Abstract

Midbrain dopamine neurons are well known for their role in reward-based reinforcement learning. We found that the activity of dopamine axons in the posterior tail of the striatum (TS) scaled with the novelty and intensity of external stimuli, but did not encode reward value. We demonstrated that the ablation of TS-projecting dopamine neurons specifically inhibited avoidance of novel or high-intensity stimuli without affecting animals’ initial avoidance responses, suggesting a role in reinforcement rather than simply in avoidance itself. Furthermore, we found that animals avoided optogenetic activation of dopamine axons in TS during a choice task and that this stimulation could partially reinstate avoidance of a familiar object. These results suggest that TS-projecting dopamine neurons reinforce avoidance of threatening stimuli. More generally, our results indicate that there are at least two axes of reinforcement learning using dopamine in the striatum: one based on value and one based on external threat.

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References

  1. 1.

    Schultz, W., Dayan, P. & Montague, P. R. A neural substrate of prediction and reward. Science 275, 1593–1599 (1997).

  2. 2.

    Cohen, J. Y., Haesler, S., Vong, L., Lowell, B. B. & Uchida, N. Neuron-type-specific signals for reward and punishment in the ventral tegmental area. Nature 482, 85–88 (2012).

  3. 3.

    Roitman, M. F., Wheeler, R. A., Wightman, R. M. & Carelli, R. M. Real-time chemical responses in the nucleus accumbens differentiate rewarding and aversive stimuli. Nat. Neurosci. 11, 1376–1377 (2008).

  4. 4.

    Matsumoto, H., Tian, J., Uchida, N. & Watabe-Uchida, M. Midbrain dopamine neurons signal aversion in a reward-context-dependent manner. eLife 5, e17328 (2016).

  5. 5.

    Tsai, H.-C. et al. Phasic firing in dopaminergic neurons is sufficient for behavioral conditioning. Science 324, 1080–1084 (2009).

  6. 6.

    Ilango, A. et al. Similar roles of substantia nigra and ventral tegmental dopamine neurons in reward and aversion. J. Neurosci. 34, 817–822 (2014).

  7. 7.

    Chang, C. Y. et al. Brief optogenetic inhibition of dopamine neurons mimics endogenous negative reward prediction errors. Nat. Neurosci. 19, 111–116 (2016).

  8. 8.

    Bayer, H. M. & Glimcher, P. W. Midbrain dopamine neurons encode a quantitative reward prediction error signal. Neuron 47, 129–141 (2005).

  9. 9.

    Matsumoto, M. & Hikosaka, O. Two types of dopamine neuron distinctly convey positive and negative motivational signals. Nature 459, 837–841 (2009).

  10. 10.

    Menegas, W., Babayan, B. M., Uchida, N. & Watabe-Uchida, M. Opposite initialization to novel cues in dopamine signaling in ventral and posterior striatum in mice. eLife 6, e21886 (2017).

  11. 11.

    Kakade, S. & Dayan, P. Dopamine: generalization and bonuses. Neural Netw. 15, 549–559 (2002).

  12. 12.

    Lerner, T. N. et al. Intact-brain analyses reveal distinct information carried by SNc dopamine subcircuits. Cell 162, 635–647 (2015).

  13. 13.

    Parker, N. F. et al. Reward and choice encoding in terminals of midbrain dopamine neurons depends on striatal target. Nat. Neurosci. 19, 845–854 (2016).

  14. 14.

    Howe, M. W. & Dombeck, D. A. Rapid signalling in distinct dopaminergic axons during locomotion and reward. Nature 535, 505–510 (2016).

  15. 15.

    Eshel, N. et al. Arithmetic and local circuitry underlying dopamine prediction errors. Nature 525, 243–246 (2015).

  16. 16.

    Bonhomme, N., De Deurwaèrdere, P., Le Moal, M. & Spampinato, U. Evidence for 5-HT4 receptor subtype involvement in the enhancement of striatal dopamine release induced by serotonin: a microdialysis study in the halothane-anesthetized rat. Neuropharmacology 34, 269–279 (1995).

  17. 17.

    Ciabatti, E., González-Rueda, A., Mariotti, L., Morgese, F. & Tripodi, M. Life-long genetic and functional access to neural circuits using self-inactivating rabies virus. Cell 170, 382–392.e14 (2017).

  18. 18.

