Functional architecture of reward learning in mushroom body extrinsic neurons of larval Drosophila

The brain adaptively integrates present sensory input, past experience, and options for future action. The insect mushroom body exemplifies how a central brain structure brings about such integration. Here we use a combination of systematic single-cell labeling, connectomics, transgenic silencing, and activation experiments to study the mushroom body at single-cell resolution, focusing on the behavioral architecture of its input and output neurons (MBINs and MBONs), and of the mushroom body intrinsic APL neuron. Our results reveal the identity and morphology of almost all of these 44 neurons in stage 3 Drosophila larvae. Upon an initial screen, functional analyses focusing on the mushroom body medial lobe uncover sparse and specific functions of its dopaminergic MBINs, its MBONs, and of the GABAergic APL neuron across three behavioral tasks, namely odor preference, taste preference, and associative learning between odor and taste. Our results thus provide a cellular-resolution study case of how brains organize behavior.The mushroom body of Drosophila integrates sensory information with past experience to guide behaviour. Here, the authors provide an atlas of the input and output neurons of the stage 3 larval mushroom body at the single-cell level, and analyse their function in learned and innate behaviours.

[1]  B. Gerber,et al.  Generalization and discrimination tasks yield concordant measures of perceived distance between odours and their binary mixtures in larval Drosophila , 2014, Journal of Experimental Biology.

[2]  William R. Stauffer,et al.  Dopamine prediction error responses integrate subjective value from different reward dimensions , 2014, Proceedings of the National Academy of Sciences.

[3]  G. Technau,et al.  Origin of Drosophila mushroom body neuroblasts and generation of divergent embryonic lineages , 2012, Development.

[4]  R. Malaka Models of classical conditioning , 1999 .

[5]  G. Nagel,et al.  Channelrhodopsin-2–XXL, a powerful optogenetic tool for low-light applications , 2014, Proceedings of the National Academy of Sciences.

[6]  T. Abrams,et al.  Temporal asymmetry in activation of Aplysia adenylyl cyclase by calcium and transmitter may explain temporal requirements of conditioning. , 1992, Proceedings of the National Academy of Sciences of the United States of America.

[7]  L. Abbott,et al.  Extending the effects of spike-timing-dependent plasticity to behavioral timescales. , 2006, Proceedings of the National Academy of Sciences of the United States of America.

[8]  R. Menzel The insect mushroom body, an experience-dependent recoding device , 2014, Journal of Physiology-Paris.

[9]  Andreas S. Thum,et al.  The neuronal and molecular basis of quinine-dependent bitter taste signaling in Drosophila larvae , 2014, Front. Behav. Neurosci..

[10]  Yoshinori Aso,et al.  Dopaminergic neurons write and update memories with cell-type-specific rules , 2016, eLife.

[11]  Bertram Gerber,et al.  Outcome expectations drive learned behaviour in larval Drosophila , 2006, Proceedings of the Royal Society B: Biological Sciences.

[12]  G. Rubin,et al.  Mushroom body efferent neurons responsible for aversive olfactory memory retrieval in Drosophila , 2011, Nature Neuroscience.

[13]  A. Fiala,et al.  Differential Associative Training Enhances Olfactory Acuity in Drosophila melanogaster , 2014, The Journal of Neuroscience.

[14]  Irina Sinakevitch,et al.  Ground plan of the insect mushroom body: Functional and evolutionary implications , 2009, The Journal of comparative neurology.

[15]  M. Gabriel,et al.  Learning and Computational Neuroscience: Foundations of Adaptive Networks , 1990 .

[16]  Marta Zlatic,et al.  Four Individually Identified Paired Dopamine Neurons Signal Reward in Larval Drosophila , 2016, Current Biology.

[17]  M. Hammer An identified neuron mediates the unconditioned stimulus in associative olfactory learning in honeybees , 1993, Nature.

[18]  Haojiang Luan,et al.  Refined Spatial Manipulation of Neuronal Function by Combinatorial Restriction of Transgene Expression , 2006, Neuron.

[19]  S. Waddell,et al.  Remembering Components of Food in Drosophila , 2016, Front. Integr. Neurosci..

[20]  G. Rubin,et al.  The neuronal architecture of the mushroom body provides a logic for associative learning , 2014, eLife.

[21]  Jeffrey A. Riffell,et al.  The neurobiology of insect olfaction: Sensory processing in a comparative context , 2011, Progress in Neurobiology.

