Colour Vision: A Fresh View of Lateral Inhibition in Drosophila

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Dispatches 6. Dominguez-Escobar, J., Chastanet, A., Crevenna, A.H., Fromion, V., Wedlich-Soldner, R., and Carballido-Lopez, R. (2011). Processive movement of MreB-associated cell wall biosynthetic complexes in bacteria. Science 333, 225–228. 7. Garner, E.C., Bernard, R., Wang, W., Zhuang, X., Rudner, D.Z., and Mitchison, T. (2011). Coupled, circumferential motions of the cell wall synthesis machinery and MreB filaments in B. subtilis. Science 333, 222–225. 8. van Teeffelen, S., Wang, S., Furchtgott, L., Huang, K.C., Wingreen, N.S., Shaevitz, J.W., and Gitai, Z. (2011). The bacterial actin MreB rotates, and rotation depends on cell-wall assembly. Proc. Natl. Acad. Sci. USA 108, 15822–15827. 9. Du, S., and Lutkenhaus, J. (2017). Assembly and activation of the Escherichia coli divisome. Mol. Microbiol. 105, 177–187. 10. Bisson-Filho, A.W., Hsu, Y.P., Squyres, G.R., Kuru, E., Wu, F., Jukes, C., Sun, Y., Dekker, C.,

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12. den Blaauwen, T., de Pedro, M.A., NguyenDisteche, M., and Ayala, J.A. (2008). Morphogenesis of rod-shaped sacculi. FEMS Microbiol. Rev. 32, 321–344. 13. Polz, M.F., Felbeck, H., Novak, R., Nebelsick, M., and Ott, J.A. (1992). Chemoautotrophic, sulfur-oxidizing symbiotic bacteria on marine nematodes: Morphological and biochemical characterization. Microb. Ecol. 24, 313–329. 14. Leisch, N., Pende, N., Weber, P.M., GruberVodicka, H.R., Verheul, J., Vischer, N.O., Abby, S.S., Geier, B., den Blaauwen, T., and Bulgheresi, S. (2016). Asynchronous division

16. Leisch, N., Verheul, J., Heindl, N.R., GruberVodicka, H.R., Pende, N., den Blaauwen, T., and Bulgheresi, S. (2012). Growth in width and FtsZ ring longitudinal positioning in a gammaproteobacterial symbiont. Curr. Biol. 22, R831–R832. 17. Kuru, E., Tekkam, S., Hall, E., Brun, Y.V., and van Nieuwenhze, M.S. (2015). Synthesis of fluorescent D-amino acids and their use for probing peptidoglycan synthesis and bacterial growth in situ. Nat. Protoc. 10, 33–52.

Colour Vision: A Fresh View of Lateral Inhibition in Drosophila Kit D. Longden Reiser Lab, HHMI Janelia Research Campus, 19700 Helix Drive, Ashburn, VA 20147, USA Correspondence: [email protected] https://doi.org/10.1016/j.cub.2018.02.052 Twitter: @TheFlySide

A recent study reports a novel form of lateral inhibition between photoreceptors supporting colour vision in the vinegar fly, Drosophila melanogaster. Take a photograph into the sun and you get a sharp lesson in just how adeptly our eyes compensate for the phenomenal range of photon intensities in daylight. While the sky may be blue and our loved ones smile patiently, the camera bleaches the colour and washes out detail. The problem solved by photoreceptors is to convey the full sweep of wavelengths and intensities of light using membrane potentials and synapses of limited scope. A key step is to remove the signal mean to encode relative rather than absolute changes. A new study by Schnaitmann et al. [1] has carefully characterised mechanisms involved in this step in the colour vision circuitry of Drosophila. This new study [1] is important because it is a first description of colour-opponent neural coding in a species with the tools

and resources to trace the circuits of colour behaviours throughout the central brain [2–4], something that would be an achievement in any species [5]. Colouropponency is the property of being excited by one wavelength of light and inhibited by another, a calling card for colour information processing. The study is also a first description of direct synaptic inhibition between photoreceptors used for daylight colour vision. It demonstrates that the photoreceptors conveying wavelengthspecific information are subject to initial contrast-enhancing filtering in flies, contrary to expectations until only very recently [2,3,6]. Thanks to their replicable circuitry and accessible cells, fly studies have a terrific track record of providing insights into the computations of contrastive coding in the visual periphery,