    Menegas, W., Uchida, N. & Watabe-Uchida, M. A self-killing rabies virus that leaves a trace on the DNA. Trends Neurosci. 40, 589–591 (2017).

  19. 19.

    Bar-Hillel, M. & Wagenaar, W. A. The perception of randomness. Adv. Appl. Math. 12, 428–454 (1991).

  20. 20.

    Soltani, A., Lee, D. & Wang, X.-J. Neural mechanism for stochastic behaviour during a competitive game. Neural Netw. 19, 1075–1090 (2006).

  21. 21.

    LeDoux, J. & Daw, N. D. Surviving threats: neural circuit and computational implications of a new taxonomy of defensive behaviour. Nat. Rev. Neurosci. 19, 269–282 (2018).

  22. 22.

    Berlyne, D. E. Curiosity and exploration. Science 153, 25–33 (1966).

  23. 23.

    Moser, E. I., Moser, M. B. & Andersen, P. Potentiation of dentate synapses initiated by exploratory learning in rats: dissociation from brain temperature, motor activity, and arousal. Learn. Mem. 1, 55–73 (1994).

  24. 24.

    Albasser, M. M. et al. Perirhinal cortex lesions uncover subsidiary systems in the rat for the detection of novel and familiar objects. Eur. J. Neurosci. 34, 331–342 (2011).

  25. 25.

    James, W. The Principles of Psychology. (Holt, New York, NY, USA, 1890).

  26. 26.

    Tong, Q. et al. Synaptic glutamate release by ventromedial hypothalamic neurons is part of the neurocircuitry that prevents hypoglycemia. Cell Metab. 5, 383–393 (2007).

  27. 27.

    Yamaguchi, T., Qi, J., Wang, H.-L., Zhang, S. & Morales, M. Glutamatergic and dopaminergic neurons in the mouse ventral tegmental area. Eur. J. Neurosci. 41, 760–772 (2015).

  28. 28.

    Menegas, W. et al. Dopamine neurons projecting to the posterior striatum form an anatomically distinct subclass. eLife 4, e10032 (2015).

  29. 29.

    Fiorillo, C. D., Song, M. R. & Yun, S. R. Multiphasic temporal dynamics in responses of midbrain dopamine neurons to appetitive and aversive stimuli. J. Neurosci. 33, 4710–4725 (2013).

  30. 30.

    Ghazizadeh, A., Griggs, W. & Hikosaka, O. Ecological origins of object salience: reward, uncertainty, aversiveness, and novelty. Front. Neurosci. 10, 378 (2016).

  31. 31.

    Krusemark, E. A. & Li, W. Do all threats work the same way? Divergent effects of fear and disgust on sensory perception and attention. J. Neurosci. 31, 3429–3434 (2011).

  32. 32.

    Lee, D. H. & Anderson, A. K. Reading what the mind thinks from how the eye sees. Psychol. Sci. 28, 494–503 (2017).

  33. 33.

    Oleson, E. B., Gentry, R. N., Chioma, V. C. & Cheer, J. F. Subsecond dopamine release in the nucleus accumbens predicts conditioned punishment and its successful avoidance. J. Neurosci. 32, 14804–14808 (2012).

  34. 34.

    Rescorla, R. A. & Wagner, A. R. A theory of Pavlovian conditioning: variations in the effectiveness of reinforcement and nonreinforcement. In: A. H. Black, W. F. Prokasy eds.. Classical Conditioning II: Current Research and Theory (pp. 64–99. Appleton-Century-Crofts, New York, NY, USA, 1972).

  35. 35.

    Lloyd, K. & Dayan, P. Safety out of control: dopamine and defence. Behav. Brain Funct. 12, 15 (2016).

  36. 36.

    Rogan, M. T., Leon, K. S., Perez, D. L. & Kandel, E. R. Distinct neural signatures for safety and danger in the amygdala and striatum of the mouse. Neuron 46, 309–320 (2005).

  37. 37.

    Guarraci, F. A., Frohardt, R. J. & Kapp, B. S. Amygdaloid D1 dopamine receptor involvement in Pavlovian fear conditioning. Brain Res. 827, 28–40 (1999).

  38. 38.

    Kishioka, A. et al. A novel form of memory for auditory fear conditioning at a low-intensity unconditioned stimulus. PLoS One 4, e4157 (2009).

  39. 39.

    Paré, D. & Quirk, G. J. When scientific paradigms lead to tunnel vision: lessons from the study of fear. NPJ Sci. Learn. 2, 6 (2017).

