A subset of dopamine neurons signals reward for odour memory in Drosophila

Journal name:
Nature
Volume:
488,
Pages:
512–516
Date published:
DOI:
doi:10.1038/nature11304
Received
Accepted
Published online

Animals approach stimuli that predict a pleasant outcome1. After the paired presentation of an odour and a reward, Drosophila melanogaster can develop a conditioned approach towards that odour2, 3. Despite recent advances in understanding the neural circuits for associative memory and appetitive motivation4, the cellular mechanisms for reward processing in the fly brain are unknown. Here we show that a group of dopamine neurons in the protocerebral anterior medial (PAM) cluster signals sugar reward by transient activation and inactivation of target neurons in intact behaving flies. These dopamine neurons are selectively required for the reinforcing property of, but not a reflexive response to, the sugar stimulus. In vivo calcium imaging revealed that these neurons are activated by sugar ingestion and the activation is increased on starvation. The output sites of the PAM neurons are mainly localized to the medial lobes of the mushroom bodies (MBs), where appetitive olfactory associative memory is formed5, 6. We therefore propose that the PAM cluster neurons endow a positive predictive value to the odour in the MBs. Dopamine in insects is known to mediate aversive reinforcement signals5, 7, 8, 9, 10, 11. Our results highlight the cellular specificity underlying the various roles of dopamine and the importance of spatially segregated local circuits within the MBs.

At a glance

Figures

  1. Thermo-activation with DDC-GAL4 induces appetitive memory.
    Figure 1: Thermo-activation with DDC-GAL4 induces appetitive memory.

    a, b, Expression patterns of TH-GAL4 (a) and DDC-GAL4 (b) in the brain with a neuropile counterstaining (magenta). Outline, MB; arrowheads, PAM neurons. Scale bars, 20μm. c, Protocol for dTRPA1(Trp)-mediated reinforcement substitution. MCH, 4-methylcyclohexanol; OCT, octan-3-ol. d, Thermo-activation with DDC-GAL4 and TH-GAL4 with or without starvation. PI, performance index. n = 16. e, Retention of induced memory. n = 10–27. f, Protocol for feeding before the training or the test of 12-h memory. Flies were satiated with a short feed (30min) or continuous feeding. g, Test of 12-h memory by thermo-activation with DDC-GAL4. n = 16. Midline, box boundaries and whiskers are median, quartiles and 10th and 90th centiles, respectively. Results in d and e are means±s.e.m. Two asterisks, P<0.01; three asterisks, P<0.001; n.s., not significant.

  2. The PAM cluster neurons signal reward for olfactory memory.
    Figure 2: The PAM cluster neurons signal reward for olfactory memory.

    a, Expression pattern of R58E02-GAL4 in the central brain revealed with UAS-mCD8::GFP. be, Co-localization of dopamine (magenta) and GAL4-expressing cells (green) in the PAM cluster of DDC-GAL4 (b, c, n = 6) and R58E02-GAL4 (d, e, n = 4). f, Thermo-activation with R58E02-GAL4/UAS-dTrpA1 induces significant appetitive memory. n = 16. gj, R58E02-GAL80 silences transgene expression in most PAM neurons of DDC-GAL4 (arrowheads in g and h) without greatly affecting other neurons (such as the suboesophageal ganglion (arrows)). k, R58E02-GAL80 suppresses the induction of appetitive memory by DDC-GAL4/UAS-dTrpA1. n = 10. l, m, Blockade of the PAM neurons in R58E02-GAL4/UAS-shits1 during memory acquisition (l) or applied after training (m). n, R58E02-GAL80 rescues the memory impairment in DDC-GAL4/UAS-shits1, n = 14–20. Upper panels in ln, protocols; lower panels, sugar conditioning. Results with error bars are means±s.e.m. Three asterisks, P<0.001; n.s., not significant. Scale bars, 20μm.

  3. The PAM neurons convey the reward signal to the MB.
    Figure 3: The PAM neurons convey the reward signal to the MB.

    ac, Presynaptic terminals (magenta) of the PAM neurons (green) in R58E02-GAL4 are localized to the MB (outline). d, The defect in sugar-induced memory in dumb2, a piggyBac insertion allowing GAL4-mediated dDA1 expression16, is rescued by MB247-GAL4. n = 16. e, Appetitive memory induced by thermo-activation in R58E02-LexA/LexAop2-dTrpA1. n = 16. f, Thermo-activation of the PAM neurons in dumb2 induces memory by rescuing dDA1 in the MB. n = 14–22. gi, No detectable overlap of R58E02-LexA-labelled neurons expressing mCD8::GFP (green) and MB-M3 neurons (magenta; arrowheads) visualized with mCD8::RFP. See also Supplementary Fig. 7. jl, Differential labelling of R58E02-LexA (green) and TH-GAL4 (magenta) in the MB lobe region. Results with error bars are means±s.e.m. Two asterisks, P<0.01; three asterisks, P<0.001; n.s., not significant. Scale bars, 20μm.

