Freeze-frame imaging of synaptic activity using SynTagMA

Pérez-Álvarez, Alberto; O’Toole, Ryan J.; Wei, Yang; Arganda‐Carreras, Ignacio; Lamothe-Molina, Paul J; Moeyaert, Benjamien; Mohr, Manuel; Panzera, Lauren C.; Schulze, Christian; Schreiter, Eric R.; Gee, Christine E.; Hoppa, Michael B.; Oertner, Thomas G.

doi:10.1101/538041

Cited by 2 publications

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References 64 publications

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“…With these tools, CaMPARI freely diffuses through the cytoplasm and so subcellular information is lost. Recently the tool SynTagMa (Synaptic Tag for Mapping Activity) was created to label active synapses by fusing CaMPARI to synaptophysin or PSD95 and labeling presynaptic boutons and dendritic spines, respectively (Perez-Alvarez et al, 2019). This tool could be further adapted to more specifically label circuitry in adult Drosophila or any other species where transgenic methods allow the creation of viable transgenic animals.…”

Section: Discussionmentioning

confidence: 99%

The Photoconvertible Fluorescent Probe, CaMPARI, Labels Active Neurons in Freely-Moving Intact Adult Fruit Flies

Edwards

Hoppa

Bosco

2020

Front. Neural Circuits

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Linking neural circuitry to behavior by mapping active neurons in vivo is a challenge. Both genetically encoded calcium indicators (GECIs) and intermediate early genes (IEGs) have been used to pinpoint active neurons during a stimulus or behavior but have drawbacks such as limiting the movement of the organism, requiring a priori knowledge of the active region or having poor temporal resolution. Calcium-modulated photoactivatable ratiometric integrator (CaMPARI) was engineered to overcome these spatial-temporal challenges. CaMPARI is a photoconvertible protein that only converts from green to red fluorescence in the presence of high calcium concentration and 405 nm light. This allows the experimenter to precisely mark active neurons within defined temporal windows. The photoconversion can then be quantified by taking the ratio of the red fluorescence to the green. CaMPARI promises the ability to trace active neurons during a specific stimulus; however, CaMPARI's uses in adult Drosophila have been limited to photoconversion during fly immobilization. Here, we demonstrate a method that allows photoconversion of multiple freely-moving intact adult flies during a stimulus. Flies were placed in a dish with filter paper wet with acetic acid (pH = 2) or neutralized acetic acid (pH = 7) and exposed to photoconvertible light (60 mW) for 30 min (500 ms on, 200 ms off). Immediately following photoconversion, whole flies were fixed and imaged by confocal microscopy. The red:green ratio was quantified for the DC4 glomerulus, a bundle of neurons expressing Ir64a, an ionotropic receptor that senses acids in the Drosophila antennal lobe. Flies exposed to acetic acid showed 1.3-fold greater photoconversion than flies exposed to neutralized acetic acid. This finding was recapitulated using a more physiological stimulus of apple cider vinegar. These results indicate that CaMPARI can be used to label neurons in intact, freely-moving adult flies and will be useful for identifying the circuitry underlying complex behaviors.

show abstract

Section: Discussionmentioning

confidence: 99%

The Photoconvertible Fluorescent Probe, CaMPARI, Labels Active Neurons in Freely-Moving Intact Adult Fruit Flies

Edwards

Hoppa

Bosco

2020

Front. Neural Circuits

Self Cite

View full text Add to dashboard Cite

show abstract

Section: Discussionmentioning

confidence: 99%

The Photoconvertible Fluorescent Probe, CaMPARI, Labels Active Neurons in Freely-Moving Intact Adult Fruit Flies

Edwards

Hoppa

Bosco

2020

Preprint

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AbstractLinking neural circuitry to behavior by mapping active neurons in vivo is a challenge. Both genetically encoded calcium indicators (GECIs) and intermediate early genes (IEGs) have been used to pinpoint active neurons during a stimulus or a behavior but have drawbacks such as limiting the movement of the organism, requiring a priori knowledge of the active region or having poor temporal resolution. CaMPARI (calcium-modulated photoactivatable ratiometric integrator) was engineered to overcome these spatial-temporal challenges. CaMPARI is a photoconvertible protein that only converts from green to red fluorescence in the presence of high calcium concentration and violet (405 nm) light. This allows the experimenter to precisely mark active neurons within defined temporal windows. The photoconversion can then be quantified by taking the ratio of the red fluorescence to the green. CaMPARI promises the ability to trace active neurons during specific stimulus; however, CaMPARI’s uses in adult Drosophila have been limited to photoconversion during fly immobilization. Here, we demonstrate a method that allows photoconversion of multiple freely-moving intact adult flies during a stimulus. Flies were placed in a dish with filter paper wet with acetic acid (pH=2) or neutralized acetic acid (pH=7) and exposed to photoconvertible light (60 mW) for 30 min (500 ms on, 200 ms off). Immediately following photoconversion, whole flies were fixed and imaged by confocal microscopy. The red:green ratio was quantified for the DC4 glomerulus, a bundle of neurons expressing Ir64a, an ionotropic receptor that senses acids in the Drosophila antennal lobe. Flies exposed to acetic acid showed 1.3-fold greater photoconversion than flies exposed to neutralized acetic acid. This finding was recapitulated using a more physiological stimulus of apple cider vinegar. These results indicate that CaMPARI can be used to label neurons in intact, freely-moving adult flies and will be useful for identifying the circuitry underlying complex behaviors.

show abstract

Freeze-frame imaging of synaptic activity using SynTagMA

Cited by 2 publications

References 64 publications

The Photoconvertible Fluorescent Probe, CaMPARI, Labels Active Neurons in Freely-Moving Intact Adult Fruit Flies

The Photoconvertible Fluorescent Probe, CaMPARI, Labels Active Neurons in Freely-Moving Intact Adult Fruit Flies

The Photoconvertible Fluorescent Probe, CaMPARI, Labels Active Neurons in Freely-Moving Intact Adult Fruit Flies

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