  40. 40.

    Kita, H. & Kitai, S. T. Amygdaloid projections to the frontal cortex and the striatum in the rat. J. Comp. Neurol. 298, 40–49 (1990).

  41. 41.

    Loughlin, S. E. & Fallon, J. H. Dopaminergic and non-dopaminergic projections to amygdala from substantia nigra and ventral tegmental area. Brain Res. 262, 334–338 (1983).

  42. 42.

    Darvas, M. & Palmiter, R. D. Restriction of dopamine signaling to the dorsolateral striatum is sufficient for many cognitive behaviors. Proc. Natl. Acad. Sci. USA 106, 14664–14669 (2009).

  43. 43.

    Schiemann, J. et al. K-ATP channels in dopamine substantia nigra neurons control bursting and novelty-induced exploration. Nat. Neurosci. 15, 1272–1280 (2012).

  44. 44.

    Choi, J.-S. & Kim, J. J. Amygdala regulates risk of predation in rats foraging in a dynamic fear environment. Proc. Natl. Acad. Sci. USA 107, 21773–21777 (2010).

  45. 45.

    Ainsworth, M. D. & Bell, S. M. Attachment, exploration, and separation: illustrated by the behavior of one-year-olds in a strange situation. Child Dev. 41, 49–67 (1970).

  46. 46.

    Belin, D., Berson, N., Balado, E., Piazza, P. V. & Deroche-Gamonet, V. High-novelty-preference rats are predisposed to compulsive cocaine self-administration. Neuropsychopharmacology 36, 569–579 (2011).

  47. 47.

    Sasson, N. J., Turner-Brown, L. M., Holtzclaw, T. N., Lam, K. S. L. & Bodfish, J. W. Children with autism demonstrate circumscribed attention during passive viewing of complex social and nonsocial picture arrays. Autism Res. 1, 31–42 (2008).

  48. 48.

    Corey, D. T. The determinants of exploration and neophobia. Neurosci. Biobehav. Rev. 2, 235–253 (1978).

  49. 49.

    Waddell, S. Reinforcement signalling in Drosophila; dopamine does it all after all. Curr. Opin. Neurobiol. 23, 324–329 (2013).

  50. 50.

    Kim, H. F., Ghazizadeh, A. & Hikosaka, O. Dopamine neurons encoding long-term memory of object value for habitual behavior. Cell 163, 1165–1175 (2015).

  51. 51.

    Bäckman, C. M. et al. Characterization of a mouse strain expressing Cre recombinase from the 3′ untranslated region of the dopamine transporter locus. Genesis 44, 383–390 (2006).

  52. 52.

    Wickersham, I. R. et al. Monosynaptic restriction of transsynaptic tracing from single, genetically targeted neurons. Neuron 53, 639–647 (2007).

  53. 53.

    Chen, T.-W. et al. Ultrasensitive fluorescent proteins for imaging neuronal activity. Nature 499, 295–300 (2013).

  54. 54.

    Thiele, S.L., Warre, R. & Nash, J.E. Development of a unilaterally-lesioned 6-OHDA mouse model of Parkinson’s disease. J. Vis. Exp. https://doi.org/10.3791/3234 (2012).

  55. 55.

    Baldo, B. A., Daniel, R. A., Berridge, C. W. & Kelley, A. E. Overlapping distributions of orexin/hypocretin- and dopamine-β-hydroxylase immunoreactive fibers in rat brain regions mediating arousal, motivation, and stress. J. Comp. Neurol. 464, 220–237 (2003).

  56. 56.

    Gittis, A. H. et al. Rapid target-specific remodeling of fast-spiking inhibitory circuits after loss of dopamine. Neuron 71, 858–868 (2011).

  57. 57.

    Schallert, T., Fleming, S. M., Leasure, J. L., Tillerson, J. L. & Bland, S. T. CNS plasticity and assessment of forelimb sensorimotor outcome in unilateral rat models of stroke, cortical ablation, parkinsonism and spinal cord injury. Neuropharmacology 39, 777–787 (2000).

  58. 58.

    Athos, J. & Storm, D. R. High precision stereotaxic surgery in mice. Curr. Protoc. Neurosci. 4 Appendix, 4A (2001).

  59. 59.

    Uchida, N. & Mainen, Z. F. Speed and accuracy of olfactory discrimination in the rat. Nat. Neurosci. 6, 1224–1229 (2003).

