A fly inspired solution to looming detection for collision avoidance.

Dodging rapidly approaching objects is a fundamental skill for both animals and intelligent robots. Flies are adept at high-speed collision avoidance. However, it remains unclear whether the fly algorithm can be extracted and is applicable to real-time machine vision. In this study, we developed a computational model inspired by the looming detection circuit recently identified in Drosophila. Our results suggest that in the face of considerably noisy local motion signals, the key for the fly circuit to achieve accurate detection is attributed to two computation strategies: population encoding and nonlinear integration. The model is further shown to be an effective algorithm for collision avoidance by virtual robot tests. The algorithm is characterized by practical flexibility, whose looming detection parameters can be modulated depending on factors such as the body size of the robots. The model sheds light on the potential of the concise fly algorithm in real-time applications.

Bioinspired figure-ground discrimination via visual motion smoothing.

Flies detect and track moving targets among visual clutter, and this process mainly relies on visual motion. Visual motion is analyzed or computed with the pathway from the retina to T4/T5 cells. The computation of local directional motion was formulated as an elementary movement detector (EMD) model more than half a century ago. Solving target detection or figure-ground discrimination problems can be equivalent to extracting boundaries between a target and the background based on the motion discontinuities in the output of a retinotopic array of EMDs. Individual EMDs cannot measure true velocities, however, due to their sensitivity to pattern properties such as luminance contrast and spatial frequency content. It remains unclear how local directional motion signals are further integrated to enable figure-ground discrimination. Here, we present a computational model inspired by fly motion vision. Simulations suggest that the heavily fluctuating output of an EMD array is naturally surmounted by a lobula network, which is hypothesized to be downstream of the local motion detectors and have parallel pathways with distinct directional selectivity. The lobula network carries out a spatiotemporal smoothing operation for visual motion, especially across time, enabling the segmentation of moving figures from the background. The model qualitatively reproduces experimental observations in the visually evoked response characteristics of one type of lobula columnar (LC) cell. The model is further shown to be robust to natural scene variability. Our results suggest that the lobula is involved in local motion-based target detection.

Classifying Drosophila olfactory projection neuron boutons by quantitative analysis of electron microscopic reconstruction.

In Drosophila melanogaster, olfactory projection neurons (PNs) convey odor information from the antenna lobe to higher brain regions. Recent transcriptomic studies reveal a large diversity of transcription factors, cell-surface molecules, neurotransmitter-coding, and neuropeptide-coding genes in PNs; however, their structural diversity remains unknown. Herein, we achieved a volumetric reconstruction of 89 PN boutons under Focused Ion Beam Scanning Electron Microscopy (FIB-SEM) and quantitatively analyzed the internal presynaptic active zones (PAZs) and dense-core vesicles (DCVs). The ultrastructure-based cluster analysis reveals three morphological distinct bouton subtypes: complex boutons, unilobed boutons, and simple boutons. The complex boutons contain the most PAZs and DCVs, which suggests that they are of the highest capability of releasing neurotransmitters and neuromodulators. By labeling a subset of boutons under FIB-SEM, we found that DCVs are preferentially distributed in certain GH146-positive subtypes. Our study demonstrates that PN boutons display distinct morphology, which may determine their capacity of releasing neurotransmitters and neuromodulators.

A neural algorithm for Drosophila linear and nonlinear decision-making.

It has been evidenced that vision-based decision-making in Drosophila consists of both simple perceptual (linear) decision and value-based (non-linear) decision. This paper proposes a general computational spiking neural network (SNN) model to explore how different brain areas are connected contributing to Drosophila linear and nonlinear decision-making behavior. First, our SNN model could successfully describe all the experimental findings in fly visual reinforcement learning and action selection among multiple conflicting choices as well. Second, our computational modeling shows that dopaminergic neuron-GABAergic neuron-mushroom body (DA-GABA-MB) works in a recurrent loop providing a key circuit for gain and gating mechanism of nonlinear decision making. Compared with existing models, our model shows more biologically plausible on the network design and working mechanism, and could amplify the small differences between two conflicting cues more clearly. Finally, based on the proposed model, the UAV could quickly learn to make clear-cut decisions among multiple visual choices and flexible reversal learning resembling to real fly. Compared with linear and uniform decision-making methods, the DA-GABA-MB mechanism helps UAV complete the decision-making task with fewer steps.

The receptor channel formed by ppk25, ppk29 and ppk23 can sense the Drosophila female pheromone 7,11-heptacosadiene.

