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A spike-timing mechanism for action selection

Abstract

We discovered a bimodal behavior in the genetically tractable organism Drosophila melanogaster that allowed us to directly probe the neural mechanisms of an action selection process. When confronted by a predator-mimicking looming stimulus, a fly responds with either a long-duration escape behavior sequence that initiates stable flight or a distinct, short-duration sequence that sacrifices flight stability for speed. Intracellular recording of the descending giant fiber (GF) interneuron during head-fixed escape revealed that GF spike timing relative to parallel circuits for escape actions determined which of the two behavioral responses was elicited. The process was well described by a simple model in which the GF circuit has a higher activation threshold than the parallel circuits, but can override ongoing behavior to force a short takeoff. Our findings suggest a neural mechanism for action selection in which relative activation timing of parallel circuits creates the appropriate motor output.

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Figure 1: Flies select between two escape sequences when confronted by a looming stimulus.
Figure 2: The GFs are necessary and sufficient for eliciting short-mode escapes.
Figure 3: GF Na+ spikes drive the short-mode escape.
Figure 4: GF spike timing determines escape mode.
Figure 5: A dual-circuit, angular size threshold model recapitulates the timing and selection between short- or long-mode escape.
Figure 6: GF-mediated short escapes confer a survival advantage within an ethologically relevant regime.

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Acknowledgements

We thank T. Ngo and A. Jenett (Janelia Farm Research Campus) for providing the GF-split GAL4 line, B.D. Pfeiffer (Janelia Farm Research Campus) for pJFRC12-10XUAS-IVS-myrGFP, pJFRC28-10XUAS-IVS-GFP, and pJFRC49-10XUAS-IVS-eGFPKir2.1 flies, and K. Wantanabe (Caltech) for the pUAS-EGFP-Kir2.1 DNA. We thank V. Jayaraman and A. Karpova (Janelia Farm Research Campus) for providing UAS-CsChrimson flies. We thank P. Herold for damselfly husbandry and assistance with data collection and R. Franconville for help troubleshooting P2X2 receptor experiments. We thank V. Jayaraman, G. Murphy and S. Huston for their comments on the manuscript. We thank the Janelia Fly Facility (T. Laverty, A. Cavallaro, K. Hibbard, D. Hall, M. Mercer, D. Fetter, J. McMachon, J.-C. Kao and D. Ruiz). We thank the Janelia Instrument Design and Fabrication Department for help with the behavioral apparatus.

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Authors and Affiliations

Authors

Contributions

C.R.v.R., A.L. and G.M.C. prepared the paper. C.R.v.R., P.B. and G.M.C. analyzed data. C.R.v.R., P.B., W.R.W. A.L.Y. and G.M.C. performed experiments. C.R.v.R., A.L. and G.M.C. designed experiments. G.Z.Z. provided genetic tools. C.R.v.R. built the electrophysiology apparatus. M.Y.P., W.R.W. and G.M.C. built the behavioral apparatus. A.L. built the damselfly tracking system.

Corresponding author

Correspondence to Gwyneth M Card.

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The authors declare no competing financial interests.

Integrated supplementary information

Supplementary Figure 1 Apparatus for video recording an individual fly's response to projected looming stimuli.

Supplementary Figure 2 Motor program durations for several genotypes.

(a) Escape sequence duration histograms (bars) and probability density functions (black lines) following Gaussian curve fits for each control genotype, combined for r/v=14, 40, and 70 ms. n are listed on histograms. Genotypes: “WT”: wild-type DL, “GF WT”: w+GF-split-GAL4 UAS-myrGFP, “AD Kir”: w+ GF-split-AD UAS-Kir2.1, “DBD Kir”: w+GF-split-DBD UAS-Kir2.1, “WT Kir”: w+ UAS-Kir2.1. (b) Escape sequence duration histogram and probability density function with GF silencing. “GF Kir”: w+GF-split-GAL4 UAS-myrGFP UAS-Kir2.1. Inset: Expressing Kir2.1 with the GF split-GAL4 driver line causes a significant hyperpolarization of the GF resting membrane potential, compared to control lines (boxplots as described in Fig. 2, n = 3 flies, two sided t-test, ** = P<0.005, t = 6.352). “GF +”: w+GF-split-GAL4 UAS-myrGFP

Supplementary Figure 3 Takeoff percentages and takeoff timing for several genotypes.

