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Current Biology : CB Dec 2020Retinal rod and cone photoreceptors mediate vision in dim and bright light, respectively, by transducing absorbed photons into neural electrical signals. Their...
Retinal rod and cone photoreceptors mediate vision in dim and bright light, respectively, by transducing absorbed photons into neural electrical signals. Their phototransduction mechanisms are essentially identical. However, one difference is that, whereas a rod visual pigment remains stable in darkness, a cone pigment has some tendency to dissociate spontaneously into apo-opsin and retinal (the chromophore) without isomerization. This cone-pigment property is long known but has mostly been overlooked. Importantly, because apo-opsin has weak constitutive activity, it triggers transduction to produce electrical noise even in darkness. Currently, the precise dark apo-opsin contents across cone subtypes are mostly unknown, as are their dark activities. We report here a study of goldfish red (L), green (M), and blue (S) cones, finding with microspectrophotometry widely different apo-opsin percentages in darkness, being ∼30% in L cones, ∼3% in M cones, and negligible in S cones. L and M cones also had higher dark apo-opsin noise than holo-pigment thermal isomerization activity. As such, given the most likely low signal amplification at the pigment-to-transducin/phosphodiesterase phototransduction step, especially in L cones, apo-opsin noise may not be easily distinguishable from light responses and thus may affect cone vision near threshold.
Topics: Animals; Darkness; Goldfish; Light Signal Transduction; Models, Animal; Opsins; Patch-Clamp Techniques; Photic Stimulation; Retinal Cone Photoreceptor Cells; Single-Cell Analysis
PubMed: 33065015
DOI: 10.1016/j.cub.2020.09.062 -
Scientific Reports Apr 2017A main challenge in chemotherapy is to determine the in cellulo parameters modulating the drug concentration required for therapeutic action. It is absolutely urgent to...
A main challenge in chemotherapy is to determine the in cellulo parameters modulating the drug concentration required for therapeutic action. It is absolutely urgent to understand membrane permeation and intracellular concentration of antibiotics in clinical isolates: passing the membrane barrier to reach the threshold concentration inside the bacterial periplasm or cytoplasm is the pivotal step of antibacterial activity. Ceftazidime (CAZ) is a key molecule of the combination therapy for treating resistant bacteria. We designed and synthesized different fluorescent CAZ derivatives (CAZ*, CAZ**) to dissect the early step of translocation-accumulation across bacterial membrane. Their activities were determined on E. coli strains and on selected clinical isolates overexpressing ß-lactamases. The accumulation of CAZ* and CAZ** were determined by microspectrofluorimetry and epifluorimetry. The derivatives were properly translocated to the periplasmic space when we permeabilize the outer membrane barrier. The periplasmic location of CAZ** was related to a significant antibacterial activity and with the outer membrane permeability. This study demonstrated the correlation between periplasmic accumulation and antibiotic activity. We also validated the method for approaching ß-lactam permeation relative to membrane permeability and paved the way for an original matrix for determining "Structure Intracellular Accumulation Activity Relationship" for the development of new therapeutic candidates.
Topics: Anti-Bacterial Agents; Ceftazidime; Cell Membrane; Gram-Negative Bacteria; Microbial Sensitivity Tests; Microspectrophotometry; Molecular Structure; Periplasm; Permeability
PubMed: 28428543
DOI: 10.1038/s41598-017-00945-8 -
Interface Focus Feb 2019The blue neck and breast feathers of the peacock are structurally coloured due to an intricate photonic crystal structure in the barbules consisting of a...
The blue neck and breast feathers of the peacock are structurally coloured due to an intricate photonic crystal structure in the barbules consisting of a two-dimensionally ordered rectangular lattice of melanosomes (melanin rodlets) and air channels embedded in a keratin matrix. We here investigate the feather coloration by performing microspectrophotometry, imaging scatterometry and angle-dependent reflectance measurements. Using previously determined wavelength-dependent refractive indices of melanin and keratin, we interpret the spectral and spatial reflection characteristics by comparing the measured spectra to calculated spectra by effective-medium multilayer and full three-dimensional finite-difference time-domain modelling. Both modelling methods yield similar reflectance spectra indicating that simple multilayer modelling is adequate for a direct understanding of the brilliant coloration of peacock feathers.
