Fly Optic Lobe Research Articles

The Understanding of ON-Edge Motion Detection Through the Simulation Based on the Connectome of Drosophila’s Optic Lobe

The optic lobe of the fly is one of the prominent model systems for the neural mechanism of the motion detection. How a fly who lives under various visual situations of the nature processes the information from at most a few thousands of ommatidia in their neural circuit for the detection of moving objects is not exactly clear though many computational models of the fly optic lobe as a moving objects detector were suggested. Here we attempted to elucidate the mechanisms of ON-edge motion detection by a simulation approach based on the TEM connectome of Drosophila. Our simulation model of the optic lobe with the NEURON simulator that covers the full scale of ommatidia, reproduced the characteristics of the receptor neurons, lamina monopolar neurons, and T4 cells in the lobula. The contribution of each neuron can be estimated by changing synaptic connection strengths in the simulation and measuring the response to the motion stimulus. Those show the paradelle pathway provide motion detection in the fly optic lobe has more robustness and is more sophisticated than a simple combination of HR and BL systems.

Vision: Space and colour meet in the fly optic lobes

Colour vision involves colour-opponent cells, which are excited and inhibited by different wavelengths. Synaptic interconnections between Drosophila Dm8 cells are required for forming spatio-chromatic receptive fields with a center and surround of opposing polarity which can invert, depending on the stimulus.

"Neur"al brain wave: Coordinating epithelial-to-neural stem cell transition in the fly optic lobe.

In the Drosophila larval optic lobe, the generation of neural stem cells involves an epithelial-to-mesenchymal-like transition of a continuous stripe of cells that sweeps across the neuroepithelium, but the dynamics at cell and tissue level were unknown until now. In this issue, Shard et al. (2020. J. Cell Biol.https://doi.org/10.1083/jcb.202005035) identify that Neuralized controls a partial epithelial-to-mesenchymal transition through regulation of the apical Crumbs complex and through the coordination of cell behaviors such as apical constriction and cell alignment.

Persistent Firing and Adaptation in Optic-Flow-Sensitive Descending Neurons

A general principle of sensory systems is that they adapt to prolonged stimulation by reducing their response over time. Indeed, in many visual systems, including higher-order motion sensitive neurons in the fly optic lobes and the mammalian visual cortex, a reduction in neural activity following prolonged stimulation occurs. In contrast to this phenomenon, the response of the motor system controlling flight maneuvers persists following the offset of visual motion. It has been suggested that this gap is caused by a lingering calcium signal in the output synapses of fly optic lobe neurons. However, whether this directly affects the responses of the post-synaptic descending neurons, leading to the observed behavioral output, is not known. We use extracellular electrophysiology to record from optic-flow-sensitive descending neurons in response to prolonged wide-field stimulation. We find that, as opposed to most sensory and visual neurons, and in particular to the motion vision sensitive neurons in the brains of both flies and mammals, the descending neurons show little adaption during stimulus motion. In addition, we find that the optic-flow-sensitive descending neurons display persistent firing, or an after-effect, following the cessation of visual stimulation, consistent with the lingering calcium signal hypothesis. However, if the difference in after-effect is compensated for, subsequent presentation of stimuli in a test-adapt-test paradigm reveals adaptation to visual motion. Our results thus show a combination of adaptation and persistent firing in the neurons that project to the thoracic ganglia and thereby control behavioral output.

Visual motion sensitivity in descending neurons in the hoverfly

Many animals use motion vision information to control dynamic behaviors. For example, flying insects must decide whether to pursue a prey or not, to avoid a predator, to maintain their current flight trajectory, or to land. The neural mechanisms underlying the computation of visual motion have been particularly well investigated in the fly optic lobes. However, the descending neurons, which connect the optic lobes with the motor command centers of the ventral nerve cord, remain less studied. To address this deficiency, we describe motion vision sensitive descending neurons in the hoverfly Eristalis tenax. We describe how the neurons can be identified based on their receptive field properties, and how they respond to moving targets, looming stimuli and to widefield optic flow. We discuss their similarities with previously published visual neurons, in the optic lobes and ventral nerve cord, and suggest that they can be classified as target-selective, looming sensitive and optic flow sensitive, based on these similarities. Our results highlight the importance of using several visual stimuli as the neurons can rarely be identified based on only one response characteristic. In addition, they provide an understanding of the neurophysiology of visual neurons that are likely to affect behavior.

