Final Report Summary - MNEMOSMELL (Unraveling the mechanisms of odor coding and sent-tracking in Drosophila larvae)
Our senses allow us to collect external signals from the environment and to convert them into coherent internal representations. Responses elicited by the internal representations of the brain form the basis of adaptive behaviours. The goal of MNEMOSMELL was to clarify the neural processes directing sensory perception and orientation behaviours. To tackle this problem, we used a simple behavioural paradigm: chemotaxis in the drosophila melanogaster larva. Chemotaxis involves movements towards attractive stimuli and away from aversive stimuli. Despite the good anatomical characterisation of the larval olfactory system, there is still no comprehensive model explaining how complex odourant stimuli are encoded in the peripheral olfactory systems of the fly and how changes in concentration are integrated to direct chemotaxis.
To address these questions, we followed an interdisciplinary approach combining electrophysiology, behavioural analysis and computational modelling. The first aim of MNEMOSMELL was to understand how naturalistic odourant stimuli are encoded and processed by the olfactory system. Using optogenetics, we created virtual olfactory signals to test hypotheses about the mechanisms controlling the integration of sensory input into behavioural output. The second aim of MNEMOSMELL was to identify neural circuits involved in the processing of olfactory information and the making of orientation decisions during chemotaxis. The research carried out in this project advances our understanding of how a simple brain integrates sensory information to control behaviour.
Peripheral encoding of dynamical olfactory stimuli
The larval 'nose' is composed of 21 olfactory sensory neurons (OSNs) expressing one type of odourant receptor along with a ubiquitously expressed co-receptor (Orco). Individual odourant receptors have overlapping, yet distinct, ligand tuning properties. Accordingly, each OSN can be viewed as a distinct information channel into the fly olfactory system. We devised a protocol to carry out single-unit extracellular recordings from the larval olfactory system to define how odours are represented by dynamical patterns of activity originating from the OSNs and transmitted to higher brain centres. Using a customised odour delivery system, we recreated the temporal changes in odour concentration that are typically experienced by a larva during chemotaxis in odour gradients. We systematically characterised the response of a single OSN to concentration time courses corresponding to episodes of behaviour in odour gradients. We discovered that the activity pattern of a single OSN captures high-order features associated with temporal changes in the stimulus intensity (e.g. first and second derivatives). This finding reveals the surprisingly high degree of signal processing that takes place in the first-order neurons of the larval olfactory system.
Navigational algorithm controlling larval chemotaxis
Bacteria navigate in chemical gradients according to an indirect orientation mechanism that consists in a biased random walk. In contrast, drosophila larvae employ a direct orientation mechanism where motion is directed toward the local odour gradient. Having ruled out an orientation mechanism purely based on stereo-olfaction (comparisons between bilateral olfactory inputs), we hypothesised that larvae rely on active sampling where lateral head movements ('head casts') permit the sampling odour intensities in space. To validate this hypothesis, we developed a computer-vision algorithm for high-resolution tracking. This tool enabled us to extract morphological and kinetic features describing the behavioural state of a larva. In addition, we combined real-time tracking and optogenetics to induce predictable behavioural time courses.
In collaboration with Janelia Farm, we built a fully automated tracking system that can resolve the posture and kinetic properties of a larva at a rate of 30 Hz. The tracking system is outfitted with a set of light emitting diodes (LEDs) that can be activated according to the behavioural history of the larva (head casting, running or turning, etc). We expressed light-gated ion channel, channelrhodopsin-2, in one targeted OSN to manipulate the olfactory inputs experienced by a freely moving larva. Using this approach, we investigated the computational bases of the spatiotemporal integration during orientation behaviour by reverse-engineering odour evoked activity patterns with light. By controlling temporal and spatial aspects of the olfactory stimulus in virtual realities, we manipulated the onset of turns in a predictable way by simulating down-the-gradient and up-the-gradient experiences. We have elaborated a mathematical model that accurately predicts the OSN activity elicited by dynamical olfactory signals and the ensuing behavioural responses.