(a) Takeoff percentages across r/v values of 14, 40, and 70 ms, from left to right (number of flies as listed, χ2-test, P = 0.015, 0.087, 0.148 and χ2 = 11.81, 6.447, and 2.371 for r/v = 14, 40, and 70 ms, Bonferroni correction post hoc where permitted, ** = P < 0.005). (b) Time of takeoff, with respect to time to contact (time of contact, TOC = 0 ms), across r/v (Kruskal-Wallis, P=<<0.0001, <<0.0001, and 0.021 and χ2 = 41.01, 42.80, and 13.27 for r/v = 14, 40, and 70 ms, Bonferroni correction post hoc, ** = P <0.005, ***=P<0.001).

Supplementary Figure 4 GF Na+ spikes can be evoked upon ATP stimulation of P2X2 receptor expressing flies.

(a, b) ATP (1mM) stimulation of P2X2 receptor expressing GFs evokes spiking in the ipsilateral (a, black trace, representative example from 5 flies) or contralateral (b, black trace, representative example from 3 flies) GF but not in GFs only expressing GFP (gray traces, representative example of 3 flies each). (c) Ipsilateral ATP pulses can also evoke both large Ca2+-mediated potentials and small, narrow spikes (inset is expanded yellow box). (d) Tetrodotoxin (TTX, 1 μm) application eliminates the small, narrow Na+ spikes while the Ca2+-mediated potentials remain (representative example from 3 flies). Genotypes: GF-split-GAL4 UAS-GFP UAS-P2X2, GF-split-GAL4 UAS-GFP (control).

Supplementary Figure 5

Example spike (top, representative response chosen from 10 flies) and subthreshold (middle, representative response chosen from 33 flies) traces in response to looms (bottom) at three separate r/v values (a) 14, (b) 40, and (c) 70 ms. Genotype: w+ GF-split-GAL4 UAS-GFP.

Supplementary Figure 6 Dual circuit, size threshold model.

(a) The time of wing elevation for GF silenced flies (red circles, mean ± standard deviation, n=57, 58, 99, 61 for r/v=8, 14, 40, and 70 ms, respectively) or GF spikes (red squares, n=8, 12, 7 for r/v=14, 40 and 70 ms, respectively) with respect to r/v. The neural delay (δ) for each pathway was taken directly from the y-intercept of the linear fit (R2 > 0.999 in both cases). Inset: The stimulus size (mean ± standard deviation) at the fixed neural delay preceding wing elevation or GF spiking. (b) The standard deviation of wing elevation or GF spiking increased linearly with r/v (R2 = >0.999 and 0.9977 while slopes = 1.353 and 1.213 for GF spike or GF Kir wing timing, respectively). The slopes were used to derive the standard deviation for the size threshold (θthresh). (c) Model schematic displaying the 4 variables, drawn from their respective distributions, used to select the circuit and determine the timing of escape on a per fly basis.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–6 (PDF 971 kb)

Supplementary Methods Checklist

(PDF 497 kb)

Supplementary Video 1

Example of short duration escape; time corresponds to start of video. (MP4 294 kb)

Supplementary Video 2

Example of long duration, wing-raised, escape; time corresponds to start of video. (MP4 432 kb)

Supplementary Video 3

Example of escape upon light activation of CsChrimson expressing GFs. (MOV 183 kb)

Supplementary Video 4

Front view of escape in a tethered prep while recording whole-cell spiking responses from the GF. (MP4 508 kb)

Supplementary Video 5

Side view of escape in a tethered prep while recording whole-cell spiking responses from the GF. (MP4 910 kb)

Supplementary Video 6

Side view of no escape in a tethered prep while recording whole-cell subthreshold responses from the GF. (AVI 1614 kb)

Supplementary Video 7

Fly capture during a damselfly predation. (MP4 256 kb)

Supplementary Video 8

Successful fly escape from a damselfly predation attempt. (MP4 681 kb)

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von Reyn, C., Breads, P., Peek, M. et al. A spike-timing mechanism for action selection. Nat Neurosci 17, 962–970 (2014). https://doi.org/10.1038/nn.3741

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