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FlyMAD: rapid thermogenetic control of neuronal activity in freely walking Drosophila


Rapidly and selectively modulating the activity of defined neurons in unrestrained animals is a powerful approach in investigating the circuit mechanisms that shape behavior. In Drosophila melanogaster, temperature-sensitive silencers and activators are widely used to control the activities of genetically defined neuronal cell types. A limitation of these thermogenetic approaches, however, has been their poor temporal resolution. Here we introduce FlyMAD (the fly mind-altering device), which allows thermogenetic silencing or activation within seconds or even fractions of a second. Using computer vision, FlyMAD targets an infrared laser to freely walking flies. As a proof of principle, we demonstrated the rapid silencing and activation of neurons involved in locomotion, vision and courtship. The spatial resolution of the focused beam enabled preferential targeting of neurons in the brain or ventral nerve cord. Moreover, the high temporal resolution of FlyMAD allowed us to discover distinct timing relationships for two neuronal cell types previously linked to courtship song.

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Figure 1: FlyMAD system overview.
Figure 2: Behavioral responses to acute neuronal silencing and activation.
Figure 3: Body part–specific targeting using TTM tracking.
Figure 4: Acute activation of courtship neurons.

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We thank S. Bidaye for data on the VT50660-GAL4 genotype, the Institute of Molecular Pathology (IMP) workshop for help fabricating the hardware, M. Palfreyman and M. Dickinson for insightful discussion, L. Fenk for technological support and P. Masek for insight into IR activation of TrpA1. We thank J.H. Simpson (Howard Hughes Medical Institute, Janelia Farm Research Campus) for providing UAS-Shibirets1 flies. The fruit fly drawings in Figure 3 are modified from versions made available by Database Center for Life Science (DBCLS) under a CC 2.1 license. This work was supported by the Natural Sciences and Engineering Research Council of Canada by a postgraduate scholarship to D.E.B., European Research Council (ERC) Starting grant 281884 and Wiener Wissenschafts-, Forschungs- und Technologiefonds (WWTF) grant CS2011-029 to A.D.S., ERC Advanced grant 233306 to B.J.D. and IMP core funding.

Author information

Authors and Affiliations



D.E.B., B.J.D. and A.D.S. conceived of the project. D.E.B., J.R.S., D.H., A.P. and A.D.S. developed the hardware and software. D.E.B. and D.H. performed experiments. All authors contributed to data analysis, interpretation and writing the manuscript.

Corresponding authors

Correspondence to Barry J Dickson or Andrew D Straw.

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Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–16 and Supplementary Tables 1–5 (PDF 5575 kb)

FlyMAD: Rapid thermogenetic control of neuronal activity in freely-walking Drosophila

Summary of FlyMAD objectives, operation and results including thermogenetic silencing and activation. (MP4 10439 kb)

Silencing motoneurons with ShibireTS

Thermogenetic silencing of motoneurons reversibly disrupts locomotion. Genotype was +; OK371-Gal4/uas-ShibireTS1. (MP4 522 kb)

Silencing photoreceptors blocks the optomotor response

Thermogenetic silencing of visual neurons disrupts optomotor response. Genotype was Rh1-Gal4; UAS-ShibireTS. (MP4 4489 kb)

Activating Moonwalker neurons with TrpA1

Thermogenetic activation of moonwalker neurons induces backwards walking. Genotype was VT50660-Gal4; UAS-TrpA1. (MP4 864 kb)

Estimating the error of Through-The-Mirror (TTM) Tracking

In TTM tracking, the mirror movement command signal is proportional to the tracking error. The lower panel of this video shows the error in X and Y directions over time, while the upper panels show the wide and TTM camera views. (MP4 2641 kb)

Head-targeted heating induces proboscis extension from dopaminergic activation

Thermogenetic activation of flies with genotype TH-Gal4; UAS-TrpA1 causes proboscis extension. (MP4 19498 kb)

Activating song neurons in the VNC is stronger when targeting thorax than head

Thermogenetic activation of flies expressing TrpA1 in thoracic song neurons induces lower latency and more frequent singing when targeting the thorax than the head. (MP4 18944 kb)

P1-dependent courtship persists after stimulus ceases

A fly expressing TrpA1 in P1 performs courtship towards plasticine balls during and long after the thermogenetic stimulus is applied. Genotype is NP2361-Gal4; UAS>stop>TrpA1myc; fruFLP. (MP4 3585 kb)

pIP10-dependent courtship is closely linked to artificial activation

A fly expressing TrpA1 in pIP10 extends wings only when the thermogenetic stimulus is applied. Genotype is VT40347-Gal4; UAS>stop>TrpA1myc; fruFLP. (MP4 687 kb)

Supplementary Software

FlyMAD 0.9 installer. The file contains an installer for FlyMAD which depends on a computer system running a 64bit version of Ubuntu 12.04. The design files for the chamber, the optomotor drum, and the circuit board are included. (ZIP 2961 kb)

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Bath, D., Stowers, J., Hörmann, D. et al. FlyMAD: rapid thermogenetic control of neuronal activity in freely walking Drosophila. Nat Methods 11, 756–762 (2014).

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