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Color vision is an important sensory capability of humans and many animals. It relies on color opponent processing in visual circuits that gradually compare the signals of photoreceptors with different spectral sensitivities. In Drosophila, this comparison begins already in the presynaptic terminals of UV-sensitive R7 and longer wavelength-sensitive R8 inner photoreceptors that inhibit each other in the medulla. How downstream neurons process their signals is unknown. Here, we report that the second order medulla interneuron Dm8 is inhibited when flies are stimulated with UV light and strongly excited in response to a broad range of longer wavelength (VIS) stimuli. Inhibition to UV light is mediated by histaminergic input from R7 and expression of the histamine receptor ort in Dm8, as previously suggested. However, two additional excitatory inputs antagonize the R7 input. First, activation of R8 leads to excitation of Dm8 by non-canonical photoreceptor signaling and cholinergic neurotransmission in the visual circuitry. Second, activation of outer photoreceptors R1-R6 with broad spectral sensitivity causes excitation in Dm8 through the cholinergic medulla interneuron Mi1, which is known for its major contribution to the detection of spatial luminance contrast and visual motion. In summary, Dm8 mediates a second step in UV/VIS color opponent processing in Drosophila by integrating input from all types of photoreceptors. Our results demonstrate novel insights into the circuit integration of R1-R6 into color opponent processing and reveal that chromatic and achromatic circuitries of the fly visual system interact more extensively than previously thought.

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.

Color vision extracts spectral information by comparing signals from photoreceptors with different visual pigments. Such comparisons are encoded by color-opponent neurons that are excited at one wavelength and inhibited at another. Here, we examine the circuit implementation of color-opponent processing in the Drosophila visual system by combining two-photon calcium imaging with genetic dissection of visual circuits. We report that coloropponent processing of UVshort/blue and UVlong/green is already implemented in R7/R8 inner photoreceptor terminals of "pale'' and "yellow'' ommatidia, respectively. R7 and R8 photoreceptors of the same type of ommatidia mutually inhibit each other directly via HisCl1 histamine receptors and receive additional feedback inhibition that requires the second histamine receptor Ort. Color-opponent processing at the first visual synapse represents an unexpected commonality between Drosophila and vertebrates; however, the differences in the molecular and cellular implementation suggest that the same principles evolved independently.

The nervous system of seeing animals derives information about optic flow in two subsequent steps. First, local motion vectors are calculated from moving retinal images, and second, the spatial distribution of these vectors is analyzed on the dendrites of large downstream neurons. In dipteran flies, this second step relies on a set of motion-sensitive lobula plate tangential cells (LPTCs), which have been studied in great detail in large fly species. Yet, studies on neurons that convey information to LPTCs and neuroanatomical investigations that enable a mechanistic understanding of the underlying dendritic computations in LPTCs are rare. We investigated the subcellular distribution of nicotinic acetylcholine receptors (nAChRs) on two sets of LPTCs: vertical system (VS) and horizontal system (HS) cells in Drosophila melanogaster. In this paper, we describe that both cell types express Dalpha7-type nAChR subunits specifically on higher order dendritic branches, similar to the expression of gamma aminobutyric acid (GABA) receptors. These findings support a model in which directional selectivity of LPTCs is achieved by the dendritic integration of excitatory, cholinergic, and inhibitory GABA-ergic input from local motion detectors with opposite preferred direction. Nonetheless, whole-cell recordings in mutant flies without Dalpha7 nAChRs revealed that direction selectivity of VS and HS cells is largely retained. In addition, mutant LPTCs were responsive to acetylcholine and remaining nAChR receptors were labeled by alpha-bungarotoxin. These results in LPTCs with genetically manipulated excitatory input synapses suggest a robust cellular implementation of dendritic processing that warrants direction selectivity. The underlying mechanism that ensures appropriate nAChR-mediated synaptic currents and the functional implications of separate sets or heteromultimeric nAChRs can now be addressed in this system.

SummaryThe crystalline-like structure of the optic lobes of the fruit fly Drosophila melanogaster has made them a model system for the study of neuronal cell-fate determination, axonal path finding, and target selection. For functional studies, however, the small size of the constituting visual interneurons has so far presented a formidable barrier. We have overcome this problem by establishing in vivo whole-cell recordings [1] from genetically targeted visual interneurons of Drosophila. Here, we describe the response properties of six motion-sensitive large-field neurons in the lobula plate that form a network consisting of individually identifiable, directionally selective cells most sensitive to vertical image motion (VS cells [2, 3]). Individual VS cell responses to visual motion stimuli exhibit all the characteristics that are indicative of presynaptic input from elementary motion detectors of the correlation type [4, 5]. Different VS cells possess distinct receptive fields that are arranged sequentially along the eye's azimuth, corresponding to their characteristic cellular morphology and position within the retinotopically organized lobula plate. In addition, lateral connections between individual VS cells cause strongly overlapping receptive fields that are wider than expected from their dendritic input. Our results suggest that motion vision in different dipteran fly species is accomplished in similar circuitries and according to common algorithmic rules. The underlying neural mechanisms of population coding within the VS cell network and of elementary motion detection, respectively, can now be analyzed by the combination of electrophysiology and genetic intervention in Drosophila.