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notably the seminal study that the title of this dispatch pays homage to [7]. To understand why the findings were surprising, we need to take in some anatomical details of the fly’s optic lobes (Figure 1A). Under every facet of the fly’s compound eye eight photoreceptors look out onto the world: the outer six, R1–R6, are sensitive to UV and cyan, and transmit luminance signals to the first region of the optic lobe, the lamina; the inner two, R7 and R8, encode the additional wavelength information needed for colour vision and project directly to the second optic lobe region, the medulla. Away from the edges of the eye and specialisations for polarisation vision, pairs of R7 and R8 cells are stochastically determined during adult development to be one of two flavours, known as ‘pale’ and ‘yellow’ for

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how they appear when illuminated with blue and yellow light: in pale pairs, R7 is selective for short wavelength UV and R8 for blue; in yellow pairs, R7 is selective for long wavelength UV and R8 for green [8]. This organisation of dedicated photoreceptor pathways for luminance and colour vision is a trait that flies share with many other insects, crustaceans and birds. It contrasts with the retinal organisation of colour-sighted mammals, reptiles and amphibians, in which individual cone signals are scaled by lateral inhibition before contributing either to the processing of luminance or of colour. Because the axons of R7 and R8 photoreceptors pass through the lamina without forming synapses [9], it was presumed that they were unfiltered and directly fed into wavelength-specific circuits and also colour vision pathways [10]. To update this picture, Schnaitmann et al. [1] first recorded simplified spectral tuning curves of the pale and yellow R7–R8s, using light emitting diodes (LEDs) with narrowband spectra and genetically encoded calcium indicators of neural activity expressed in cell-specific driver lines. To pick out whether the cells are inhibited by non-preferred wavelengths, the authors re-recorded the spectral tuning curve while an LED close to their preferred wavelength was also on. For all four pale and yellow R7–R8 cells, responses to their preferred LED were suppressed by light from the opposing end of the spectrum: green and cyan suppressed R7, and UV suppressed R8. The question now was to identify the mechanisms. There is spectral inhibition between photoreceptors in a wide variety of insects, including the honeybee, butterfly, locust and dragonfly [11–14]. A leading candidate mechanism to account for these effects was the local field potential: the current from a photoreceptor axon depolarises the extracellular space, and so alters the driving force of return currents across either high resistance glial membranes or less active photoreceptors [14,15]. A mechanism of this type plays a role in the horizontal cell inhibition of photoreceptors in many vertebrate retinas [16]. By recording from lamina monopolar cells with chemically permeabilised membranes, Weckstro¨m and Laughlin [15] neatly measured the

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Figure 1. Drosophila visual circuitry and schematic illustrations of signal interactions between photoreceptors. (A) A single lamina cartridge is innervated by R1–R6 cells occupying separate, neighbouring retinal cartridges and sharing the same optical axis, an organisation known as the neural superposition principle [8]. In the lamina, R1–R6 axons synapse with glia and lamina monopolar cells (LMCs), in a complex synaptic column that also includes feedback from medulla cells (Lawf, T & C cells) and lamina neurons (Lai, Lat) [9]. Glia form high resistance barriers at the retina basal membrane (thick black line) and between retina and lamina cartridges (dashed lines) [15]. Pairs of pale (p) or yellow (y) R7–R8 axons travel together through the lamina in junctions between glia inter-cartridge barriers. Various distal medulla cells (Dm) are well placed to form synapses with R7-8 and provide feedback [4]. Transmedulla (Tm), visual projection neurons to the mushroom bodies (VPN-MB), and other cells integrate processed R7–R8 signals in the medulla and pass them to the central brain [5]. (B) Screening of R8 spectral tuning by R7 [19]. R7s absorb UV light before it reaches the R8, reducing the UV sensitivity of R8 cells. (C) Effect of field potential on photoreceptor signalling. Curves are redrawn from Figure 3 of [15], recordings in the blowfly Calliphora vicina. Dashed line: R1–R6 cell membrane potential at the axon terminal. Solid line: the difference between the potential in the axon terminal and the extracellular space. The direct current component of the signal is much reduced. (D) Contributions of R7–R8 to the spectral tuning of LMCs. R1–R6 lack sensitivity to blue light (dashed curve). R7–R8 extend the spectral range of LMCs into blue, short UV and green (solid curve) [6]. (E) Impact of lateral inhibition on the spectral tuning of pale R7 and R8, as indicated by the study of Schnaitmann et al. [1].