  60. 60.

    Wang, A. Y., Miura, K. & Uchida, N. The dorsomedial striatum encodes net expected return, critical for energizing performance vigor. Nat. Neurosci. 16, 639–647 (2013).

  61. 61.

    Yizhar, O., Fenno, L. E., Davidson, T. J., Mogri, M. & Deisseroth, K. Optogenetics in neural systems. Neuron 71, 9–34 (2011).

  62. 62.

    Stujenske, J. M., Spellman, T. & Gordon, J. A. Modeling the spatiotemporal dynamics of light and heat propagation for in vivo optogenetics. Cell Rep. 12, 525–534 (2015).

  63. 63.

    Nakamura, K. & Hikosaka, O. Role of dopamine in the primate caudate nucleus in reward modulation of saccades. J. Neurosci. 26, 5360–5369 (2006).

  64. 64.

    Kudo, Y. et al. A single optical fiber fluorometric device for measurement of intracellular Ca2+concentration: its application to hippocampal neurons in vitro and in vivo. Neuroscience 50, 619–625 (1992).

  65. 65.

    Gunaydin, L. A. et al. Natural neural projection dynamics underlying social behavior. Cell 157, 1535–1551 (2014).

  66. 66.

    Kim, C. K. et al. Simultaneous fast measurement of circuit dynamics at multiple sites across the mammalian brain. Nat. Methods 13, 325–328 (2016).

  67. 67.

    Akerboom, J. et al. Optimization of a GCaMP calcium indicator for neural activity imaging. J. Neurosci. 32, 13819–13840 (2012).

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Acknowledgements

We thank F. Morgese, M. Tripodi, and E. Ciabatti at MRC Laboratory of Molecular Biology in Cambridge, UK, for the self-inactivating rabies virus amplification kit. We thank B. Lowell at Harvard University in Boston, MA, for the Vglut2flox mouse. We thank E. Boyden at MIT in Cambridge, MA, for pAAV-FLEX-GFP, K. Deisseroth at Stanford University in Stanford, CA, for pAAV-DIO-ChR2 and pAAV-fDIO-EYFP, and V. Jayaraman, R. Kerr, D. Kim, L. Looger, and K. Svoboda from the GENIE Project, Janelia Farm Research Campus, Howard Hughes Medical Institute for pGP-GCaMP6f and AAV-FLEX-GCaMP6m. We also appreciate C. Dulac and J. Assad for critically reading the manuscript. Finally, we thank H. Kim for help setting up the closed-loop optogenetic stimulation system. This work was supported by National Institutes of Health grants R01MH095953 (N.U.), R01MH101207 (N.U.), R01MH110404 (N.U.), Harvard Mind Brain and Behavior faculty grant (N.U.) and JSPS Overseas Research Fellowships (R.A.).

Author information

Affiliations

  1. Center for Brain Science, Department of Molecular and Cellular Biology, Harvard University, Cambridge, MA, USA

    • William Menegas
    • , Korleki Akiti
    • , Ryunosuke Amo
    • , Naoshige Uchida
    •  & Mitsuko Watabe-Uchida

Authors

  1. Search for William Menegas in:

  2. Search for Korleki Akiti in:

  3. Search for Ryunosuke Amo in:

  4. Search for Naoshige Uchida in:

  5. Search for Mitsuko Watabe-Uchida in:

Contributions

W.M.: conceptualization, investigation, formal analysis, writing (original draft); K.A.: methodology (novel object); R.A.: methodology (rabies virus); N.U.: conceptualization, supervision, writing (review and editing); M.W.-U.: conceptualization, supervision, writing (original draft).

Competing interests

The authors declare no competing interests.

Corresponding author

Correspondence to Mitsuko Watabe-Uchida.

Integrated supplementary information

  1. Supplementary Figure 1 Summary of GCaMP recording fiber implant sites.

    Location of optic fibers used to collect GCaMP signals in VS (top) and TS (bottom) in head fixed (Fig. 1) and freely moving (Fig. 7) animals. Reference slices are depicted with 0.5 mm spacing, and fiber positions are plotted on the nearest reference slice. The approximate location of the tip of each 200 µm diameter fiber is indicated (green). Light emitted by axons directly below these sites could be detected.