Neurons in the nervous system can change their functional properties over time. At present, there are no techniques that allow reliable monitoring of changes within identified neurons over repeated experimental sessions. We increased the signal strength of troponin C-based calcium biosensors in the low-calcium regime by mutagenesis and domain rearrangement within the troponin C calcium binding moiety to generate the indicator TN-XXL. Using in vivo two-photon ratiometric imaging, we show that TN-XXL exhibits enhanced fluorescence changes in neurons of flies and mice. TN-XXL could be used to obtain tuning curves of orientation-selective neurons in mouse visual cortex measured repeatedly over days and weeks. Thus, the genetically encoded calcium indicator TN-XXL allows repeated imaging of response properties from individual, identified neurons in vivo, which will be crucial for gaining new insights into cellular mechanisms of plasticity, regeneration and disease.

At the Drosophila melanogaster larval neuromuscular junction (NMJ), a motor neuron releases glutamate from 30–100 boutons onto the muscle it innervates. How transmission strength is distributed among the boutons of the NMJ is unknown. To address this, we created synapcam, a version of the Ca2+ reporter Cameleon. Synapcam localizes to the postsynaptic terminal and selectively reports Ca2+ influx through glutamate receptors (GluRs) with single-impulse and single-bouton resolution. GluR-based Ca2+ signals were uniform within a given connection (that is, a given bouton/postsynaptic terminal pair) but differed considerably among connections of an NMJ. A steep gradient of transmission strength was observed along axonal branches, from weak proximal connections to strong distal ones. Presynaptic imaging showed a matching axonal gradient, with higher Ca2+ influx and exocytosis at distal boutons. The results suggest that transmission strength is mainly determined presynaptically at the level of individual boutons, possibly by one or more factors existing in a gradient. *Note: In the version of this article initially published online, the second author’s name was misspelled. The correct spelling should be Dierk F Reiff. The error has been corrected in the HTML version of the article. This correction has been appended to the PDF and print versions.

Long-term synaptic plasticity may be associated with structural rearrangements within the neuronal circuitry1,2. Although the molecular mechanisms governing such activity-controlled morphological alterations are mostly elusive, polysomal accumulations at the base of developing dendritic spines3 and the activity-induced synthesis of synaptic components suggest that localized translation is involved during synaptic plasticity4,5. Here we show that large aggregates of translational components as well as messenger RNA of the postsynaptic glutamate receptor subunit DGluR-IIA6 are localized within subsynaptic compartments of larval neuromuscular junctions of Drosophila melanogaster. Genetic models of junctional plasticity7 and genetic manipulations using the translation initiation factors eIF4E8 and poly(A)-binding protein9 showed an increased occurrence of subsynaptic translation aggregates. This was associated with a significant increase in the postsynaptic DGluR-IIA protein levels and a reduction in the junctional expression of the cell-adhesion molecule Fasciclin II. In addition, the efficacy of junctional neurotransmission and the size of larval neuromuscular junctions were significantly increased. Our results therefore provide evidence for a postsynaptic translational control of long-term junctional plasticity.

In flies, the large tangential cells of the lobula plate represent an important processing center for visual navigation based on optic flow. Although the visual response properties of these cells have been well studied in blowflies, information on their synaptic organization is mostly lacking. Here we study the distribution of presynaptic release and postsynaptic inhibitory sites in the same set of cells in Drosophila melanogaster. By making use of transgenic tools and immunohistochemistry, our results suggest that HS and VS cells of Drosophila express gamma-aminobutyric acid (GABA) receptors in their dendritic region within the lobula plate, thus being postsynaptic to inhibitory input there. At their axon terminals in the protocerebrum, both cell types express synaptobrevin, suggesting the presence of presynaptic specializations there. HS- and VS-cell terminals additionally show evidence for postsynaptic GABAergic input, superimposed on this synaptic polarity. Our findings are in line with the general circuit for visual motion detection and receptive field properties as postulated from electrophysiological and optical recordings in blowflies, suggesting a similar functional organization of lobula plate tangential cells in the two species.