extracellular potential without disturbing the glial compartment, and showed that it is effective in reducing the directcurrent component of the photoreceptor depolarisation (Figure 1C). The anticipated consequence for photoreceptors with different spectral

sensitivities is to reduce the responses to common wavelengths [12,14]. Whether the field potential modulates R7–R8 is an open question, because these cells occupy their own glial compartment, set apart from the R1–R6 axons (Figure 1A). In flies, the only previous hint of spectral

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Dispatches lateral inhibition between photoreceptors had been an intriguing report that UV and green light excite R1–R6 more individually than together [17]. To identify the mechanisms in Drosophila, Schnaitmann et al. [1] switched to genetically rescuing the function of specific R1–R8 cells in mutants with otherwise blind photoreceptors. They found that functional pairs of pale or yellow R7–R8s were necessary to clearly rescue lateral inhibition. In particular, rescuing the function of R1–R6 did not affect the antagonistic responses in R7–R8, indicating that the field potential from R1–R6 and lamina cell activity alone is insufficient. Could R7–R8 inhibit each other? There are synapses between R7 and R8 that are revealed by electron microscope (EM) images [2], but the puzzle here was the postsynaptic receptors. All R1–R8 cells use histamine as a neurotransmitter, and only two histamine-gated chloride channels have been found in the optic lobes, Ort and HisCl1 (also known as HCLA and HCLB): Ort is expressed in a number of cells, but not photoreceptors, while a genetic reporter of HisCl1 shows up in lamina glia, but not the medulla [3,18]. Schnaitmann et al. [1] generated a new, more sensitive fosmid-based genetic reporter of HisCl1 that is driven by more of the regulatory sequences of the hiscl1 gene [1]. With this powerfully sensitive tool, they could show that both R7 and R8 express HisCl1, opening up the possibility that they mutually inhibit each other. If so, the prediction is that direct inhibition between R7 and R8 should require HisCl1 receptors, while indirect R7–R8 inhibition should require the Ort receptors found in other postsynaptic cell partners. To test this idea, they measured responses of individual R7–R8s in mutants that lacked functional copies of one or both of the ort or hiscl1 genes. They found that R7s could be inhibited by R8 or another pathway, because they required either ort or hiscl1 to be inhibited by blue and green. In contrast R8s were only inhibited by the simultaneous activity of R7 and other cells, because they needed both ort and hiscl1 to be inhibited by UV. These results fit with the number of synapses between R7 and R8. In the most recent EM medulla dataset [2], a landmark effort covering seven adjoining columns,

there are 20–31 synapses from R8 to R7 per column, and 0–2 synapses from R7 to R8. Although the synapses from R7 to R8 are few, they are functional. To show this, Schnaitmann et al. [1] restored HisCl1 receptors in R7-8s of ort or hiscl1 mutants — for these flies there can be no other source of lateral inhibition other than direct input, because they otherwise lack functional ort and hiscl1 genes. When they restored HisCl1 function in R8 cells, the spectral response to combined LEDs was consistent with R7 inhibiting R8 [1]. It is also helpful to remember that other mechanisms are at play here: R7 suppresses the responses of R8 to UV light by absorbing the UV light itself, a process known as screening [19] (Figure 1B). So R7 encodes more redundant spectral information than R8, and as a result, R8 can usefully reduce redundant responses to long wavelengths in R7, but there is little UV signal in R8 to be squashed by R7 inhibition. Overall, these data support direct inhibition from R8 to R7, and indirect pathways for inhibition of R8 by R7, an allocation that makes sense given the imbalance of shared wavelength redundancy in the R7–R8 cells. Why do R7–R8 wait until layer 3 of the medulla to inhibit each other [2]? Photoreceptors are energy-intensive cells, and there are plenty of opportunities to subtract costly redundancies as they travel in close concert across the lamina. One answer may be that spectral information is transferred from R7–R8 to the lamina monopolar cells, which drive the luminance-based motion pathway [6]. The spectral input from R7–R8 gives motion responses sensitivity to blue light and a broader range of wavelengths (Figure 1D). The mechanism for this connection is not known — there are no known lamina synaptic contacts [9] — so feedback of the uninhibited signal from the distal layers of the medulla is a possible route. What does the fly do that depends on this lateral inhibition between photoreceptors? Intuitively, one would expect that the first recorded neural correlates of colour-opponent coding in flies would have consequences for colour behaviour, but there is more going on. To make a colour-opponent signal useful for distinguishing wavelengths regardless of intensity — a benchmark for colour

