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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 Jul 2023Dystrophic muscle is characterized by necrosis/regeneration cycles, inflammation, and fibro-adipogenic development. Conventional histological stainings provide essential...
Dystrophic muscle is characterized by necrosis/regeneration cycles, inflammation, and fibro-adipogenic development. Conventional histological stainings provide essential topographical data of this remodeling but may be limited to discriminate closely related pathophysiological contexts. They fail to mention microarchitecture changes linked to the nature and spatial distribution of tissue compartment components. We investigated whether label-free tissue autofluorescence revealed by Synchrotron deep ultraviolet (DUV) radiation could serve as an additional tool for monitoring dystrophic muscle remodeling. Using widefield microscopy with specific emission fluorescence filters and microspectroscopy defined by high spectral resolution, we analyzed samples from healthy dogs and two groups of dystrophic dogs: naïve (severely affected) and MuStem cell-transplanted (clinically stabilized) animals. Multivariate statistical analysis and machine learning approaches demonstrated that autofluorescence emitted at 420-480 nm by the Biceps femoris muscle effectively discriminates between healthy, dystrophic, and transplanted dog samples. Microspectroscopy showed that dystrophic dog muscle displays higher and lower autofluorescence due to collagen cross-linking and NADH respectively than that of healthy and transplanted dogs, defining biomarkers to evaluate the impact of cell transplantation. Our findings demonstrate that DUV radiation is a sensitive, label-free method to assess the histopathological status of dystrophic muscle using small amounts of tissue, with potential applications in regenerative medicine.
Topics: Animals; Dogs; Random Forest; Support Vector Machine; Muscular Dystrophies; Ultraviolet Rays; Microspectrophotometry; Microscopy; Stem Cell Transplantation; Male; Biopsy
PubMed: 37402811
DOI: 10.1038/s41598-023-37762-1 -
Insects Aug 2023Quantifying the growth of entomopathogenic fungi is crucial for understanding their virulence and pathogenic potential. Traditional methods for determining growth, such...
Quantifying the growth of entomopathogenic fungi is crucial for understanding their virulence and pathogenic potential. Traditional methods for determining growth, such as biomass determination or colony growth area, are time-consuming and quantitatively and spatially limited in scope. In this study, we introduce a high-throughput method for rapidly measuring fungal growth using spectrophotometry in small-volume, liquid media cultures in 96-well microplates. Optical density (OD) changes were directly correlated with dry weight of samples for six isolates from three species of the genus to validate spectrophotometric growth measurements, and investigate species- and isolate-specific effects. We quantified fungal biomass from the microcultures by extracting, drying, and weighing mycelial mats. From the relationship established between OD and biomass, we generated standard curves for predicting biomass based on the OD values. The OD measurements clearly distinguished growth patterns among six isolates from three species. The logistic growth phase, as captured by the OD measurements, could be accurately assessed within a span of 80 h. Using isolates of , , and , this technique was demonstrated to be an effective, reproducible, and simple method for rapidly measuring filamentous fungal growth with high precision. This technique offers a valuable tool for studying the growth dynamics of entomopathogenic fungi and investigating the factors that influence their growth.
PubMed: 37623413
DOI: 10.3390/insects14080703 -
Journal of the American Chemical Society Jul 2023Single-molecule localization microscopy (SMLM) at cryogenic temperature opens new avenues to investigate intact biological samples at the nanoscale and perform...
Single-molecule localization microscopy (SMLM) at cryogenic temperature opens new avenues to investigate intact biological samples at the nanoscale and perform cryo-correlative studies. Genetically encoded fluorescent proteins (FPs) are markers of choice for cryo-SMLM, but their reduced conformational flexibility below the glass-transition temperature hampers efficient cryo-photoswitching. We investigated cryo-switching of rsEGFP2, one of the most efficient reversibly switchable fluorescent proteins at ambient temperature due to facile isomerization of the chromophore. UV-visible microspectrophotometry and X-ray crystallography revealed a completely different switching mechanism at ∼110 K. At this cryogenic temperature, on-off photoswitching involves the formation of two off-states in conformation with blue-shifted absorption relative to that of the protonated chromophore populated at ambient temperature. Only one of these off-states can be switched back to the fluorescent on-state by 405 nm light, while both of them are sensitive to UV light at 355 nm. Superior recovery to the fluorescent on-state by 355 nm light was confirmed at the single-molecule level. This suggests, as also shown by simulations, that employing 355 nm light in cryo-SMLM experiments using rsEGFP2 and possibly other FPs could improve the effective labeling efficiency achievable with this technique. The rsEGFP2 photoswitching mechanism discovered in this work adds to the panoply of known switching mechanisms in fluorescent proteins.