In Drosophila, pheromones play a crucial role in regulating courtship behaviors. In males, female aphrodisiac pheromones promote male-female courtship, and male antiaphrodisiac pheromones inhibit male-male courtship. Previous studies have reported that receptor proteins belonging to the pickpocket (ppk) family, ionotropic receptor family and gustatory receptor family are required for pheromone detection and normal courtship. However, none of them has been shown to be sufficient for sensing pheromones after ectopic expression in originally unresponsive cells. "M" cells are activated by male antiaphrodisiac pheromones but not female aphrodisiac pheromones, and the activated cells inhibit male-male courtship. In our study, male flies with ectopic expression of ppk25, ppk29 and ppk23 in "M" cells showed decreased male-female courtship. Using an in vivo calcium imaging approach, we found that the "M" cells expressing these three ppks were significantly activated by the female aphrodisiac pheromone 7,11-heptacosadiene (7,11-HD). Our results indicate that a sodium channel consisting, at minimum, of ppk25, ppk29 and ppk23, can sense 7,11-HD, most likely as a receptor. Our findings may help us gain insights into the molecular mechanisms of pheromonal functions.

Suppression of GABAergic neurons through D2-like receptor secures efficient conditioning in Drosophila aversive olfactory learning.

The GABAergic system serves as a vital negative modulator in cognitive functions, such as learning and memory, while the mechanisms governing this inhibitory system remain to be elucidated. In Drosophila, the GABAergic anterior paired lateral (APL) neurons mediate a negative feedback essential for odor discrimination; however, their activity is suppressed by learning via unknown mechanisms. In aversive olfactory learning, a group of dopaminergic (DA) neurons is activated on electric shock (ES) and modulates the Kenyon cells (KCs) in the mushroom body, the center of olfactory learning. Here we find that the same group of DA neurons also form functional synaptic connections with the APL neurons, thereby emitting a suppressive signal to the latter through Drosophila dopamine 2-like receptor (DD2R). Knockdown of either DD2R or its downstream molecules in the APL neurons results in impaired olfactory learning at the behavioral level. Results obtained from in vivo functional imaging experiments indicate that this DD2R-dependent DA-to-APL suppression occurs during odor-ES conditioning and discharges the GABAergic inhibition on the KCs specific to the conditioned odor. Moreover, the decrease in odor response of the APL neurons persists to the postconditioning phase, and this change is also absent in DD2R knockdown flies. Taken together, our findings show that DA-to-GABA suppression is essential for restraining the GABAergic inhibition during conditioning, as well as for inducing synaptic modification in this learning circuit. Such circuit mechanisms may play conserved roles in associative learning across species.

Functional organization of intrinsic and feedback presynaptic inputs in the primary visual cortex.

In the primary visual cortex (V1) of many mammalian species, neurons are spatially organized according to their preferred orientation into a highly ordered map. However, whether and how the various presynaptic inputs to V1 neurons are organized relative to the neuronal orientation map remain unclear. To address this issue, we constructed genetically encoded calcium indicators targeting axon boutons in two colors and used them to map the organization of axon boutons of V1 intrinsic and V2-V1 feedback projections in tree shrews. Both connections are spatially organized into maps according to the preferred orientations of axon boutons. Dual-color calcium imaging showed that V1 intrinsic inputs are precisely aligned to the orientation map of V1 cell bodies, while the V2-V1 feedback projections are aligned to the V1 map with less accuracy. Nonselective integration of intrinsic presynaptic inputs around the dendritic tree is sufficient to reproduce cell body orientation preference. These results indicate that a precisely aligned map of intrinsic inputs could reinforce the neuronal map in V1, a principle that may be prevalent for brain areas with function maps.

Generation of low-gamma oscillations in a GABAergic network model of the striatum.

Striatal oscillations in the low-gamma frequency range have been consistently recorded in a number of experimental studies. However, whether these rhythms are locally generated in the striatum circuit, which is mainly composed of GABAergic neurons, remains an open question. GABAergic medium spiny projection neurons represent the great majority of striatal neurons, but they fire at very low rates. GABAergic fast-spiking interneurons typically show firing rates that are approximately 10 times higher than those of principal neurons, but they are a very small minority of the total neuronal population. In this study, based on physiological constraints we developed a computational network model of these neurons and dissected the oscillations. Simulations showed that the population of medium spiny projection neurons, and not the GABAergic fast-spiking interneurons, determines the frequency range of the oscillations. D2-type dopamine receptor-expressing neurons dominate the generation of low-gamma rhythms. Feedforward inputs from GABAergic fast-spiking interneurons promote the oscillations by strengthening the inhibitory interactions between medium spiny projection neurons. The promotion effect is independent of the degree of synchronization in the fast-spiking interneuron population but affected by the strength of their feedforward inputs to medium spiny projection neurons. Our results provide a theoretical explanation for how firing properties and connections of the three types of GABAergic neurons, which are susceptible to on-going behaviors, experience, and dopamine disruptions, sculpt striatal oscillations.