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vision — the opponent signals are scaled first, and then an appropriate weight of inhibition can be struck so that the rectified signal gives the correct wavelength classification. It is for this reason, for example, that vertebrate colour-opponent cones support colour vision when bipolar cells integrate similar inputs, and drive luminance processing in other cells [16]. From this perspective, colour-opponency in photoreceptors reduces signal redundancy to ensure the efficient use of synapses, and may or may not be used by a colour-opponent cell to successfully discriminate colours. R7–R8 outputs support wavelength-specific behaviours as well as colour vision [3], so the counterintuitive prediction is that wavelength-specific phototaxis should be modulated by other wavelengths. For colour vision, flies compare different combinations of photoreceptors [10], and the corresponding circuits may have the capacity to compensate for a loss of direct inhibition between R7–R8. The large range of signals experienced by photoreceptors means that the relative contribution of any one filtering mechanism will not be fixed. Rather, mechanisms will be flexibly recruited to match the degree of redundancy in the visual scene, across space, time, wavelengths, polarisation and illumination intensity [7]. In addition, the filtering required in the visual periphery needs to match the demands of the behavioural task, the physiological state of the animal, and the affordances of the visual scene. As in the vertebrate retina, the visual periphery of flies has a sufficient variety of cell types to do far more than just statically filter the visual scene [20]. Addressing how different mechanisms are recruited will be key to understanding the basis of colour vision in flies, and Schnaitmann et al. [1] have laid a foundational stone for this enterprise. REFERENCES 1. Schnaitmann, C., Haikala, V., Abraham, E., Oberhauser, V., Thestrup, T., Griesbeck, O., and Reiff, D.F. (2018). Color processing in the early visual system of Drosophila. Cell 172, 318–330.e18. 2. Takemura, S.Y., Xu, C.S., Lu, Z., Rivlin, P.K., Parag, T., Olbris, D.J., Plaza, S., Zhao, T., Katz, W.T., Umayam, L., et al. (2015). Synaptic circuits and their variations within different columns in the visual system of Drosophila. Proc. Natl. Acad. Sci. USA 112, 13711–13716.

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9. Meinertzhagen, I.A., and O’Neil, S.D. (1991). Synaptic organization of columnar elements in the lamina of the wild type in Drosophila melanogaster. J. Comp. Neurol. 305, 232–263.

4. Nern, A., Pfeiffer, B.D., and Rubin, G.M. (2015). Optimized tools for multicolor stochastic labeling reveal diverse stereotyped cell arrangements in the fly visual system. Proc. Natl. Acad. Sci. USA 112, E2967–E2976.

10. Schnaitmann, C., Garbers, C., Wachtler, T., and Tanimoto, H. (2013). Color discrimination with broadband photoreceptors. Curr. Biol. 23, 2375–2382.

5. Longden, K.D. (2016). Central brain circuitry for color-vision-modulated behaviors. Curr. Biol. 26, R981–R988. 6. Wardill, T.J., List, O., Li, X., Dongre, S., McCulloch, M., Ting, C.-Y., O’Kane, C.J., Tang, S., Lee, C.-H., Hardie, R.C., et al. (2012). Multiple spectral inputs improve motion discrimination in the Drosophila visual system. Science 336, 925–931. 7. Srinivasan, M.V., Laughlin, S.B., and Dubs, A. (1982). Predictive coding: a fresh view of inhibition in the retina. Proc. R. Soc. Lond. B 216, 427–459. 8. Kolodkin, A.L., and Hiesinger, P.R. (2017). Wiring visual systems: common and divergent mechanisms and principles. Curr. Opin. Neurobiol. 42, 128–135.