[22]  T. Tanimura,et al.  Learning the specific quality of taste reinforcement in larval Drosophila , 2015, eLife.

[23]  Andreas S. Thum,et al.  Diversity, variability, and suboesophageal connectivity of antennal lobe neurons in D. melanogaster larvae , 2011, The Journal of comparative neurology.

[24]  B. Gerber,et al.  Cellular site and molecular mode of synapsin action in associative learning. , 2011, Learning & memory.

[25]  B. Gerber,et al.  A molecular and neuronal basis for amino acid sensing in the Drosophila larva , 2016, Scientific Reports.

[26]  G. Nagel,et al.  Light-Induced Activation of Distinct Modulatory Neurons Triggers Appetitive or Aversive Learning in Drosophila Larvae , 2006, Current Biology.

[27]  A. Guo,et al.  A GABAergic Inhibitory Neural Circuit Regulates Visual Reversal Learning in Drosophila , 2012, The Journal of Neuroscience.

[28]  Kei Ito,et al.  Systematic Analysis of Neural Projections Reveals Clonal Composition of the Drosophila Brain , 2013, Current Biology.

[29]  Andreas S. Thum,et al.  A map of sensilla and neurons in the taste system of drosophila larvae , 2017, The Journal of comparative neurology.

[30]  Bertram Gerber,et al.  Olfactory learning in individually assayed Drosophila larvae. , 2003, Learning & memory.

[31]  Aljoscha Nern,et al.  Optimized tools for multicolor stochastic labeling reveal diverse stereotyped cell arrangements in the fly visual system , 2015, Proceedings of the National Academy of Sciences.

[32]  Andreas S. Thum,et al.  Characterization of the octopaminergic and tyraminergic neurons in the central brain of Drosophila larvae , 2014, The Journal of comparative neurology.

[33]  R. Menzel,et al.  Mushroom body extrinsic neurons in the honeybee (Apis mellifera) brain integrate context and cue values upon attentional stimulus selection. , 2015, Journal of neurophysiology.

[34]  Omotara Ogundeyi,et al.  A GAL4 driver resource for developmental and behavioral studies on the larval CNS of Drosophila. , 2014, Cell reports.

[35]  Simon G. Sprecher,et al.  The Serotonergic Central Nervous System of the Drosophila Larva: Anatomy and Behavioral Function , 2012, PloS one.

[36]  Fabio Benfenati,et al.  Pavlovian Conditioning of Larval Drosophila: An Illustrated, Multilingual, Hands-On Manual for Odor-Taste Associative Learning in Maggots , 2017, Front. Behav. Neurosci..

[37]  Y. Hotta,et al.  Proliferation pattern of postembryonic neuroblasts in the brain of Drosophila melanogaster. , 1992, Developmental biology.

[38]  Leslie C Griffith,et al.  A single pair of neurons links sleep to memory consolidation in Drosophila melanogaster , 2015, eLife.

[39]  Bertram Gerber,et al.  Adaptive Adjustment of the Generalization-Discrimination Balance in Larval Drosophila , 2010, Journal of neurogenetics.

[40]  Evren Pamir,et al.  Common microbehavioral "footprint" of two distinct classes of conditioned aversion. , 2017, Learning & memory.

[41]  Gerald M. Rubin,et al.  A Higher Brain Circuit for Immediate Integration of Conflicting Sensory Information in Drosophila , 2015, Current Biology.

[42]  Kei Ito,et al.  A single GABAergic neuron mediates feedback of odor-evoked signals in the mushroom body of larval Drosophila , 2014, Front. Neural Circuits.

[43]  P. Tobler,et al.  Identity-specific coding of future rewards in the human orbitofrontal cortex , 2015, Proceedings of the National Academy of Sciences.

[44]  B. Grünewald,et al.  Morphology of feedback neurons in the mushroom body of the honeybee, Apis mellifera , 1999, The Journal of comparative neurology.

[45]  Michael Bate,et al.  Altered Electrical Properties in DrosophilaNeurons Developing without Synaptic Transmission , 2001, The Journal of Neuroscience.

[46]  T. Miyamoto,et al.  The Molecular Basis of Sugar Sensing in Drosophila Larvae , 2013, Current Biology.

[47]  R. Stocker,et al.  A central neural circuit for experience-independent olfactory and courtship behavior in Drosophila melanogaster , 2001, Proceedings of the National Academy of Sciences of the United States of America.