  2. Supplementary Figure 2 GCaMP normalization and GFP control signals.

    (a) Left: schematic of strategy for expressing GCaMP in dopamine axons (green) and collecting signal using an implanted optic fiber. Middle: example raw voltage traces (green), running median (blue), and normalized “dF/F” signal (black). Right: Comparison of dF/F signals (one line per session) sorted from smallest to largest dF/F value. (b) GFP control schematic (left), example signals (middle), and comparison across sessions (right). (c) Average signals following unexpected water delivery (left) or tone delivery (right) in GFP expressing control mice.

  3. Supplementary Figure 3 Responses in dopamine axons in TS scale with external stimulus novelty and intensity.

    Left: average responses to different tone sizes in dopamine axons in TS (top). Data was plotted in bins of 15 trials to demonstrate the effect of novelty (middle) and displayed as a heat map (bottom). Right: average responses to different air puff sizes in dopamine axons in TS (top). Data was plotted in bins of 15 trials (middle) and displayed as a heat map (bottom). Note that “trial number” refers to the total number of each stimulus rather than the number of stimuli of that intensity prior to that point. * p < 0.05, 1-way ANOVA. For tone: f(8) = 3.05, p = 0.0275, 1-way ANOVA. For air puff: f(10) = 4.93, p = 0.00081, 1-way ANOVA.

  4. Suppplementary Figure 4 Comparison of reward-omission signals in dopamine axons in VS and TS, and responses to external stimuli.

    (a) Top: trial structure for classical conditioning. Odor A is 90% followed by water (green, expected water) and 10% followed by nothing (red, omitted water). Odor B (black) is 100% followed by nothing. A small number ( < 5%) of all trials consisted of unexpected water (blue). Bottom: example single trial responses to unexpected water of different sizes simultaneously recorded from VS (cyan) and TS (magenta). (b) Example recording sites in VS (top) and TS (bottom). (c) Average responses across animals recorded from dopamine axons in VS (top) and TS (bottom). (d) Average difference (mean ± s.e.m. across n = 11 animals) between responses to expected and omitted water (expected minus omitted) across animals in dopamine axons in VS (top; t = 4.98, p = 0.00076, n = 11 animals, paired t-test) and TS (bottom; t = -0.83, p = 0.43, n = 11 animals, paired t-test). TS dopamine axon responses (mean ± s.e.m. across n = 11 animals) are also shown for (e) odor: t = 6.49, p = 0.0029, paired t-test, (f) tone: t = 5.057, p = 0.0072, paired t-test, (g) light: t = 2.83, p = 0.018, paired t-test, and (h) air puff: t = 5.82, p = 0.00025, paired t-test. For statistical tests, peak signals in a 1 second window during the inter-trial interval (“Baseline”) is compared to the peak signal in the 1 second window following stimulus presentation (red). Solid dots indicate mean and transparent dots indicate each animal. * P< 0.01, two-sided t-test.

  5. Supplementary Figure 5 Distribution of TS-projecting dopamine neurons in the midbrain.

    TS-projecting dopamine neurons labelled with rabies-GFP (Fig. 4). (a) Example midbrain section (approximately Bregma −3.10) from each injected animal. Left: dopamine neurons labelled with anti-TH antibody (magenta) and TS-projecting dopamine neurons labelled with rabies-GFP (green). Right: TS-projecting dopamine neurons labelled with rabies-GFP (green) and anterior striatum projecting dopamine neurons labeled with rabies-mCherry. (b) GFP-labelled neurons (TS-projecting dopamine neurons) from all animals plotted to summarize their distribution.

  6. Supplementary Figure 6 TS-projecting dopamine neurons in SNL respond to novel and high-intensity external stimuli.

    (a) Responses to a novel odor (black, first 5 presentations) compared to all presentations of that odor in TS-projecting dopamine neurons in SNL. (b) Responses to first 5 presentations of sucrose (black) compared to all presentations of sucrose (grey). (c) Responses to a loud tone (black, also see Fig. 1d) and a quiet tone (grey, also see Fig. 1d). (d) Responses to a high concentration of sucrose (black, 10%) compared to a low concentration of sucrose (grey, 2%). Solid dots indicate mean and transparent dots indicate each animal. Transparent error bars depict s.e.m. n= 6 animals.