New circuits participating in the control of chemotaxis
Larvae use a strategy based on active sampling to track odours. During forward runs and lateral head casts, larvae are capable of inferring the direction of the local odour gradient. How are these decisions made? To identify the circuits participating in this process, we carried out a large-scale behavioural screen. We took advantage of the Gal4-drivers of the Kyoto collection to study the effects of disabling sub-regions of the larval brain by genetically expressing the tetanus toxin light chain (inhibitor of synaptic transmission). While specific neurons were silenced, we tested the orientation capabilities of mutants to directional sensory stimuli. We identified several lines with strong phenotypic defects and sparse expression patterns. We decided to concentrate on the two most promising lines. The first one shows altered integration of odour information during forward locomotion, which leads to a defective initiation of reorientation manoeuvres. The second is severely impaired in the execution of active sampling and shows prolonged run episodes. By means of controlled gain-of-function experiments, we investigated the potential contribution of neurons located in the sub-oesophagus ganglion and Mushroom bodies in the modulation of run persistency and the execution of head casts during chemotaxis. We are currently conducting a physiological characterisation of these neurons to clarify their function in the processing of olfactory inputs.
Outlook
The navigational strategy employed by larvae to orient in odour gradients bears striking similarities with the trail-tracking behaviour of vertebrates. In spite of their high evolutionary divergence, drosophila larvae, rats and dogs track odours by actively sampling their odorant environment through lateral head movements. Orientation results from the differentiation and the temporal integration of the sensory stimulus. In the MNEMOSMELL project, we exploited the genetic tools available in the drosophila larva to advance of understanding of the molecular and cellular mechanisms underlying the sensorimotor integration of naturalistic olfactory signals.
References related to the work of the lab
Gomez-Marin, A. and M. Louis (2012). 'Active sensation during orientation behavior in the Drosophila larva: more sense than luck.' Curr Opin Neurobiol 22(2): 208-215.
Gomez-Marin, A., N. Partoune, et al. (2012). 'Automated Tracking of Animal Posture and Movement during Exploration and Sensory Orientation Behaviors.' PLoS One 7(8): e41642.
Gomez-Marin, A., G. J. Stephens, et al. (2011). 'Active sampling and decision making in Drosophila chemotaxis.' Nat Commun 2: 441.
Louis, M., T. Huber, et al. (2008). 'Bilateral olfactory sensory input enhances chemotaxis behavior.' Nat Neurosci 11(2): 187-199.
To address these questions, we followed an interdisciplinary approach combining electrophysiology, behavioural analysis and computational modelling. The first aim of MNEMOSMELL was to understand how naturalistic odourant stimuli are encoded and processed by the olfactory system. Using optogenetics, we created virtual olfactory signals to test hypotheses about the mechanisms controlling the integration of sensory input into behavioural output. The second aim of MNEMOSMELL was to identify neural circuits involved in the processing of olfactory information and the making of orientation decisions during chemotaxis. The research carried out in this project advances our understanding of how a simple brain integrates sensory information to control behaviour.
Peripheral encoding of dynamical olfactory stimuli
The larval 'nose' is composed of 21 olfactory sensory neurons (OSNs) expressing one type of odourant receptor along with a ubiquitously expressed co-receptor (Orco). Individual odourant receptors have overlapping, yet distinct, ligand tuning properties. Accordingly, each OSN can be viewed as a distinct information channel into the fly olfactory system. We devised a protocol to carry out single-unit extracellular recordings from the larval olfactory system to define how odours are represented by dynamical patterns of activity originating from the OSNs and transmitted to higher brain centres. Using a customised odour delivery system, we recreated the temporal changes in odour concentration that are typically experienced by a larva during chemotaxis in odour gradients. We systematically characterised the response of a single OSN to concentration time courses corresponding to episodes of behaviour in odour gradients. We discovered that the activity pattern of a single OSN captures high-order features associated with temporal changes in the stimulus intensity (e.g. first and second derivatives). This finding reveals the surprisingly high degree of signal processing that takes place in the first-order neurons of the larval olfactory system.