Persistent Firing and Adaptation in Optic Flow Sensitive Descending Neurons

A general principle of sensory systems is that they adapt to prolonged stimulation by reducing their response over time. Indeed, many visual systems, including higher-order motion sensitive neurons in the fly optic lobes and the mammalian visual cortex, a reduction in neural activity following prolonged stimulation occurs. In contrast to this phenomenon, the response of the motor system controlling flight maneuvers persists following the offset of visual motion. It has been suggested that this gap is caused by a lingering calcium signal in the output synapses of optic lobe neurons. However, whether this directly affects the responses of the post-synaptic descending neurons, leading to the observed behavioral output, is not known. We use extracellular electrophysiology to record from optic flow sensitive descending neurons in response to prolonged wide-field stimulation. We find that, as opposed to most sensory and visual neurons, and in particular to the motion vision sensitive neurons in the brains of flies and mammals, the descending neurons show little adaption during stimulus motion. In addition, we find that the optic flow sensitive descending neurons display persistent firing, or an after-effect, following the cessation of visual stimulation, consistent with the lingering calcium signal hypothesis. However, if the difference in after-effect is compensated for, subsequent presentation of stimuli in a test-adapt-test paradigm reveals adaptation to visual motion. Our results thus show an interesting combination of adaptation and persistent firing, in the neurons that project to the thoracic ganglia, and thereby control behavioral output.

Modality-Specific Circuits for Skylight Orientation in the Fly Visual System.

In the fly optic lobe, ∼800 highly stereotypical columnar microcircuits are arranged retinotopically to process visual information. Differences in cellular composition and synaptic connectivity within functionally specialized columns remain largely unknown. Here, we describe the cellular and synaptic architecture in medulla columns located downstream of photoreceptors in the dorsal rim area (DRA), where linearly polarized skylight is detected for guiding orientation responses. We show that only in DRA medulla columns both R7 and R8 photoreceptors target to the bona fide R7 target layerwhere they form connections with previously uncharacterized, modality-specific Dm neurons: two morphologically distinct DRA-specific cell types (termed Dm-DRA1 and Dm-DRA2) stratify in separate sublayers and exclusively contact polarization-sensitive DRA inputs, while avoiding overlaps with color-sensitive Dm8 cells. Using the activity-dependent GRASP and trans-Tango techniques, we confirm thatDRA R7 cells are synaptically connected to Dm-DRA1, whereas DRA R8 form synapses with Dm-DRA2. Finally, using live imaging of ingrowing pupal photoreceptor axons, we show that DRA R7 and R8 termini reach layer M6 sequentially, thus separating the establishment of different synaptic connectivity in time. We propose that a duplication of R7→Dm circuitry in DRA ommatidia serves as an ideal adaptation for detecting linearly polarized skylight using orthogonal e-vector analyzers.

Full reconstruction of large lobula plate tangential cells in Drosophila from a 3D EM dataset.

With the advent of neurogenetic methods, the neural basis of behavior is presently being analyzed in more and more detail. This is particularly true for visually driven behavior of Drosophila melanogaster where cell-specific driver lines exist that, depending on the combination with appropriate effector genes, allow for targeted recording, silencing and optogenetic stimulation of individual cell-types. Together with detailed connectomic data of large parts of the fly optic lobe, this has recently led to much progress in our understanding of the neural circuits underlying local motion detection. However, how such local information is combined by optic flow sensitive large-field neurons is still incompletely understood. Here, we aim to fill this gap by a dense reconstruction of lobula plate tangential cells of the fly lobula plate. These neurons collect input from many hundreds of local motion-sensing T4/T5 neurons and connect them to descending neurons or central brain areas. We confirm all basic features of HS and VS cells as published previously from light microscopy. In addition, we identified the dorsal and the ventral centrifugal horizontal, dCH and vCH cell, as well as three VSlike cells, including their distinct dendritic and axonal projection area.

A common directional tuning mechanism of Drosophila motion-sensing neurons in the ON and in the OFF pathway.

In the fruit fly optic lobe, T4 and T5 cells represent the first direction-selective neurons, with T4 cells responding selectively to moving brightness increments (ON) and T5 cells to brightness decrements (OFF). Both T4 and T5 cells comprise four subtypes with directional tuning to one of the four cardinal directions. We had previously found that upward-sensitive T4 cells implement both preferred direction enhancement and null direction suppression (Haag et al., 2016). Here, we asked whether this mechanism generalizes to OFF-selective T5 cells and to all four subtypes of both cell classes. We found that all four subtypes of both T4 and T5 cells implement both mechanisms, that is preferred direction enhancement and null direction inhibition, on opposing sides of their receptive fields. This gives rise to the high degree of direction selectivity observed in both T4 and T5 cells within each subpopulation.

Visual Circuits for Direction Selectivity.