  7. Supplementary Figure 7 Summary of ChR2 stimulation fiber sites.

    (a) Location of optic fiber implants used to stimulate ChR2-expressing dopamine axons in VS (top) and TS (bottom) in freely moving animals (Fig. 3). Reference slices are depicted with 0.5 mm spacing, and fiber positions are plotted on the nearest reference slice. The approximate location of the tip of each 200 µm diameter fiber is indicated (green). (b) Schematic of light spread from fiber. (c) Position of fibers relative to dorsal tip of amygdala. (d) Correlation between distance away from tip of amygdala and effect size (Fig. 3).

  8. Supplementary Figure 8 Example 6-OHDA histology.

    Example slices stained with anti-TH (green) antibody to label dopamine cell bodies and axons. (a) Axons of dopamine neurons (green) in an example intact animal (top) and lesioned animal (bottom) in VS, DMS, DLS, TS, and Ce. (b) Cell bodies of dopamine neurons (green) in an example intact animal (top) and lesioned animal (bottom) in VTA, SNC, SNL, and RRF. Also shown are TH-labelled noradrenaline neurons in LC (right). (c) Average fluorescence in dopamine axon target regions in intact (blue) and lesion (red) animals (mean ± s.e.m. across n = 20 per condition, each hemisphere quantified separately). For anterior TS: t = 3.86, p = 0.00054, n = 20 hemispheres per condition, unpaired t-test. For posterior TS: t = 7.20, p = 0.00000091, n = 20 hemispheres per condition, unpaired t-test. (d) Average cell body counts per section in intact (blue) and lesion (red) animals (mean ± s.e.m. across n = 20 per condition, each hemisphere quantified separately). For SNL: t = 4.61, p = 0.00049, n = 20 hemispheres per condition, unpaired t-test. VS: ventral striatum, DMS: dorsomedial striatum, DLS: dorsolateral striatum, aTS: anterior tail of striatum, pTS: posterior tail of striatum, Ce: central amygdala, BLA: basolateral amygdala, LHb: lateral habenula, VTA: ventral tegmental area, SNC: substantia nigra pars compacta, SNL: substantia nigra pars lateralis, RRF: retrorubral field, LC: locus coeruleus. ** P < 0.005, unpaired t-test.

  9. Supplementary Figure 9 Average velocity is not affected by 6-OHDA lesion.

    (a) Left: example cumulative center of mass traces for saline (blue) and lesion (red) mice during a 10-minute session. Right: example velocity plot for saline (blue) and lesion (red) mice during one minute of free movement. (b) Average time spent moving (top) and average velocity (bottom) of saline and lesion mice (mean + s.e.m. across n = 10 animals per condition) for first ten minutes of free movement For average velocity: Saline x 6OHDA: t = -0.45, p = 0.66, n = 10 animals per condition. Solid dots indicate mean and transparent dots indicate each animal. (c) Time spend in center (top) and maximum velocity (bottom) of saline and lesion mice (mean + s.e.m. across n = 10 animals per condition) during the first ten minutes of free movement. For maximum velocity: Saline x 6OHDA: t = -0.31, p = 0.76, n = 10 animals per condition). Solid dots indicate mean and transparent dots indicate each animal. All panels in this figure use data from mice not performing any choice task or engaging in novel object exploration.

  10. Supplementary Figure 10 Time-course of choice-bias, retreat, and TS dopamine axons signals elicited by airpuff in saline control and 6-OHDA lesion animals.

    (a) Time course of choice of water + puff in saline (blue) and lesion (red) animals. Here, trial number indicates the total trial number (note that Fig. 4 shows the time course of retreats based on the water + air puff choice number). (b) All choices from saline (blue) and lesion (red) animals are plotted by trial number. (c) Left: average stay (%) in saline and 6OHDA mice when choosing between ports of equal value (i.e. water only). Stay (%) defined as the fraction of trials in which mice did not switch choice ports on the next trial. Saline x 6-OHDA: t = 0.010, p = 0.99, n = 10 animals in each group, unpaired t-test. Right: average stay (%) in saline and 6OHDA mice when choosing between water (cyan) and water + air puff (magenta). Saline: t = 3.57, p = 0.0022, n = 10 animals, paired t-test. 6-OHDA: t = -0.54, p = 0.60, n = 10 animals, paired t-test. ** P < 0.005, paired t-test. n = 10 animals for each condition. (d) Retreats from air puff plotted as a function of total trial number. (e) Peak GCaMP responses to air puff in dopamine axons in TS as a function of total trial number. In all panels, solid dots indicate mean and transparent dots indicate each animal. Asterisks indicate that several animals did not choose air puff at all within this time bin, so an average could not be defined. For all time courses in this figure, data was analyzed in bins of 5 trials.

