Mechanisms and Function of Neural Synchronization in an Insect Olfactory System

One of the fundamental questions in modem integrative neurobiology relates to the encoding of sensory information by populations of neurons, and to the significance of this activity for perception, learning, memory and behavior. Synchronization of activity across a population of neurons has been observed many times over, but has never been demonstrated to be a necessary component of this coding process. Neural synchronization has been found in many brain areas in animals across several phyla, from molluscs to mammals. Studies in mammals have correlated the degree of neural synchronization with specific behavioral or cognitive states, such as sensorimotor tasks, segmentation and binocular rivalry suggesting a functional link. In the locust olfactory system, oscillatory synchronization is a prominent feature of the odor-evoked neural activity. Stimulation of the antenna by odors evokes synchronized firing in dynamic and odor-specific ensembles of the projection neurons of the antennal lobe, the principal neurons of the first-order olfactory relay in insects. The coherent activity of these projection neurons underlies an odor-evoked oscillatory field potential which can be recorded in the mushroom body, the second-order olfactory relay to which they project. In this dissertation, we investigated two important questions raised by these findings: how are such stimulus-evoked synchronous ensembles generated, and what is their functional significance? To address these questions, we performed electrophysiological experiments and recorded odor responses from neurons of the antennal lobes and mushroom bodies of locusts, in vivo and using natural odor stimulation in an unanesthetized, semi-intact preparation. We demonstrated the critical mechanism involved in neural synchronization of the antennal lobe neurons. The synchronization of the projection neurons relies critically on fast GABA (γ-aminobutyric acid) -mediated inhibition from the local interneurons. Projection neuron synchronization could be selectively blocked by local injection of the GABA receptor antagonist, picrotoxin. Picrotoxin spared the odor-specific, slow modulation of individual projection neuron responses, but desynchronized the firing of the odor-activated projection neuron assemblies. The oscillatory activity of the local intemeurons was also blocked by picrotoxin, which indicates that such activity depends on network synaptic dynamics. We also showed that the mushroom body networks are capable of generating oscillatory behavior of a similar frequency as that of its projection neuron inputs, and that they may thus be "tuned" to accept synchronized, oscillatory inputs of that frequency range. Our understanding of this mechanism, in tum, made possible the functional investigation of neural synchronization by selective disruption of projection neuron synchronization. We studied a population of neurons downstream from the antennal lobe projection neurons, the extrinsic neurons of the β-lobe of the mushroom body (βLNs). These βLNs were chosen for investigation because they were found to be odor-responsive and because their position in the olfactory pathway makes them a suitable "read-out" of population activity in the antennal lobe. We characterized βLN odor responses before and after selective disruption of the synchronization of the projection neuron ensembles with local picrotoxin injection into the antennal lobe. We showed that the tuning of these βLN responses was altered by PN desynchronization by changing existing responses and inducing new responses. This alteration in tuning resulted in a significant loss of odor specificity in individual βLN responses, an effect that never occurred in the responses of individual, desynchronized projection neurons. We thus propose that neural synchronization is indeed important for information processing in the brain: it serves, at least in part, as a temporal substrate for the transmission of information that is contained across co-activated neurons (relational code) early in the pathway.</p

Behavioral and Neurophysiological Study of Olfactory Perception and Learning in Honeybees

The honeybee Apis mellifera has been a central insect model in the study of olfactory perception and learning for more than a century, starting with pioneer work by Karl von Frisch. Research on olfaction in honeybees has greatly benefited from the advent of a range of behavioral and neurophysiological paradigms in the Lab. Here I review major findings about how the honeybee brain detects, processes, and learns odors, based on behavioral, neuroanatomical, and neurophysiological approaches. I first address the behavioral study of olfactory learning, from experiments on free-flying workers visiting artificial flowers to laboratory-based conditioning protocols on restrained individuals. I explain how the study of olfactory learning has allowed understanding the discrimination and generalization ability of the honeybee olfactory system, its capacity to grant special properties to olfactory mixtures as well as to retain individual component information. Next, based on the impressive amount of anatomical and immunochemical studies of the bee brain, I detail our knowledge of olfactory pathways. I then show how functional recordings of odor-evoked activity in the brain allow following the transformation of the olfactory message from the periphery until higher-order central structures. Data from extra- and intracellular electrophysiological approaches as well as from the most recent optical imaging developments are described. Lastly, I discuss results addressing how odor representation changes as a result of experience. This impressive ensemble of behavioral, neuroanatomical, and neurophysiological data available in the bee make it an attractive model for future research aiming to understand olfactory perception and learning in an integrative fashion