Locomotor Assay in Drosophila melanogaster.

This protocol describes a simple locomotor assay in Drosophila melanogaster. In brief, the locomotor of each single fly in the culture dish is recorded by a web camera. The moving time, walking length, speed and the locomotor trails of the single fly could be quantitatively analyzed.

Nicotine-induced acute hyperactivity is mediated by dopaminergic system in a sexually dimorphic manner.

Short-term exposure to nicotine induces positive effects in mice, monkeys and humans, including mild euphoria, hyperactivity, and enhanced cognition. However, the underlying neural basis and molecular mechanisms for these effects remain poorly understood. Here, using a video recording system, we find that acute nicotine administration induces locomotor hyperactivity in Drosophila, similar to observations made in higher model organisms. Suppressing dopaminergic neurons or down-regulating dopamine 1-like receptor (DopR) abolishes this acute nicotine response, but surprisingly, does so only in male flies. Using a GFP reconstitution across synaptic partners (GRASP) approach, we show that dopaminergic neurons possess potential synaptic connections with acetylcholinergic neurons in wide regions of the brain. Furthermore, dopaminergic neurons are widely activated upon nicotine perfusion in both sexes, while the response curve differs significantly between the sexes. Moreover, knockdown of the ß1 nicotine acetylcholine receptor (nAChR) in dopaminergic neurons abolishes the acute nicotine response only in male flies, while panneural knock-down occurs in both sexes. Taken together, our results reveal that in fruit flies, dopaminergic neurons mediate nicotine-induced acute locomotor hyperactivity in a sexually dimorphic manner, and Drosophila ß1 nAChR subunit plays a crucial role in this nicotine response. These findings provide important insights into the molecular and neural basis of acute nicotine effects, and the underlying mechanisms may play conserved roles across species.

Gap junction networks in mushroom bodies participate in visual learning and memory in Drosophila.

Gap junctions are widely distributed in the brains across species and play essential roles in neural information processing. However, the role of gap junctions in insect cognition remains poorly understood. Using a flight simulator paradigm and genetic tools, we found that gap junctions are present in Drosophila Kenyon cells (KCs), the major neurons of the mushroom bodies (MBs), and showed that they play an important role in visual learning and memory. Using a dye coupling approach, we determined the distribution of gap junctions in KCs. Furthermore, we identified a single pair of MB output neurons (MBONs) that possess a gap junction connection to KCs, and provide strong evidence that this connection is also required for visual learning and memory. Together, our results reveal gap junction networks in KCs and the KC-MBON circuit, and bring new insight into the synaptic network underlying fly's visual learning and memory.

CRASP: CFP reconstitution across synaptic partners.

Mapping the pattern of connectivity between neurons is widely regarded to be critical for understanding the nervous system. GRASP (GFP reconstitution across synaptic partners) has been used as a promising method for mapping neuronal connectivity, but is currently available in the green color only, limiting its potential applications. Here we demonstrate CRASP (CFP reconstitution across synaptic partners), a cyan-colored version of GRASP. We validated the system in HEK 293T cells, and generated transgenic Drosophila lines to show that the system could reliably detect neuronal contacts in the brain. Furthermore, we showed that the CRASP signal could be selectively amplified using standard immunohistochemistry methods. The CRASP system adds to the toolkit available to researchers for mapping neuronal connectivity, and substantially expands the potential application of GRASP-like strategies.

A Novel Method for Tracking Individuals of Fruit Fly Swarms Flying in a Laboratory Flight Arena.

The growing interest in studying social behaviours of swarming fruit flies, Drosophila melanogaster, has heightened the need for developing tools that provide quantitative motion data. To achieve such a goal, multi-camera three-dimensional tracking technology is the key experimental gateway. We have developed a novel tracking system for tracking hundreds of fruit flies flying in a confined cubic flight arena. In addition to the proposed tracking algorithm, this work offers additional contributions in three aspects: body detection, orientation estimation, and data validation. To demonstrate the opportunities that the proposed system offers for generating high-throughput quantitative motion data, we conducted experiments on five experimental configurations. We also performed quantitative analysis on the kinematics and the spatial structure and the motion patterns of fruit fly swarms. We found that there exists an asymptotic distance between fruit flies in swarms as the population density increases. Further, we discovered the evidence for repulsive response when the distance between fruit flies approached the asymptotic distance. Overall, the proposed tracking system presents a powerful method for studying flight behaviours of fruit flies in a three-dimensional environment.