  11. Supplementary Figure 11 Example video clips of airpuff responses in saline control and 6-OHDA lesion animals.

    (a) Example of a control (saline) animal’s air puff response in early (top) and later (bottom) trials. (b) Example of a lesion (6OHDA) animal’s air puff response in early (top) and later (bottom) trials.

  12. Supplementary Figure 12 D1-antagonist in TS eliminates choice bias and modulates airpuff responses.

    (a) Average choice bias (mean ± s.e.m. across n = 6 animals) on days with saline injection with no air puff trials (black), with saline injection with air puff on one port (blue), and with D1 antagonist injection with air puff on one port (red) in the same set of cannula-implanted mice. Saline x D1 antagonist: t = 4.44, p = 0.0068, paired t-test. Solid dots indicate mean and transparent dots indicate each animal. (b) Examples of retreat distance over time for each condition. (c) Retreat distance (mean ± s.e.m. across n = 6 animals) on trial 1 (left; Saline x Saline + puff: f = 4.67, p = 0.00088, n = 6, paired t-test; Saline x D1 + puff: f = -2.88, p = 0.016, n = 6, paired t-test) and trial 1-10 (right; Saline x Saline + puff: f = 2.51, p = 0.031, n = 6, paired t-test, Saline + puff x D1 + puff: f = 2.23, p = 0.045, n = 6, paired t-test) for each condition. Solid dots indicate mean and transparent dots indicate each animal. (d) Schematic of choice task. (e) Average velocity (left; Saline x D1: t = 1.72, p = 0.12, n = 6, paired t-test) and time spent moving (right; Saline x D1: t = 1.09, p = 0.30, n = 6, paired t-test) while mice engage in the choice task (mean + s.e.m. across n = 6 animals). Solid dots indicate mean and transparent dots indicate each animal. (f) Example center of mass traces from mice engaged in the choice task. * P < 0.01, two-tailed t-test.

  13. Supplementary Figure 13 Example video clips of novel object investigation in saline control and 6-OHDA lesion animals.

    (a) Example of a control (saline) animal’s early (top) and later (bottom) investigation of a novel object. (b) Example of a lesion (6OHDA) animal’s early (top) and later (bottom) investigation of a novel object.

  14. Supplementary Figure 14 Responses in dopamine axons in TS are not locked to initiation of movement.

    (a) Example velocity traces (grey) and GCaMP traces (green) as animals move freely within a behavior box (not performing a task or interacting with a novel object). Three 10 second example traces are shown. (b) Animal velocity was aligned to movement initiation and separated based on the minimum gain of velocity (top). Corresponding GCaMP traces aligned to movement initiation (middle). Quantification of peak GCaMP signal for movements of each size (bottom). (c) Top: bouts of novel object exploration were aligned to the nearest point of investigation. Middle: average GCaMP (green) and GFP control (grey) traces locked to retreat from novel object. Bottom: GCaMP signals (green) and GFP signals (grey) locked to retreat from novel object (average + s.e.m. across n = 6 animals).

  15. Supplementary Figure 15 Time-course of responses to novel objects in dopamine axons in TS; removing Vglut2 from dopamine neurons does not diminish avoidance of novel objects.

    (a) Responses in dopamine axons in TS plotted by bout (i.e. trial) for the first 30 interactions with a novel object. Solid dots indicate mean and transparent dots indicate each animal. (b) Control animal that does not express DAT-cre, meaning that Vglut2 is not knocked out in any neurons (black), and knock out animal expressing DAT-cre to specifically knock out Vglut2 in dopamine neurons (magenta). (c) Example center of mass traces of control (black) and knock-out (magenta) mice not performing any task or engaging in novel object interaction. Right: average velocity (Control x Vglut2 -/-: t = 1.16, p = 0.26, n = 8 animals per group, unpaired t-test). Solid dots indicate mean and transparent dots indicate each animal. (d) Example center of mass traces of control (black) and knock-out (magenta) mice interacting with a novel object. Right: average fraction of time spent near object in a session (Control x Vglut2 -/-: t = -0.31, p = 0.76, n = 8 animals per group, unpaired t-test). Solid dots indicate mean and transparent dots indicate each animal.

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https://doi.org/10.1038/s41593-018-0222-1

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