11. Menzel, R., and Blakers, M. (1976). Colour receptors in the bee eye - morphology and spectral sensitivity. J. Comp. Physiol. A 108, 11–13. elja, L., Jahnke, R., and 12. Horridge, G.A., Marc , T. (1983). Single electrode studies on Matic the retina of the butterfly Papilio. J. Comp. Physiol. A 150, 271–294. 13. Yang, E.C., and Osorio, D. (1991). Spectral sensitivities of photoreceptors and lamina monopolar cells in the dragonfly, Hemicordulia tau. J. Comp. Physiol. A 169, 663–669. 14. Shaw, S.R. (1975). Retinal resistance barriers and electrical lateral inhibition. Nature 255, 480–483. 15. Weckstro¨m, M., and Laughlin, S. (2010). Extracellular potentials modify the transfer of information at photoreceptor output synapses

in the blowfly compound eye. J. Neurosci. 30, 9557–9566. 16. Thoreson, W.B., and Mangel, S.C. (2012). Lateral interactions in the outer retina. Prog. Retin. Eye Res. 31, 407–441. 17. McCann, G.D., Fargason, R.D., and Shantz, V.T. (1977). The response properties of retinula cells in the fly Calliphora erythrocephala as a function of the wavelength and polarization properties of visible and ultraviolet light. Biol. Cybern. 26, 93–107. 18. Pantazis, A., Segaran, A., Liu, C.H., Nikolaev, A., Rister, J., Thum, A.S., Roeder, T., Semenov, E., Juusola, M., and Hardie, R.C. (2008). Distinct roles for two histamine receptors (hclA and hclB) at the Drosophila photoreceptor synapse. J. Neurosci. 28, 7250–7259. , G. 19. Stavenga, D.G., Wehling, M.F., and Belusic (2017). Functional interplay of visual, sensitizing and screening pigments in the eyes of Drosophila and other red-eyed dipteran flies. J. Physiol. 595, 5481–5494. 20. Gollisch, T., and Meister, M. (2010). Eye smarter than scientists believed: neural computations in circuits of the retina. Neuron 65, 150–164.

Paleoneurology: A Sight for Four Eyes Lawrence M. Witmer Department of Biomedical Sciences, Ohio University Heritage College of Osteopathic Medicine, Ohio Center for Ecology and Evolutionary Studies, Athens, OH 45701, USA Correspondence: [email protected] https://doi.org/10.1016/j.cub.2018.02.071

The ‘third eye’ of the pineal complex is a curious component of the vertebrate brain associated with light sensation and melatonin production. A fossil lizard with a ‘fourth eye’ now calls for a reinterpretation of pineal evolution. ‘‘The part of the body in which the soul directly exercises its functions is not the heart at all, or the whole of the brain. It is rather.a certain very small gland [the pineal organ] situated in the middle of the brain’s substance.’’  Descartes, The Passions of Rene the Soul, 1649 [1] It’s perhaps understandable that our knowledge of the function and evolution of the pineal organ has been murky, because, as Descartes noted, it is indeed a very small structure buried deep within the brain of humans and other mammals. Although

perhaps few people today subscribe to Descartes’ view of the pineal organ as the ‘seat of the soul’, it retains an almost mystical quality in some quarters. Pop cultural references abound, suggesting ways (and selling products, of course) to ‘activate the pineal’ with cannabis or ‘harmonic sound wave technology’ to achieve enlightenment (psychedelic or spiritual). Fortunately, there is also a rich scientific literature on the neuroscience and clinical neuroendocrinology of the pineal gland, revealing that the organ may have less to do with the soul and enlightenment and more to do with responding to light/dark cycles and

mediating circadian and seasonal rhythms via production of the hormone melatonin [2]. The mammalian pineal organ is complicated enough, but when other vertebrates are taken into account, we now speak of a ‘pineal complex’ involving multiple organs that emphasize the photosensory component. That is, they often are ‘eyes’ with structures comparable to the cornea, lens, retina, and visual pigments (opsins) that we typically associate with our normal, lateral eyes and which breach the skull to appear on the top of the head [3,4]. In fact, when considering the pineal complex of vertebrates we now need to distinguish between ‘lateral eyes’

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