PubMed: 30603065
DOI: 10.1098/rsfs.2018.0043 -
Proceedings. Biological Sciences Sep 2016Previous investigations of vision and visual pigment evolution in aquatic predators have focused on fish and crustaceans, generally ignoring the cephalopods. Since the... (Comparative Study)
Comparative Study
Previous investigations of vision and visual pigment evolution in aquatic predators have focused on fish and crustaceans, generally ignoring the cephalopods. Since the first cephalopod opsin was sequenced in late 1980s, we now have data on over 50 cephalopod opsins, prompting this functional and phylogenetic examination. Much of this data does not specifically examine the visual pigment spectral absorbance position (λmax) relative to environment or lifestyle, and cephalopod opsin functional adaptation and visual ecology remain largely unknown. Here we introduce a new protocol for photoreceptor microspectrophotometry (MSP) that overcomes the difficulty of bleaching the bistable visual pigment and that reveals eight coastal coleoid cephalopods to be monochromatic with λmax varying from 484 to 505 nm. A combination of current MSP results, the λmax values previously characterized using cephalopod retinal extracts (467-500 nm) and the corresponding opsin phylogenetic tree were used for systematic comparisons with an end goal of examining the adaptations of coleoid visual pigments to different light environments. Spectral tuning shifts are described in response to different modes of life and light conditions. A new spectral tuning model suggests that nine amino acid substitution sites may determine the direction and the magnitude of spectral shifts.
Topics: Animals; Cephalopoda; Ecosystem; Phylogeny; Retinal Pigments; Rod Opsins
PubMed: 27629028
DOI: 10.1098/rspb.2016.1346 -
Journal of Evolutionary Biology Jul 2015The dominant hypothesis for the evolutionary origin of snakes from 'lizards' (non-snake squamates) is that stem snakes acquired many snake features while passing through...
The dominant hypothesis for the evolutionary origin of snakes from 'lizards' (non-snake squamates) is that stem snakes acquired many snake features while passing through a profound burrowing (fossorial) phase. To investigate this, we examined the visual pigments and their encoding opsin genes in a range of squamate reptiles, focusing on fossorial lizards and snakes. We sequenced opsin transcripts isolated from retinal cDNA and used microspectrophotometry to measure directly the spectral absorbance of the photoreceptor visual pigments in a subset of samples. In snakes, but not lizards, dedicated fossoriality (as in Scolecophidia and the alethinophidian Anilius scytale) corresponds with loss of all visual opsins other than RH1 (λmax 490-497 nm); all other snakes (including less dedicated burrowers) also have functional sws1 and lws opsin genes. In contrast, the retinas of all lizards sampled, even highly fossorial amphisbaenians with reduced eyes, express functional lws, sws1, sws2 and rh1 genes, and most also express rh2 (i.e. they express all five of the visual opsin genes present in the ancestral vertebrate). Our evidence of visual pigment complements suggests that the visual system of stem snakes was partly reduced, with two (RH2 and SWS2) of the ancestral vertebrate visual pigments being eliminated, but that this did not extend to the extreme additional loss of SWS1 and LWS that subsequently occurred (probably independently) in highly fossorial extant scolecophidians and A. scytale. We therefore consider it unlikely that the ancestral snake was as fossorial as extant scolecophidians, whether or not the latter are para- or monophyletic.
Topics: Animals; Biological Evolution; Evolution, Molecular; Lizards; Molecular Sequence Data; Opsins; Phylogeny; Retina; Snakes
PubMed: 26012745
DOI: 10.1111/jeb.12663 -
The Journal of Experimental Biology Apr 2024In many animals, ultraviolet (UV) vision guides navigation, foraging, and communication, but few studies have addressed the contribution of UV signals to colour vision,...
In many animals, ultraviolet (UV) vision guides navigation, foraging, and communication, but few studies have addressed the contribution of UV signals to colour vision, or measured UV discrimination thresholds using behavioural experiments. Here, we tested UV colour vision in an anemonefish (Amphiprion ocellaris) using a five-channel (RGB-V-UV) LED display. We first determined that the maximal sensitivity of the A. ocellaris UV cone was ∼386 nm using microspectrophotometry. Three additional cone spectral sensitivities had maxima at ∼497, 515 and ∼535 nm. We then behaviourally measured colour discrimination thresholds by training anemonefish to distinguish a coloured target pixel from grey distractor pixels of varying intensity. Thresholds were calculated for nine sets of colours with and without UV signals. Using a tetrachromatic vision model, we found that anemonefish were better (i.e. discrimination thresholds were lower) at discriminating colours when target pixels had higher UV chromatic contrast. These colours caused a greater stimulation of the UV cone relative to other cone types. These findings imply that a UV component of colour signals and cues improves their detectability, which likely increases the prominence of anemonefish body patterns for communication and the silhouette of zooplankton prey.
