in drosophila during social behaviors and optogenetic stimulation |
| topic | immediate early gene Hr38 activity-dependent mapping social behavior hybridization chain reaction HCR |
| url | https://elifesciences.org/articles/92380 |
| work_keys_str_mv | AT kiichiwatanabe wholebraininsitumappingofneuronalactivationindrosophiladuringsocialbehaviorsandoptogeneticstimulation AT huichiu wholebraininsitumappingofneuronalactivationindrosophiladuringsocialbehaviorsandoptogeneticstimulation AT davidjanderson wholebraininsitumappingofneuronalactivationindrosophiladuringsocialbehaviorsandoptogeneticstimulation |