  4. The PAM neurons integrate reward and relevant signals.
    Figure 4: The PAM neurons integrate reward and relevant signals.

    a, Thermo-activation of the PAM neurons induces appetitive memory without octopamine. n = 16–22. bd, Octopamine and PAM neurons contact each other (arrows) in a protocerebral region (c) and the spur of the γ lobe (d). eg, The PAM neurons respond selectively to the sugar reward. e, Representative images (anterior down) and calcium responses of the PAM neurons at different levels of the MB. f, Time course of responses. n = 10. Black bar: stimulus application. g, Average response to different gustatory substances. Terminals in the β′ lobe responded significantly to water and caffeine (dagger, P<0.05; two daggers, P<0.01 in one-sample t-test from zero). h, Response of the PAM neurons without starvation or with tarsal stimulation only. n>7. i, Model of a reward circuit in the fly brain. Dashed arrows indicate hypothetical pathways. The reward value integrated by the PAM neurons is signalled by dopamine (DA) to the MB (green shading), thus they may drive associative plasticity in odour-representing Kenyon cells (KC, grey). OA, octopamine; PN, projection neurons. Results with error bars are means±s.e.m. Asterisk, P<0.05; two asterisks, P<0.01; three asterisks, P<0.001. Scale bars, 20μm.

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Author information

Affiliations

  1. Max-Planck-Institut für Neurobiologie, Martinsried 82152, Germany

    • Chang Liu,
    • Nobuhiro Yamagata,
    • Yoshinori Aso,
    • Anja B. Friedrich,
    • Igor Siwanowicz &
    • Hiromu Tanimoto
  2. Laboratory of Primate Cognitive Neuroscience, Kunming Institute of Zoology, The Chinese Academy of Sciences, Kunming, Yunnan 650223, China

    • Chang Liu
  3. Graduate University of the Chinese Academy of Sciences, Beijing 100049, China

    • Chang Liu
  4. Genes and Dynamics of Memory Systems, Neurobiology Unit, Centre National de la Recherche Scientifique, École Supérieure de Physique et de Chimie Industrielles, 75005 Paris, France

    • Pierre-Yves Plaçais &
    • Thomas Preat
  5. Janelia Farm Research Campus, Howard Hughes Medical Institute, Ashburn, Virginia 20147, USA

    • Barret D. Pfeiffer,
    • Yoshinori Aso &
    • Gerald M. Rubin

Contributions

C.L., N.Y., Y.A. and H.T. designed and C.L. and N.Y. performed all the behavioural experiments in this study. P.Y.P., T.P. and H.T. designed in vivo imaging experiments, and P.Y.P. and T.P. devised a new gustatory stimulation method. P.Y.P. performed imaging experiments and analysed the data. B.D.P. and G.M.R. designed and generated the new transgenic flies (GAL4, GAL80, LexA and LexAop2-dTrpA1 lines). Y.A. and H.T. identified R58E02 by using a database of GAL4 expression patterns created by G.M.R. and the Janelia Farm Fly Light Project Team. A.B.F. and I.S. performed immunohistochemistry, and C.L., A.B.F. and H.T. analysed the microscopic data. C.L. and H.T. made the figures and wrote the paper with the help of all the other authors.

Competing financial interests

The authors declare no competing financial interests.

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Supplementary information

PDF files

  1. Supplementary Information (2.4M)

    This file contains Supplementary Figures 1-9 and full legends for Supplementary Movies 1-5.

Movies

  1. Supplementary Movie 1 (5.4M)

    This file contains a movie showing the expression pattern of R58E02-GAL4 in the brain.

  2. Supplementary Movie 2 (4.2M)

    This file contains a movie showing the expression pattern of DDC-GAL4 in the central brain.

  3. Supplementary Movie 3 (3.7M)

    This file contains a movie showing the expression pattern of DDC-GAL4 with R58E02-GAL80 in the central brain.

  4. Supplementary Movie 4 (116K)

    This file contains a movie showing differential labelling of NP5272-GAL4 and R58E02-LexA in the cell body region of the PAM cluster.

  5. Supplementary Movie 5 (3.7M)

    This file contains a movie showing differential labelling of TH-GAL4 and R58E02-LexA in the MB.

Additional data