[48]  Thomas Preat,et al.  PKA Dynamics in a Drosophila Learning Center: Coincidence Detection by Rutabaga Adenylyl Cyclase and Spatial Regulation by Dunce Phosphodiesterase , 2010, Neuron.

[49]  Stefan R. Pulver,et al.  Independent Optical Excitation of Distinct Neural Populations , 2014, Nature Methods.

[50]  V. Hartenstein,et al.  The development of the Drosophila larval brain. , 2008, Advances in experimental medicine and biology.

[51]  Johannes Felsenberg,et al.  Activity of Defined Mushroom Body Output Neurons Underlies Learned Olfactory Behavior in Drosophila , 2015, Neuron.

[52]  Ronald L. Davis,et al.  Olfactory Learning , 2004, Neuron.

[53]  Edward M. Reingold,et al.  Graph drawing by force‐directed placement , 1991, Softw. Pract. Exp..

[54]  G. Rubin,et al.  A subset of dopamine neurons signals reward for odour memory in Drosophila , 2012, Nature.

[55]  B. Gerber,et al.  A Behavioral Odor Similarity “Space” in Larval Drosophila , 2011, Chemical senses.

[56]  M. Heisenberg,et al.  Dopamine and Octopamine Differentiate between Aversive and Appetitive Olfactory Memories in Drosophila , 2003, The Journal of Neuroscience.

[57]  Feng Li,et al.  The complete connectome of a learning and memory centre in an insect brain , 2017, Nature.

[58]  G. Rubin,et al.  Refinement of Tools for Targeted Gene Expression in Drosophila , 2010, Genetics.

[59]  K. Han,et al.  D1 Dopamine Receptor dDA1 Is Required in the Mushroom Body Neurons for Aversive and Appetitive Learning in Drosophila , 2007, The Journal of Neuroscience.

[60]  Ronald L. Davis,et al.  Dynamics of Learning-Related cAMP Signaling and Stimulus Integration in the Drosophila Olfactory Pathway , 2009, Neuron.

[61]  Andreas S. Thum,et al.  Smelling, tasting, learning: Drosophila as a study case. , 2009, Results and problems in cell differentiation.

[62]  Vikram Chandra,et al.  Neural correlates of water reward in thirsty Drosophila , 2014, Nature Neuroscience.

[63]  M. Heisenberg,et al.  Experimental psychology: Event timing turns punishment to reward , 2004, Nature.

[64]  Bertram Gerber,et al.  The Drosophila larva as a model for studying chemosensation and chemosensory learning: a review. , 2007, Chemical senses.

[65]  M. Heisenberg Mushroom body memoir: from maps to models , 2003, Nature Reviews Neuroscience.

[66]  Casey M. Schneider-Mizell,et al.  Synaptic transmission parallels neuromodulation in a central food-intake circuit , 2016, bioRxiv.

[67]  A. Dahanukar,et al.  Molecular neurobiology of Drosophila taste , 2015, Current Opinion in Neurobiology.

[68]  Mark Stopfer,et al.  A Temporal Channel for Information in Sparse Sensory Coding , 2014, Current Biology.

[69]  A. Guo,et al.  The GABAergic anterior paired lateral neurons facilitate olfactory reversal learning in Drosophila. , 2012, Learning & memory.

[70]  L. Vosshall,et al.  Molecular architecture of smell and taste in Drosophila. , 2007, Annual review of neuroscience.

[71]  Hiromu Tanimoto,et al.  Two pairs of mushroom body efferent neurons are required for appetitive long-term memory retrieval in Drosophila. , 2013, Cell reports.

[72]  N. K. Tanaka,et al.  Stereotypic and random patterns of connectivity in the larval mushroom body calyx of Drosophila. , 2005, Proceedings of the National Academy of Sciences of the United States of America.

[73]  T. Kitamoto Conditional modification of behavior in Drosophila by targeted expression of a temperature-sensitive shibire allele in defined neurons. , 2001, Journal of neurobiology.

[74]  M. Hammer,et al.  Backward inhibitory learning in honeybees: a behavioral analysis of reinforcement processing. , 1998, Learning & memory.

[75]  Tzumin Lee,et al.  Serotonin–mushroom body circuit modulating the formation of anesthesia-resistant memory in Drosophila , 2011, Proceedings of the National Academy of Sciences.