Navigational algorithm controlling larval chemotaxis
Bacteria navigate in chemical gradients according to an indirect orientation mechanism that consists in a biased random walk. In contrast, drosophila larvae employ a direct orientation mechanism where motion is directed toward the local odour gradient. Having ruled out an orientation mechanism purely based on stereo-olfaction (comparisons between bilateral olfactory inputs), we hypothesised that larvae rely on active sampling where lateral head movements ('head casts') permit the sampling odour intensities in space. To validate this hypothesis, we developed a computer-vision algorithm for high-resolution tracking. This tool enabled us to extract morphological and kinetic features describing the behavioural state of a larva. In addition, we combined real-time tracking and optogenetics to induce predictable behavioural time courses.
In collaboration with Janelia Farm, we built a fully automated tracking system that can resolve the posture and kinetic properties of a larva at a rate of 30 Hz. The tracking system is outfitted with a set of light emitting diodes (LEDs) that can be activated according to the behavioural history of the larva (head casting, running or turning, etc). We expressed light-gated ion channel, channelrhodopsin-2, in one targeted OSN to manipulate the olfactory inputs experienced by a freely moving larva. Using this approach, we investigated the computational bases of the spatiotemporal integration during orientation behaviour by reverse-engineering odour evoked activity patterns with light. By controlling temporal and spatial aspects of the olfactory stimulus in virtual realities, we manipulated the onset of turns in a predictable way by simulating down-the-gradient and up-the-gradient experiences. We have elaborated a mathematical model that accurately predicts the OSN activity elicited by dynamical olfactory signals and the ensuing behavioural responses.
New circuits participating in the control of chemotaxis
Larvae use a strategy based on active sampling to track odours. During forward runs and lateral head casts, larvae are capable of inferring the direction of the local odour gradient. How are these decisions made? To identify the circuits participating in this process, we carried out a large-scale behavioural screen. We took advantage of the Gal4-drivers of the Kyoto collection to study the effects of disabling sub-regions of the larval brain by genetically expressing the tetanus toxin light chain (inhibitor of synaptic transmission). While specific neurons were silenced, we tested the orientation capabilities of mutants to directional sensory stimuli. We identified several lines with strong phenotypic defects and sparse expression patterns. We decided to concentrate on the two most promising lines. The first one shows altered integration of odour information during forward locomotion, which leads to a defective initiation of reorientation manoeuvres. The second is severely impaired in the execution of active sampling and shows prolonged run episodes. By means of controlled gain-of-function experiments, we investigated the potential contribution of neurons located in the sub-oesophagus ganglion and Mushroom bodies in the modulation of run persistency and the execution of head casts during chemotaxis. We are currently conducting a physiological characterisation of these neurons to clarify their function in the processing of olfactory inputs.
Outlook
The navigational strategy employed by larvae to orient in odour gradients bears striking similarities with the trail-tracking behaviour of vertebrates. In spite of their high evolutionary divergence, drosophila larvae, rats and dogs track odours by actively sampling their odorant environment through lateral head movements. Orientation results from the differentiation and the temporal integration of the sensory stimulus. In the MNEMOSMELL project, we exploited the genetic tools available in the drosophila larva to advance of understanding of the molecular and cellular mechanisms underlying the sensorimotor integration of naturalistic olfactory signals.
References related to the work of the lab
Gomez-Marin, A. and M. Louis (2012). 'Active sensation during orientation behavior in the Drosophila larva: more sense than luck.' Curr Opin Neurobiol 22(2): 208-215.
Gomez-Marin, A., N. Partoune, et al. (2012). 'Automated Tracking of Animal Posture and Movement during Exploration and Sensory Orientation Behaviors.' PLoS One 7(8): e41642.
Gomez-Marin, A., G. J. Stephens, et al. (2011). 'Active sampling and decision making in Drosophila chemotaxis.' Nat Commun 2: 441.
Louis, M., T. Huber, et al. (2008). 'Bilateral olfactory sensory input enhances chemotaxis behavior.' Nat Neurosci 11(2): 187-199.