Images projected onto the retina of an animal eye are rarely still. Instead, they usually contain motion signals originating either from moving objects or from retinal slip caused by self-motion. Accordingly, motion signals tell the animal in which direction a predator, prey, or the animal itself is moving. At the neural level, visual motion detection has been proposed to extract directional information by a delay-and-compare mechanism, representing a classic example of neural computation. Neurons responding selectively to motion in one but not in the other direction have been identified in many systems, most prominently in the mammalian retina and the fly optic lobe. Technological advances have now allowed researchers to characterize these neurons' upstream circuits in exquisite detail. Focusing on these upstream circuits, we review and compare recent progress in understanding the mechanisms that generate direction selectivity in the early visual system of mammals and flies.

Analyzing Dendritic Morphology in Columns and Layers.

In many regions of the central nervous systems, such as the fly optic lobes and the vertebrate cortex, synaptic circuits are organized in layers and columns to facilitate brain wiring during development and information processing in developed animals. Postsynaptic neurons elaborate dendrites in type-specific patterns in specific layers to synapse with appropriate presynaptic terminals. The fly medulla neuropil is composed of 10 layers and about 750 columns; each column is innervated by dendrites of over 38 types of medulla neurons, which match with the axonal terminals of some 7 types of afferents in a type-specific fashion. This report details the procedures to image and analyze dendrites of medulla neurons. The workflow includes three sections: (i) the dual-view imaging section combines two confocal image stacks collected at orthogonal orientations into a high-resolution 3D image of dendrites; (ii) the dendrite tracing and registration section traces dendritic arbors in 3D and registers dendritic traces to the reference column array; (iii) the dendritic analysis section analyzes dendritic patterns with respect to columns and layers, including layer-specific termination and planar projection direction of dendritic arbors, and derives estimates of dendritic branching and termination frequencies. The protocols utilize custom plugins built on the open-source MIPAV (Medical Imaging Processing, Analysis, and Visualization) platform and custom toolboxes in the matrix laboratory language. Together, these protocols provide a complete workflow to analyze the dendritic routing of Drosophila medulla neurons in layers and columns, to identify cell types, and to determine defects in mutants.

Dscam2 affects visual perception in Drosophila melanogaster.

Dscam2, a cell surface protein that mediates cellular repulsion, plays a crucial role in the development of the Drosophila melanogaster visual system. Dscam2 generates boundaries between neighboring modules in the fly optic lobe; in Dscam2 mutants this visual system modularity is compromised. Although developmental wiring defects have been well described in the Dscam2 mutant, behavioral consequences have not been investigated. To address this, we examined the visual behavior of Dscam2 mutant flies. Using a phototaxis assay, we ascertained that these flies are not blind, but have a reduced phototaxic response. Through population-based and single fly optomotor assays, we found that Dscam2 mutant flies can track motion but that their response is opposite to control flies under defined experimental conditions. In a fixation paradigm, which allows tethered flies to control the angular position of a visual stimulus, mutant flies' responses were diametrically opposed to those seen in control flies. These data suggest that modest changes in the modularity of the fly visual system in the Dscam2 mutant can dramatically change the perception of specific visual cues and modify behavior.

Neurons forming optic glomeruli compute figure-ground discriminations in Drosophila.

Many animals rely on visual figure-ground discrimination to aid in navigation, and to draw attention to salient features like conspecifics or predators. Even figures that are similar in pattern and luminance to the visual surroundings can be distinguished by the optical disparity generated by their relative motion against the ground, and yet the neural mechanisms underlying these visual discriminations are not well understood. We show in flies that a diverse array of figure-ground stimuli containing a motion-defined edge elicit statistically similar behavioral responses to one another, and statistically distinct behavioral responses from ground motion alone. From studies in larger flies and other insect species, we hypothesized that the circuitry of the lobula--one of the four, primary neuropiles of the fly optic lobe--performs this visual discrimination. Using calcium imaging of input dendrites, we then show that information encoded in cells projecting from the lobula to discrete optic glomeruli in the central brain group these sets of figure-ground stimuli in a homologous manner to the behavior; "figure-like" stimuli are coded similar to one another and "ground-like" stimuli are encoded differently. One cell class responds to the leading edge of a figure and is suppressed by ground motion. Two other classes cluster any figure-like stimuli, including a figure moving opposite the ground, distinctly from ground alone. This evidence demonstrates that lobula outputs provide a diverse basis set encoding visual features necessary for figure detection.

Neurogenesis and neuronal circuit formation in the Drosophila visual center.