Aging accelerates memory extinction and impairs memory restoration in Drosophila.

Age-related memory impairment (AMI) is a phenomenon observed from invertebrates to human. Memory extinction is proposed to be an active inhibitory modification of memory, however, whether extinction is affected in aging animals remains to be elucidated. Employing a modified paradigm for studying memory extinction in fruit flies, we found that only the stable, but not the labile memory component was suppressed by extinction, thus effectively resulting in higher memory loss in aging flies. Strikingly, young flies were able to fully restore the stable memory component 3 h post extinction, while aging flies failed to do so. In conclusion, our findings reveal that both accelerated extinction and impaired restoration contribute to memory impairment in aging animals.

Astrocyte-like glial cells physiologically regulate olfactory processing through the modification of ORN-PN synaptic strength in Drosophila.

Astrocyte-like glial cells are abundant in the central nervous system of adult Drosophila and exhibit morphology similar to astrocytes of mammals. Previous evidence has shown that astrocyte-like glial cells are strongly associated with synapses in the antennal lobe (AL), the first relay of the olfactory system, where olfactory receptor neurons (ORNs) transmit information into projection neurons (PNs). However, the function of astrocyte-like glia in the AL remains obscure. In this study, using in vivo calcium imaging, we found that astrocyte-like glial cells exhibited spontaneous microdomain calcium elevations. Using simultaneous manipulation of glial activity and monitoring of neuronal function, we found that the astrocyte-like glial activation, but not ensheathing glial activation, could inhibit odor-evoked responses of PNs. Ensheathing glial cells are another subtype of glia, and are of functional importance in the AL. Electrophysiological experiments indicated that astrocyte-like glial activation decreased the amplitude and slope of excitatory postsynaptic potentials evoked through electrical stimulation of the antennal nerve. These results suggest that astrocyte-like glial cells may regulate olfactory processing through negative regulation of ORN-PN synaptic strength. Beyond the antennal lobe we observed astrocyte-like glial spontaneous calcium activities in the ventromedial protocerebrum, indicating that astrocyte-like glial spontaneous calcium elevations might be general in the adult fly brain. Overall, our study demonstrates a new function for astrocyte-like glial cells in the physiological modulation of olfactory information transmission, possibly through regulating ORN-PN synapse strength.

A subset of cholinergic mushroom body neurons requires Go signaling to regulate sleep in Drosophila.

STUDY OBJECTIVES: Identifying the neurochemistry and neural circuitry of sleep regulation is critical for understanding sleep and various sleep disorders. Fruit flies display sleep-like behavior, sharing essential features with sleep of vertebrate. In the fruit fly's central brain, the mushroom body (MB) has been highlighted as a sleep center; however, its neurochemical nature remains unclear, and whether it promotes sleep or wake is still a topic of controversy. DESIGN: We used a video recording system to accurately monitor the locomotor activity and sleep status. Gene expression was temporally and regionally manipulated by heat induction and the Gal4/UAS system. MEASUREMENTS AND RESULTS: We found that expressing pertussis toxin (PTX) in the MB by c309-Gal4 to block Go activity led to unique sleep defects as dramatic sleep increase in daytime and fragmented sleep in nighttime. We narrowed down the c309-Gal4 expressing brain regions to the MB α/ß core neurons that are responsible for the Go-mediated sleep effects. Using genetic tools of neurotransmitter-specific Gal80 and RNA interference approach to suppress acetylcholine signal, we demonstrated that these MB α/ß core neurons were cholinergic and sleep-promoting neurons, supporting that Go mediates an inhibitory signal. Interestingly, we found that adjacent MB α/ß neurons were also cholinergic but wake-promoting neurons, in which Go signal was also required. CONCLUSION: Our findings in fruit flies characterized a group of sleep-promoting neurons surrounded by a group of wake-promoting neurons. The two groups of neurons are both cholinergic and use Go inhibitory signal to regulate sleep.

Transformation of odor selectivity from projection neurons to single mushroom body neurons mapped with dual-color calcium imaging.

