Patterning the Retina of Drosophila Melanogaster for Color and Polarized Light Vision

Across the animal kingdom, specialized sensory epithelia are used for photoreception, allowing individuals to interact with their environment based on visual cues. Generally, neuronal photoreceptor cells (PRs) are organized in the retina, a specialized part of body tissue exposed to the outside world, and transform the energy of incoming electromagnetic radiation into neuronal excitation. This process depends on the large family of opsin proteins which are required in PRs of all animal species. This lead to the theory, that the very divergent eye structures may share a common ancestor, although they most likely arose several times independently during evolution. PRs transmit their electrical excitation to higher order neurons, which are organized in the brain of the animal. How the brain then integrates the incoming signals from a multitude of PRs to reproduce a reliable representation of the world remains one of the central questions of neurobiology. Most animals can extract different kinds of visual information from their environment. Besides detecting the shape and movement of objects, additional qualities like color or degree of polarization can also be distinguished. In most cases, different classes of PRs are used for each of these visual tasks. For instance, color discrimination is achieved by comparing the outputs of PRs having different spectral sensitivity, as they express different opsin molecules. In humans, three different subclasses of so-called cone PRs, are specialized to absorb light of either short, medium or long wavelengths, corresponding to blue, green or red colors, respectively. Loss of any one of these PR classes leads to a dramatic impairment in the ability to discriminate between colors. Cones are most highly concentrated in the center of the retina (fovea), where the three subclasses form a random mosaic. Much remains to be understood about how different PR subtypes choose expression of their opsin and how they distribute in the retina. The developing eye of the fruitfly Drosophila melanogaster was used here as a model system to investigate both nature and regulation of the different strategies involved in retinal patterning. The adult Drosophila eye consists of ~800 stereotypical unit eyes (ommatidia), each containing exactly 8 PRs (R1-R8). The six 'outer PRs' (R1-R6) are molecularly identical in all ommtidia as they always express the same opsin. They form a separate visual system contributing to the detection of shapes and motion. The morphological and molecular differences between inner PRs (R7 and R8) from different ommatidia leads to the formation of a retinal mosaic in Drosophila. Three ommatidial subtypes can be distinguished: while the ommatidia of the 'dorsal rim area' (DRA) are always found precisely localized in the dorsal periphery, the remaining 'pale' and yellow' ommatidia are distributed stochastically through the rest of the retina. Only DRA ommatidia can be identified based on morphologic criteria, as these ommatidia form a polarizing filter which the fly uses to measure e-vector orientation of polarized sunlight for navigational purposes. The remaining two ommatidial subtypes are believed to serve color discrimination. They can only be identified based on the combination of opsins their inner PRs express. In order to identify genes and pathways involved in generating the retinal mosaic in Drosophila, a GAL4 enhancer trap screen was performed. Genes exhibiting expression patterns similar to inner PR opsins were analyzed genetically. The homeodomain transcription factor Homothorax (Hth) was identified as the key regulator of DRA specification. Hth is both necessary and sufficient for the formation of the polarization sensors. During pupal development, positional information provided by the diffusible morphogen Wingless (Wg), as well as the dorsal selector genes of the Iroquois complex (IRO-C) and the gene optomotorblind (omb) get integrated, leading to the specific induction of Hth expression in inner PRs of prospective DRA ommatidia. In contrast to this localized specification approach, stochastic expression of the Drosophila arylhydrocarbon receptor Spineless (Ss) in a large subset of pupal R7 cells is responsible for the specification of color ommtidia. Ss is both necessary and sufficient to induce the 'yellow' R7 fate (yR7). Ss was therefore identified as the key effector of a stochastic specification approach. How stochastic expression of Ss is regulated, remains obscure. However, an activating effect of the Notch (N) pathway on yR7 specification indicates that retinal patterning in Drosophila might combine the inductive effects of both wg and N signaling once again during pupal development. Further investigation of the regulatory relationship between Hth and Ss (or Wg and N) will provide a better understanding how retinal patterning contributes to the integration of different kinds of visual information