The Drosophila optic lobe is composed of a wide variety of neurons that form laminar structures and columnar units. The fly optic lobe shares structural features with the mammalian brain, and fly genetics allow precise genetic manipulations. Thus, the Drosophila visual center is an excellent model for studying the mechanisms underlying the establishment of a functional neuronal circuit during brain development. However, little is understood about the developmental mechanisms that produce neuronal diversity and establish neuronal circuits in the medulla, the largest component of the optic lobe. Our recent research revealed key features of medulla development, such as birth-order-dependent specification of neuronal types and the subdivision of the medulla primordium into concentric zones, which is characterized by the expression of four transcription factors. Here, we review recent investigations into the development of the medulla and discuss the mechanisms that establish functional neuronal circuits.

A directional tuning map of Drosophila elementary motion detectors

The extraction of directional motion information from changing retinal images is one of the earliest and most important processing steps in any visual system. In the fly optic lobe, two parallel processing streams have been anatomically described, leading from two first-order interneurons, L1 and L2, via T4 and T5 cells onto large, wide-field motion-sensitive interneurons of the lobula plate. Therefore, T4 and T5 cells are thought to have a pivotal role in motion processing; however, owing to their small size, it is difficult to obtain electrical recordings of T4 and T5 cells, leaving their visual response properties largely unknown. We circumvent this problem by means of optical recording from these cells in Drosophila, using the genetically encoded calcium indicator GCaMP5 (ref. 2). Here we find that specific subpopulations of T4 and T5 cells are directionally tuned to one of the four cardinal directions; that is, front-to-back, back-to-front, upwards and downwards. Depending on their preferred direction, T4 and T5 cells terminate in specific sublayers of the lobula plate. T4 and T5 functionally segregate with respect to contrast polarity: whereas T4 cells selectively respond to moving brightness increments (ON edges), T5 cells only respond to moving brightness decrements (OFF edges). When the output from T4 or T5 cells is blocked, the responses of postsynaptic lobula plate neurons to moving ON (T4 block) or OFF edges (T5 block) are selectively compromised. The same effects are seen in turning responses of tethered walking flies. Thus, starting with L1 and L2, the visual input is split into separate ON and OFF pathways, and motion along all four cardinal directions is computed separately within each pathway. The output of these eight different motion detectors is then sorted such that ON (T4) and OFF (T5) motion detectors with the same directional tuning converge in the same layer of the lobula plate, jointly providing the input to downstream circuits and motion-driven behaviours.

Activity and coexpression of Drosophila black with ebony in fly optic lobes reveals putative cooperative tasks in vision that evade electroretinographic detection

AbstractDrosophila mutants black and ebony show pigmentation defects in the adult cuticle, which disclose their cooperative activity in β‐alanyl‐dopamine formation. In visual signal transduction, Ebony conjugates β‐alanine to histamine, forming β‐alanyl‐histamine or carcinine. Mutation of ebony disrupts signal transduction and reveals an electroretinogram (ERG) phenotype. In contrast to the corresponding cuticle phenotype of black and ebony, there is no ERG phenotype observed when black expression is disrupted. This discrepancy calls into question the longstanding assumption of Black and Ebony interaction. The purpose of this study was to investigate the role of Black and Ebony in fly optic lobes. We excluded a presynaptic histamine uptake pathway and confirmed histamine recycling via carcinine formation in glia. β‐Alanine supply for this pathway is independent of enzymatic synthesis by Black and β‐alanine synthase Pyd3. Two versions of Black are expressed in vivo. Black is a specific aspartate decarboxylase with no activity on glutamate. RNA in situ hybridization and anti‐Black antisera localized Black expression in the head. Immunolabeling revealed expression in lamina glia, in large medulla glia, in glia of the ocellar ganglion, and in astrocyte‐like glia below the ocellar ganglion. In these glia types, Black expression is strictly accompanied by Ebony expression. Activity, localization, and strict coexpression with Ebony strongly indicate a specific mode of functional interaction that, however, evades ERG detection. © 2013 Wiley Periodicals, Inc.

Activity and coexpression of Drosophila black with ebony in fly optic lobes reveals putative cooperative tasks in vision that evade electroretinographic detection