[76]  Bertram Gerber,et al.  Maggot learning and Synapsin function , 2013, Journal of Experimental Biology.

[77]  KyeongJin Kang,et al.  Behavioral Analysis of Bitter Taste Perception in Drosophila Larvae. , 2016, Chemical senses.

[78]  Daryl M. Gohl,et al.  Layered reward signaling through octopamine and dopamine in Drosophila , 2012, Nature.

[79]  A. Fiala,et al.  Induction of aversive learning through thermogenetic activation of Kenyon cell ensembles in Drosophila , 2014, Front. Behav. Neurosci..

[80]  Andreas V. M. Herz,et al.  Event Timing in Associative Learning: From Biochemical Reaction Dynamics to Behavioural Observations , 2012, PloS one.

[81]  John R. Carlson,et al.  Molecular and Cellular Organization of the Taste System in the Drosophila Larva , 2011, The Journal of Neuroscience.

[82]  G. Rubin,et al.  Neuron hemilineages provide the functional ground plan for the Drosophila ventral nervous system , 2015, eLife.

[83]  Aravinthan D. T. Samuel,et al.  The wiring diagram of a glomerular olfactory system , 2016, bioRxiv.

[84]  Andreas S. Thum,et al.  Taste processing in Drosophila larvae , 2015, Frontiers in Integrative Neuroscience.

[85]  G. Rubin,et al.  Mushroom body output neurons encode valence and guide memory-based action selection in Drosophila , 2014, eLife.

[86]  Nils Otto,et al.  The Ol1mpiad: concordance of behavioural faculties of stage 1 and stage 3 Drosophila larvae , 2017, Journal of Experimental Biology.

[87]  Scott Waddell,et al.  Olfactory learning skews mushroom body output pathways to steer behavioral choice in Drosophila , 2015, Current Opinion in Neurobiology.

[88]  Bertram Gerber,et al.  Innate Attractiveness and Associative Learnability of Odors Can Be Dissociated in Larval Drosophila , 2011, Chemical senses.

[89]  M. Pankratz,et al.  Central relay of bitter taste to the protocerebrum by peptidergic interneurons in the Drosophila brain , 2016, Nature Communications.

[90]  G. Laurent,et al.  Hebbian STDP in mushroom bodies facilitates the synchronous flow of olfactory information in locusts , 2007, Nature.

[91]  Bertram Gerber,et al.  A behavior-based circuit model of how outcome expectations organize learned behavior in larval Drosophila. , 2011, Learning & memory.

[92]  H. Tanimoto,et al.  Reversing Stimulus Timing in Visual Conditioning Leads to Memories with Opposite Valence in Drosophila , 2015, PloS one.

[93]  W. Quinn,et al.  The amnesiac Gene Product Is Expressed in Two Neurons in the Drosophila Brain that Are Critical for Memory , 2000, Cell.

[94]  R. Stocker,et al.  Architecture of the primary taste center of Drosophila melanogaster larvae , 2007, The Journal of comparative neurology.

[95]  Raphael Cohn,et al.  Coordinated and Compartmentalized Neuromodulation Shapes Sensory Processing in Drosophila , 2015, Cell.

[96]  Andreas S. Thum,et al.  Drosophila Larvae Establish Appetitive Olfactory Memories via Mushroom Body Neurons of Embryonic Origin , 2010, The Journal of Neuroscience.

[97]  M. Fendt,et al.  Pain-relief learning in flies, rats, and man: basic research and applied perspectives , 2014, Learning & memory.

[98]  Andreas S. Thum,et al.  The Role of Dopamine in Drosophila Larval Classical Olfactory Conditioning , 2009, PloS one.

[99]  Ronald L. Davis,et al.  The GABAergic anterior paired lateral neuron suppresses and is suppressed by olfactory learning , 2008, Nature Neuroscience.

[100]  Gerald M. Rubin,et al.  Heterosynaptic Plasticity Underlies Aversive Olfactory Learning in Drosophila , 2015, Neuron.

[101]  Jason Sih-Yu Lai,et al.  Heterotypic Gap Junctions between Two Neurons in the Drosophila Brain Are Critical for Memory , 2011, Current Biology.

[102]  Yoshinori Aso,et al.  Three Dopamine Pathways Induce Aversive Odor Memories with Different Stability , 2012, PLoS genetics.