Although the response properties of most neurons are, to a large extent, determined by the presynaptic inputs that they receive, comprehensive functional characterization of the presynaptic inputs of a single neuron remains elusive. Toward this goal, we introduce a dual-color calcium imaging approach that simultaneously monitors the responses of a single postsynaptic neuron together with its presynaptic axon terminal inputs in vivo. As a model system, we applied the strategy to the feed-forward connections from the projection neurons (PNs) to the Kenyon cells (KCs) in the mushroom body of Drosophila and functionally mapped essentially all PN inputs for some of the KCs. We found that the output of single KCs could be well predicted by a linear summation of the PN input signals, indicating that excitatory PN inputs play the major role in generating odor-selective responses in KCs. When odors failed to activate KC output, local calcium transients restricted to individual postsynaptic sites could be observed in the KC dendrites. The response amplitudes of the local transients often correlated linearly with the presynaptic response amplitudes, allowing direct assay of the strength of single synaptic sites. Furthermore, we found a scaling relationship between the total number of PN terminals that a single KC received and the average synaptic strength of these PN-KC synapses. Our strategy provides a unique perspective on the process of information transmission and integration in a model neural circuit and may be broadly applicable for the study of the origin of neuronal response properties.

Parallel pathways for cross-modal memory retrieval in Drosophila.

Memory-retrieval processing of cross-modal sensory preconditioning is vital for understanding the plasticity underlying the interactions between modalities. As part of the sensory preconditioning paradigm, it has been hypothesized that the conditioned response to an unreinforced cue depends on the memory of the reinforced cue via a sensory link between the two cues. To test this hypothesis, we studied cross-modal memory-retrieval processing in a genetically tractable model organism, Drosophila melanogaster. By expressing the dominant temperature-sensitive shibire(ts1) (shi(ts1)) transgene, which blocks synaptic vesicle recycling of specific neural subsets with the Gal4/UAS system at the restrictive temperature, we specifically blocked visual and olfactory memory retrieval, either alone or in combination; memory acquisition remained intact for these modalities. Blocking the memory retrieval of the reinforced olfactory cues did not impair the conditioned response to the unreinforced visual cues or vice versa, in contrast to the canonical memory-retrieval processing of sensory preconditioning. In addition, these conditioned responses can be abolished by blocking the memory retrieval of the two modalities simultaneously. In sum, our results indicated that a conditioned response to an unreinforced cue in cross-modal sensory preconditioning can be recalled through parallel pathways.

The GABA system regulates the sparse coding of odors in the mushroom bodies of Drosophila.

In the mushroom bodies (MBs) of Drosophila, an analogue of the mammalian olfactory cortex, olfactory stimuli are sparsely encoded by Kenyon cells (KCs) that exhibit a high level of odor selectivity. Sparse coding of olfactory stimuli has significant advantages for maximizing the discrimination power and storage capacity of MBs. The inhibitory gamma-aminobutyric acid (GABA) system is important for regulating information processing in MBs, but its specific role in the sparse coding of odors is unclear. In this study, we investigated the role of the GABA system in the sparse coding of odors using an in vivo calcium imaging strategy, which allowed us to measure the activity of the KC population at single cell resolution while the components of the GABA system were genetically manipulated. We found that the down-regulation of GABAA but not GABAB receptors in KCs reduced the sparseness of odor representations in the MB, as shown by an increase in the population response probability and decrease in the odor selectivity of single KCs. Furthermore, the down-regulation of GABA synthesis in a pair of large GABAergic neurons innervating the entire MB reduced the sparseness of odor representations in KCs. In conclusion, the sparse coding of odors in MBs is regulated by a pair of GABAergic neurons through the GABAA receptors on KCs, thus demonstrating a specific role of the inhibitory GABA system on information processing in the MB.

Distinct acute zones for visual stimuli in different visual tasks in Drosophila.

The fruit fly Drosophila melanogaster has a sophisticated visual system and exhibits complex visual behaviors. Visual responses, vision processing and higher cognitive processes in Drosophila have been studied extensively. However, little is known about whether the retinal location of visual stimuli can affect fruit fly performance in various visual tasks. We tested the response of wild-type Berlin flies to visual stimuli at several vertical locations. Three paradigms were used in our study: visual operant conditioning, visual object fixation and optomotor response. We observed an acute zone for visual feature memorization in the upper visual field when visual patterns were presented with a black background. However, when a white background was used, the acute zone was in the lower visual field. Similar to visual feature memorization, the best locations for visual object fixation and optomotor response to a single moving stripe were in the lower visual field with a white background and the upper visual field with a black background. The preferred location for the optomotor response to moving gratings was around the equator of the visual field. Our results suggest that different visual processing pathways are involved in different visual tasks and that there is a certain degree of overlap between the pathways for visual feature memorization, visual object fixation and optomotor response.