Heading choices of flying Drosophila under changing angles of polarized light

Many navigating insects include the celestial polarization pattern as an additional visual cue to orient their travels. Spontaneous orientation responses of both walking and flying fruit flies (Drosophila melanogaster) to linearly polarized light have previously been demonstrated. Using newly designed modular flight arenas consisting entirely of off-the-shelf parts and 3D-printed components we present individual flying flies with a slow and continuous rotational change in the incident angle of linear polarization. Under such open-loop conditions, single flies choose arbitrary headings with respect to the angle of polarized light and show a clear tendency to maintain those chosen headings for several minutes, thereby adjusting their course to the slow rotation of the incident stimulus. Importantly, flies show the tendency to maintain a chosen heading even when two individual test periods under a linearly polarized stimulus are interrupted by an epoch of unpolarized light lasting several minutes. Finally, we show that these behavioral responses are wavelength-specific, existing under polarized UV stimulus while being absent under polarized green light. Taken together, these findings provide further evidence supporting Drosophila’s abilities to use celestial cues for visually guided navigation and course correction

Parallel Visual Pathways with Topographic versus Nontopographic Organization Connect the Drosophila Eyes to the Central Brain

One hallmark of the visual system is a strict retinotopic organization from the periphery toward the central brain, where functional imaging in Drosophila revealed a spatially accurate representation of visual cues in the central complex. This raised the question how, on a circuit level, the topographic features are implemented, as the majority of visual neurons enter the central brain converge in optic glomeruli. We discovered a spatial segregation of topographic versus non-topographic projections of distinct classes of medullo-tubercular (MeTu) neurons into a specific visual glomerulus, the anterior optic tubercle (AOTU). These parallel channels synapse onto different tubercular-bulbar (TuBu) neurons, which in turn relay visual information onto specific central complex ring neurons in the bulb neuropil. Hence, our results provide the circuit basis for spatially accurate representation of visual information and highlight the AOTU's role as a prominent relay station for spatial information from the retina to the central brain

Behavioral responses of free-flying Drosophila melanogaster to shiny, reflecting surfaces

Active locomotion plays an important role in the life of many animals, permitting them to explore the environment, find vital resources, and escape predators. Most insect species rely on a combination of visual cues such as celestial bodies, landmarks, or linearly polarized light to navigate or orient themselves in their surroundings. In nature, linearly polarized light can arise either from atmospheric scattering or from reflections off shiny non-metallic surfaces like water. Multiple reports have described different behavioral responses of various insects to such shiny surfaces. Our goal was to test whether free-flying Drosophila melanogaster, a molecular genetic model organism and behavioral generalist, also manifests specific behavioral responses when confronted with such polarized reflections. Fruit flies were placed in a custom-built arena with controlled environmental parameters (temperature, humidity, and light intensity). Flight detections and landings were quantified for three different stimuli: a diffusely reflecting matt plate, a small patch of shiny acetate film, and real water. We compared hydrated and dehydrated fly populations, since the state of hydration may change the motivation of flies to seek or avoid water. Our analysis reveals for the first time that flying fruit flies indeed use vision to avoid flying over shiny surfaces

Synaptic targets of photoreceptors specialized to detect color and skylight polarization in Drosophila

Color and polarization provide complementary information about the world and are detected by specialized photoreceptors. However, the downstream neural circuits that process these distinct modalities are incompletely understood in any animal. Using electron microscopy, we have systematically reconstructed the synaptic targets of the photoreceptors specialized to detect color and skylight polarization in Drosophila, and we have used light microscopy to confirm many of our findings. We identified known and novel downstream targets that are selective for different wavelengths or polarized light, and followed their projections to other areas in the optic lobes and the central brain. Our results revealed many synapses along the photoreceptor axons between brain regions, new pathways in the optic lobes, and spatially segregated projections to central brain regions. Strikingly, photoreceptors in the polarization-sensitive dorsal rim area target fewer cell types, and lack strong connections to the lobula, a neuropil involved in color processing. Our reconstruction identifies shared wiring and modality-specific specializations for color and polarization vision, and provides a comprehensive view of the first steps of the pathways processing color and polarized light inputs

Iroquois Complex Genes Induce Co-Expression of rhodopsins in Drosophila

The Drosophila eye is a mosaic that results from the stochastic distribution of two ommatidial subtypes. Pale and yellow ommatidia can be distinguished by the expression of distinct rhodopsins and other pigments in their inner photoreceptors (R7 and R8), which are implicated in color vision. The pale subtype contains ultraviolet (UV)-absorbing Rh3 in R7 and blue-absorbing Rh5 in R8. The yellow subtype contains UV-absorbing Rh4 in R7 and green-absorbing Rh6 in R8. The exclusive expression of one rhodopsin per photoreceptor is a widespread phenomenon, although exceptions exist. The mechanisms leading to the exclusive expression or to co-expression of sensory receptors are currently not known. We describe a new class of ommatidia that co-express rh3 and rh4 in R7, but maintain normal exclusion between rh5 and rh6 in R8. These ommatidia, which are localized in the dorsal eye, result from the expansion of rh3 into the yellow-R7 subtype. Genes from the Iroquois Complex (Iro-C) are necessary and sufficient to induce co-expression in yR7. Iro-C genes allow photoreceptors to break the “one receptor–one neuron” rule, leading to a novel subtype of broad-spectrum UV- and green-sensitive ommatidia