Drosophila mutants black and ebony show pigmentation defects in the adult cuticle, which disclose their cooperative activity in β-alanyl-dopamine formation. In visual signal transduction, Ebony conjugates β-alanine to histamine, forming β-alanyl-histamine or carcinine. Mutation of ebony disrupts signal transduction and reveals an electroretinogram (ERG) phenotype. In contrast to the corresponding cuticle phenotype of black and ebony, there is no ERG phenotype observed when black expression is disrupted. This discrepancy calls into question the longstanding assumption of Black and Ebony interaction. The purpose of this study was to investigate the role of Black and Ebony in fly optic lobes. We excluded a presynaptic histamine uptake pathway and confirmed histamine recycling via carcinine formation in glia. β-Alanine supply for this pathway is independent of enzymatic synthesis by Black and β-alanine synthase Pyd3. Two versions of Black are expressed in vivo. Black is a specific aspartate decarboxylase with no activity on glutamate. RNA in situ hybridization and anti-Black antisera localized Black expression in the head. Immunolabeling revealed expression in lamina glia, in large medulla glia, in glia of the ocellar ganglion, and in astrocyte-like glia below the ocellar ganglion. In these glia types, Black expression is strictly accompanied by Ebony expression. Activity, localization, and strict coexpression with Ebony strongly indicate a specific mode of functional interaction that, however, evades ERG detection.

Optogenetic Stimulation of Escape Behavior in <em>Drosophila melanogaster</em>

A growing number of genetically encoded tools are becoming available that allow non-invasive manipulation of the neural activity of specific neurons in Drosophila melanogaster. Chief among these are optogenetic tools, which enable the activation or silencing of specific neurons in the intact and freely moving animal using bright light. Channelrhodopsin (ChR2) is a light-activated cation channel that, when activated by blue light, causes depolarization of neurons that express it. ChR2 has been effective for identifying neurons critical for specific behaviors, such as CO(2) avoidance, proboscis extension and giant-fiber mediated startle response. However, as the intense light sources used to stimulate ChR2 also stimulate photoreceptors, these optogenetic techniques have not previously been used in the visual system. Here, we combine an optogenetic approach with a mutation that impairs phototransduction to demonstrate that activation of a cluster of loom-sensitive neurons in the fly's optic lobe, Foma-1 neurons, can drive an escape behavior used to avoid collision. We used a null allele of a critical component of the phototransduction cascade, phospholipase C-β, encoded by the norpA gene, to render the flies blind and also use the Gal4-UAS transcriptional activator system to drive expression of ChR2 in the Foma-1 neurons. Individual flies are placed on a small platform surrounded by blue LEDs. When the LEDs are illuminated, the flies quickly take-off into flight, in a manner similar to visually driven loom-escape behavior. We believe that this technique can be easily adapted to examine other behaviors in freely moving flies.

Optogenetic Stimulation of Escape Behavior in <em>Drosophila melanogaster</em>

A growing number of genetically encoded tools are becoming available that allow non-invasive manipulation of the neural activity of specific neurons in Drosophila melanogaster1. Chief among these are optogenetic tools, which enable the activation or silencing of specific neurons in the intact and freely moving animal using bright light. Channelrhodopsin (ChR2) is a light-activated cation channel that, when activated by blue light, causes depolarization of neurons that express it. ChR2 has been effective for identifying neurons critical for specific behaviors, such as CO2 avoidance, proboscis extension and giant-fiber mediated startle response2-4. However, as the intense light sources used to stimulate ChR2 also stimulate photoreceptors, these optogenetic techniques have not previously been used in the visual system. Here, we combine an optogenetic approach with a mutation that impairs phototransduction to demonstrate that activation of a cluster of loom-sensitive neurons in the fly's optic lobe, Foma-1 neurons, can drive an escape behavior used to avoid collision. We used a null allele of a critical component of the phototransduction cascade, phospholipase C-β, encoded by the norpA gene, to render the flies blind and also use the Gal4-UAS transcriptional activator system to drive expression of ChR2 in the Foma-1 neurons. Individual flies are placed on a small platform surrounded by blue LEDs. When the LEDs are illuminated, the flies quickly take-off into flight, in a manner similar to visually driven loom-escape behavior. We believe that this technique can be easily adapted to examine other behaviors in freely moving flies.

Two-photon calcium imaging from head-fixed Drosophila during optomotor walking behavior.

Drosophila melanogster is a model organism rich in genetic tools to manipulate and identify neural circuits involved in specific behaviors. Here we present a novel technique for two-photon calcium imaging in the central brain of head-fixed Drosophila walking on an air-supported ball. The ball’s motion is tracked at high resolution and can be treated as a proxy for the fly’s own movements. We used the genetically encoded calcium sensor, GCaMP3.0, to record from important elements of the motion-processing pathway, the horizontal-system (HS) lobula plate tangential cells (LPTCs) in the fly optic lobe. We presented motion stimuli to the tethered fly and found that calcium transients in HS-neurons correlated with robust optomotor behavior during walking. Our technique allows an entirely new set of questions to be addressed by monitoring behavior and physiology in identified neurons in a powerful genetic model organism with an extensive repertoire of walking behaviors.