Topics: Animals; Color; Color Vision; Retinal Cone Photoreceptor Cells; Color Perception; Perciformes; Ultraviolet Rays
PubMed: 38586934
DOI: 10.1242/jeb.247425 -
Proceedings. Biological Sciences May 2016The coloration of flowers is due to the wavelength-selective absorption by pigments of light backscattered by structures inside the petals. We investigated the optical...
The coloration of flowers is due to the wavelength-selective absorption by pigments of light backscattered by structures inside the petals. We investigated the optical properties of flowers using (micro)spectrophotometry and anatomical methods. To assess the contribution of different structures to the overall visual signal of flowers, we used an optical model, where a petal is considered as a stack of differently pigmented and structured layers and we interpreted the visual signals of the model petals with insect vision models. We show that the reflectance depends, in addition to the pigmentation, on the petal's thickness and the inhomogeneity of its interior. We find large between-species differences in floral pigments, pigment concentration and localization, as well as floral interior structure. The fractions of reflected and transmitted light are remarkably similar between the studied species, suggesting common selective pressures of pollinator visual systems. Our optical model highlights that pigment localization crucially determines the efficiency of pigmentary filtering and thereby the chromatic contrast and saturation of the visual signal. The strongest visual signal occurs with deposition of pigments only on the side of viewing. Our systematic approach and optical modelling open new perspectives on the virtues of flower colour.
Topics: Animals; Bees; Flowers; Models, Biological; Photoreceptor Cells, Invertebrate; Pigmentation; Pigments, Biological; Pollination; Spectrophotometry
PubMed: 27170723
DOI: 10.1098/rspb.2016.0429 -
Scientific Reports Mar 2016Monitoring the effect of the substrate on the local surface plasmon resonance (LSPR) of metallic nanoparticles is key for deepening our understanding of light-matter...
Monitoring the effect of the substrate on the local surface plasmon resonance (LSPR) of metallic nanoparticles is key for deepening our understanding of light-matter interactions at the nanoscale. This coupling gives rise to shifts of the LSPR as well as changes in the scattering pattern shape. The problem requires of high-throughput techniques that present both high spatial and spectral resolution. We present here a technique, referred to as Spatially Multiplexed Micro-Spectrophotometry (SMMS), able to perform polarization-resolved spectral and spatial analysis of the scattered light over large surface areas. The SMMS technique provides three orders of magnitude faster spectroscopic analysis than conventional dark-field microspectrophotometry, with the capability for mapping the spatial distribution of the scattered light intensity with lateral resolution of 40 nm over surface areas of 0.02 mm(2). We show polarization-resolved dark-field spectral analysis of hundreds of gold nanoparticles deposited on a silicon surface. The technique allows determining the effect of the substrate on the LSPR of single nanoparticles and dimers and their scattering patterns. This is applied for rapid discrimination and counting of monomers and dimers of nanoparticles. In addition, the diameter of individual nanoparticles can be rapidly assessed with 1 nm accuracy.
PubMed: 26953042
DOI: 10.1038/srep22836 -
PloS One 2014The howler monkeys (Alouatta sp.) are the only New World primates to exhibit routine trichromacy. Both males and females have three cone photopigments. However, in...