A Flight Sensory-Motor to Olfactory Histamine Circuit Mediates Olfactory Processing of Ecologically and Behaviorally Natural Stimuli

Environmental pressures have conferred species specific behavioral and morphological traits to optimize reproductive success. To optimally interact with their environment, nervous systems have evolved motor-to-sensory circuits that mediate the processing of its own reafference. Moth flight behavioral patterns to odor sources are stereotyped, presumably to optimize the likelihood of interacting with the odor source. In the moth Manduca sexta wing beating causes oscillatory flow of air over the antenna; because of this, odorant-antennal interactions are oscillatory in nature. Electroantennogram recordings on antennae show that the biophysical properties of their spiking activity can effectively track odors presented at the wing beat frequency. Psychophysical experiments using Manduca show that when odors are pulsed, as opposed to presented as a continuous stream, detection and discrimination thresholds are lowered. In this study, we characterized histamine immunoreactivity in the thoracic ganglia and brain of Manduca. We generated antibodies for and characterized the distribution of the histamine B receptor, the first known antibody for this receptor protein. Our results show an elaborate pair of neurons projecting from the mesothoracic ganglion to the brain, including axon innervation of the antennal lobe and antennal mechanosensory and motor centers. Additionally, histamine B receptor labeling overlapped with a subset of GABAergic and peptidergic local interneurons. Next, we characterized the response properties of these cells within the context of fictive flight behavior and found a tonic increase in activity. Furthermore, disrupting this circuit, with surgical ablation and pharmacology, disrupts antennal lobe projection neurons from entraining to odors presented at a natural 20 Hz frequency, as well as behavioral measures of detection and discrimination thresholds. Finally, we characterized the relationship between motor patterns/behaviors, and circuit structure of this pair of histamine immunoreactive neurons. Specifically, presence of MDHn axon collaterals entering the antennal lobe is correlated with olfactory-guided target approach behaviors in crepuscular and nocturnal moths who require stereotyped zigzagging and wing beating behaviors for locating an olfactory target have axonal ramifications in the antennal lobe. This study is the first characterization of a motor to olfactory corollary discharge circuit in invertebrates and may represent the first characterization of a higher order corollary discharge circuit in an invertebrate model

Neural Circuit Dynamics and Ensemble Coding in the Locust and Fruit Fly Olfactory System

Raw sensory information is usually processed and reformatted by an organism’s brain to carry out tasks like identification, discrimination, tracking and storage. The work presented in this dissertation focuses on the processing strategies of neural circuits in the early olfactory system in two insects, the locust and the fruit fly. Projection neurons (PNs) in the antennal lobe (AL) respond to an odor presented to the locust’s antennae by firing in slow information-carrying temporal patterns, consistent across trials. Their downstream targets, the Kenyon cells (KCs) of the mushroom body (MB), receive input from large ensembles of transiently synchronous PNs at a time. The information arrives in slices of time corresponding to cycles of oscillatory activity originating in the AL. In the first part of the thesis, ensemble-level analysis techniques are used to understand how the AL-MB system deals with the problem of identifying odors across different concentrations. Individual PN odor responses can vary dramatically with concentration, but invariant patterns in PN ensemble responses are shown to allow odor identity to be extracted across a wide range of intensities by the KCs. Second, the sensitivity of the early olfactory system to stimulus history is examined. The PN ensemble and the KCs are found capable of tracking an odor in most conditions where it is pulsed or overlapping with another, but they occasionally fail (are masked) or reach intermediate states distinct from those seen for the odors presented alone or in a static mixture. The last part of the thesis focuses on the development of new recording techniques in the fruit fly, an organism with well-studied genetics and behavior. Genetically expressed fluorescent sensors of calcium offer the best available option to study ensemble activity in the fly. Here, simultaneous electrophysiology and two-photon imaging are used to estimate the correlation between G-CaMP, a popular genetically expressible calcium sensor, and electrical activity in PNs. The sensor is found to have poor temporal resolution and to miss significant spiking activity. More generally, this combination of electrophysiology and imaging enables explorations of functional connectivity and calibrated imaging of ensemble activity in the fruit fly.</p