The development and function of neuronal subtypes processing color and skylight polarization in the optic lobes of Drosophila melanogaster

The retinal mosaics of many insects contain different ommatidial subtypes harboring photoreceptors that are both molecularly and morphologically specialized for comparing between different wavelengths versus detecting the orientation of skylight polarization. The neural circuits underlying these different inputs and the characterization of their specific cellular elements are the subject of intense research. Here we review recent progress on the description of both assembly and function of color and skylight polarization circuitry, by focusing on two cell types located in the distal portion of the medulla neuropil of the fruit fly Drosophila melanogaster's optic lobes, called Dm8 and Dm9. In the main part of the retina, Dm8 cells fall into two molecularly distinct subtypes whose center becomes specifically connected to either one of randomly distributed ‘pale’ or ‘yellow’ R7 photoreceptor fates during development. Only in the ‘dorsal rim area’ (DRA), both polarization-sensitive R7 and R8 photoreceptors are connected to different Dm8-like cell types, called Dm-DRA1 and Dm-DRA2, respectively. An additional layer of interommatidial integration is introduced by Dm9 cells, which receive input from multiple neighboring R7 and R8 cells, as well as providing feedback synapses back into these photoreceptors. As a result, the response properties of color-sensitive photoreceptor terminals are sculpted towards being both maximally decorrelated, as well as harboring several levels of opponency (both columnar as well as intercolumnar). In the DRA, individual Dm9 cells appear to mix both polarization and color signals, thereby potentially serving as the first level of integration of different celestial stimuli. The molecular mechanisms underlying the establishment of these synaptic connections are beginning to be revealed, by using a combination of live imaging, developmental genetic studies, and cell type-specific transcriptomics

Insect Responses to Linearly Polarized Reflections: Orphan Behaviors Without Neural Circuits

The e-vector orientation of linearly polarized light represents an important visual stimulus for many insects. Especially the detection of polarized skylight by many navigating insect species is known to improve their orientation skills. While great progress has been made towards describing both the anatomy and function of neural circuit elements mediating behaviors related to navigation, relatively little is known about how insects perceive non-celestial polarized light stimuli, like reflections off water, leaves, or shiny body surfaces. Work on different species suggests that these behaviors are not mediated by the “Dorsal Rim Area” (DRA), a specialized region in the dorsal periphery of the adult compound eye, where ommatidia contain highly polarization-sensitive photoreceptor cells whose receptive fields point towards the sky. So far, only few cases of polarization-sensitive photoreceptors have been described in the ventral periphery of the insect retina. Furthermore, both the structure and function of those neural circuits connecting to these photoreceptor inputs remain largely uncharacterized. Here we review the known data on non-celestial polarization vision from different insect species (dragonflies, butterflies, beetles, bugs and flies) and present three well-characterized examples for functionally specialized non-DRA detectors from different insects that seem perfectly suited for mediating such behaviors. Finally, using recent advances from circuit dissection in Drosophila melanogaster, we discuss what types of potential candidate neurons could be involved in forming the underlying neural circuitry mediating non-celestial polarization vision

Non-celestial polarization vision in arthropods

Most insects can detect the pattern of polarized light in the sky with the dorsal rim area in their compound eyes and use this visual information to navigate in their environment by means of 'celestial' polarization vision. 'Non-celestial polarization vision', in contrast, refers to the ability of arthropods to analyze polarized light by means of the 'main' retina, excluding the dorsal rim area. The ability of using the main retina for polarization vision has been attracting sporadic, but steady attention during the last decade. This special issue of the Journal of Comparative Physiology A presents recent developments with a collection of seven original research articles, addressing different aspects of non-celestial polarization vision in crustaceans and insects. The contributions cover different sources of linearly polarized light in nature, the underlying retinal and neural mechanisms of object detection using polarization vision and the behavioral responses of arthropods to polarized reflections from water.</p