The howler monkeys (Alouatta sp.) are the only New World primates to exhibit routine trichromacy. Both males and females have three cone photopigments. However, in contrast to Old World monkeys, Alouatta has a locus control region upstream of each opsin gene on the X-chromosome and this might influence the retinal organization underlying its color vision. Post-mortem microspectrophotometry (MSP) was performed on the retinae of two male Alouatta to obtain rod and cone spectral sensitivities. The MSP data were consistent with only a single opsin being expressed in each cone and electrophysiological data were consistent with this primate expressing full trichromacy. To study the physiological organization of the retina underlying Alouatta trichromacy, we recorded from retinal ganglion cells of the same animals used for MSP measurements with a variety of achromatic and chromatic stimulus protocols. We found MC cells and PC cells in the Alouatta retina with similar properties to those previously found in the retina of other trichromatic primates. MC cells showed strong phasic responses to luminance changes and little response to chromatic pulses. PC cells showed strong tonic response to chromatic changes and small tonic response to luminance changes. Responses to other stimulus protocols (flicker photometry; changing the relative phase of red and green modulated lights; temporal modulation transfer functions) were also similar to those recorded in other trichromatic primates. MC cells also showed a pronounced frequency double response to chromatic modulation, and with luminance modulation response saturation accompanied by a phase advance between 10-20 Hz, characteristic of a contrast gain mechanism. This indicates a very similar retinal organization to Old-World monkeys. Cone-specific opsin expression in the presence of a locus control region for each opsin may call into question the hypothesis that this region exclusively controls opsin expression.
Topics: Alouatta; Animals; Color; Color Perception; Color Vision; Electrophysiology; Female; Light; Male; Microspectrophotometry; Neurons; Opsins; Retinal Cone Photoreceptor Cells; Retinal Ganglion Cells; Retinal Pigments; Vision, Ocular
PubMed: 25405863
DOI: 10.1371/journal.pone.0113321 -
The Journal of Neuroscience : the... Jun 2016Rhodopsin is a prototypical G-protein-coupled receptor (GPCR) that is activated when its 11-cis-retinal moiety is photoisomerized to all-trans retinal. This step...
UNLABELLED
Rhodopsin is a prototypical G-protein-coupled receptor (GPCR) that is activated when its 11-cis-retinal moiety is photoisomerized to all-trans retinal. This step initiates a cascade of reactions by which rods signal changes in light intensity. Like other GPCRs, rhodopsin is deactivated through receptor phosphorylation and arrestin binding. Full recovery of receptor sensitivity is then achieved when rhodopsin is regenerated through a series of steps that return the receptor to its ground state. Here, we show that dephosphorylation of the opsin moiety of rhodopsin is an extremely slow but requisite step in the restoration of the visual pigment to its ground state. We make use of a novel observation: isolated mouse retinae kept in standard media for routine physiologic recordings display blunted dephosphorylation of rhodopsin. Isoelectric focusing followed by Western blot analysis of bleached isolated retinae showed little dephosphorylation of rhodopsin for up to 4 h in darkness, even under conditions when rhodopsin was completely regenerated. Microspectrophotometeric determinations of rhodopsin spectra show that regenerated phospho-rhodopsin has the same molecular photosensitivity as unphosphorylated rhodopsin and that flash responses measured by trans-retinal electroretinogram or single-cell suction electrode recording displayed dark-adapted kinetics. Single quantal responses displayed normal dark-adapted kinetics, but rods were only half as sensitive as those containing exclusively unphosphorylated rhodopsin. We propose a model in which light-exposed retinae contain a mixed population of phosphorylated and unphosphorylated rhodopsin. Moreover, complete dark adaptation can only occur when all rhodopsin has been dephosphorylated, a process that requires >3 h in complete darkness.
SIGNIFICANCE STATEMENT
G-protein-coupled receptors (GPCRs) constitute the largest superfamily of proteins that compose ∼4% of the mammalian genome whose members share a common membrane topology. Signaling by GPCRs regulate a wide variety of physiological processes, including taste, smell, hearing, vision, and cardiovascular, endocrine, and reproductive homeostasis. An important feature of GPCR signaling is its timely termination. This normally occurs when, after their activation, GPCRs are rapidly phosphorylated by specific receptor kinases and subsequently bound by cognate arrestins. Recovery of receptor sensitivity to the ground state then requires dephosphorylation of the receptor and unbinding of arrestin, processes that are poorly understood. Here we investigate in mouse rod photoreceptors the relationship between rhodopsin dephosphorylation and recovery of visual sensitivity.
Topics: Animals; Biophysics; Dark Adaptation; Electroretinography; G-Protein-Coupled Receptor Kinase 1; Heterotrimeric GTP-Binding Proteins; In Vitro Techniques; Isoelectric Focusing; Light; Mice; Mice, Inbred C57BL; Mice, Transgenic; Microspectrophotometry; Mutation; Opsins; Phosphorylation; Retina; Retinal Rod Photoreceptor Cells; Retinaldehyde; Rhodopsin
PubMed: 27358455
DOI: 10.1523/JNEUROSCI.3544-15.2016