Intrinsic and Extrinsic Neuronal Mechanisms in Temporal Coding: A Further Look at Neuronal Oscillations

Many studies in recent years have been devoted to the detection of fast oscillations in the Central Nervous System (CNS), interpreting them as synchronizing devices. We should, however, refrain from associating too closely the two concepts of synchronization and oscillation. Whereas synchronization is a relatively well-defined concept, by contrast oscillation of a population of neurones in the CNS looks loosely defined, in the sense that both its frequency sharpness and the duration of the oscillatory episodes vary widely from case to case. Also, the functions of oscillations in the brain are multiple and are not confined to synchronization. The paradigmatic instantiation of oscillation in physics is given by the harmonic oscillator, a device particularly suited to tell the time, as in clocks. We will thus examine first the case of oscillations or cycling discharges of neurones, which provide a clock or impose a “tempo” for various kinds of information processing. Neuronal oscillators are rarely just clocks clicking at a fixed frequency. Instead, their frequency is often adjustable and controllable, as in the example of the “chattering cells” discovered in the superficial layers of the visual cortex. Moreover, adjustable frequency oscillators are suitable for use in “phase locked loops” (PLL) networks, a device that can convert time coding to frequency coding; such PLL units have been found in the somatosensory cortex of guinea pigs. Finally, are oscillations stricto sensu necessary to induce synchronization in the discharges of downstream neurones? We know that this is not the case, at least not for local populations of neurones. As a contribution to this question, we propose that repeating patterns in neuronal discharges production may be looked at as one such alternative solution in relation to the processing of information. We review here the case of precisely repeating triplets, detected in the discharges of olfactory mitral cells of a freely breathing rat under odor stimulation

Short-term memory and olfactory signal processing

Modern neural recording methodologies, including multi-electrode and optical recordings, allow us to monitor the large population of neurons with high temporal resolution. Such recordings provide rich datasets that are expected to understand better how information about the external world is internally represented and how these representations are altered over time. Achieving this goal requires the development of novel pattern recognition methods and/or the application of existing statistical methods in novel ways to gain insights into basic neural computational principles. In this dissertation, I will take this data-driven approach to dissect the role of short-term memory in olfactory signal processing in two relatively simple models of the olfactory system: fruit fly (Drosophila melanogaster) and locust (Schistocerca americana). First, I will focus on understanding how odor representations within a single stimulus exposure are refined across different populations of neurons (faster dynamics; on the order seconds) in the early olfactory circuits. Using light-sheet imaging datasets from transgenic flies expressing calcium indicators in select populations of neurons, I will reveal how odor representations are decorrelated over time in different neural populations. Further, I will examine how this computation is altered by short-term memory in this neural circuitry. Next, I will examine how neural representations for odorants at an ensemble level are altered across different exposures (slower dynamics; on the order of tens of seconds to minutes). I will examine the role of this short-term adaptation in altering neural representations for odor identity and intensity. Lastly, I will present approaches to help achieve robustness against both extrinsic and intrinsic perturbations of odor-evoked neural responses. I will conclude with a Boolean neural network inspired by the insect olfactory system and compare its performance against other state-of-the-art methods on standard machine learning benchmark datasets. In sum, this work will provide deeper insights into how short-term plasticity alters sensory neural representations and their computational significance