Topics: Temperature; Luminescent Proteins; Isomerism; Protein Conformation; Ultraviolet Rays
PubMed: 37389576
DOI: 10.1021/jacs.3c01500 -
Small (Weinheim An Der Bergstrasse,... Feb 2024Thermochromic materials have been widely investigated due to their relevance in technological applications, including anti-counterfeiting materials, fashion accessories,...
Thermochromic materials have been widely investigated due to their relevance in technological applications, including anti-counterfeiting materials, fashion accessories, displays, and temperature sensors. While many organisms exhibit color changes, few studies have explored the potential of the responsive natural materials for temperature sensing, especially given the often limited and irreversible nature of these changes in live specimens. Here, it is shown that the hindwings of the blue-winged grasshopper Coloracris azureus can act as a reversible, power-free bio-thermometer, transitioning from blue to purple/red in a 30-100°C temperature range. Using microspectrophotometry, light microscopy and Raman microscopy, it is found that the blue color of the wings originates from pigmentary coloration, based on a complex of astaxanthin and proteins. The thermochromic shift from blue to red, induced by a temperature increase, is attributed to a denaturation of this carotenoprotein complex, upon which astaxanthin is released. This process is reversible upon a subsequent temperature decrease. The color changes are both swift and consistent upon temperature change, making the grasshopper's wings suitable as direct visual sensors on thermally dynamic, curved surfaces. The potential possibilities of sustainable, power-free temperature sensors or microthermometers based on biomaterials are demonstrated.
PubMed: 38366281
DOI: 10.1002/smll.202310193 -
Forensic Science International Jan 2021Soil is a very important type of trace evidence. The iron content of soil is of great significance in distinguishing soil types, discriminating among different soils,...
Soil is a very important type of trace evidence. The iron content of soil is of great significance in distinguishing soil types, discriminating among different soils, and tracing soils. However, conventional methods for analyzing the iron content of soil are expensive, laborious, and time-consuming. Previous studies have shown that the color of soil correlates well with its hematite content. This article thus deals with the indirect determination of iron content using soil color as a proxy. Soil color measurements were conducted using microspectrophotometry (MSP), and resulting data were transformed into chromaticity value (L*, a*, and b*). Predictions using the redness index in conjunction with a linear regression model were compared with those using the chromaticity value and a back propagation neural network (BPNN) model. The influences of different modeling conditions on the modeling accuracy were compared, and more accurate predictions were achieved when the iron content was higher than 2.13%. The BPNN model produced predictions with R and RMSE values of 0.955 and 0.336%, which were better than the predictions of the linear regression model (R: 0.859, RMSE, 1.07%). We thus demonstrated that MSP can be used for fast, accurate, and non-destructive measurements of soil color and prediction of its iron content. Although the results may not be as precise as conventional laboratory analysis, they still provide more information with acceptable accuracy, which should have promising applications in forensic applications.
PubMed: 33278697
DOI: 10.1016/j.forsciint.2020.110600 -
Journal of Fish Biology Sep 2022The main goal of this study was to clarify whether the spectral properties of retinal photoreceptors reflect the features of behaviour of closely related fish species...
The main goal of this study was to clarify whether the spectral properties of retinal photoreceptors reflect the features of behaviour of closely related fish species cohabiting shallow marine and fresh waters. The spectral sensitivity of photoreceptors was compared between two smelt species, Hypomesus japonicus and Japanese smelt Hypomesus nipponensis. The spectral absorption of the visual pigments was measured using microspectrophotometry. In H. japonicus, a mostly marine species, all photoreceptors contained visual pigments based on retinal and were distributed differently in specific retinal areas. The absorbance maxima (λ ) of rods and long-wave-sensitive members of double cones throughout the retina amounted to 507 and 573 nm, respectively, but the λ value of the short-wave-sensitive members of double cones and single cones in the temporal hemiretina showed a significant blue shift compared to the nasal hemiretina: 485 vs. 516 nm and 375 vs. 412 nm, respectively, thus enhancing the short-wave sensitivity of the temporal hemiretina. In H. nipponensis, an euryhaline species, the estimated λ value of both rods and cones significantly varied between the groups caught in different localities (sea, river or estuary) because of the presence of rhodopsin/porphyropsin mixtures. The long-wavelength shift in rod and cone photoreceptors was observed because of changes in the chromophore complement in closely related but ecologically different species dwelling in freshened bodies of water. Considering the data available in the literature, several putative common opsin genes have been suggested for species under study.
Topics: Animals; Opsins; Osmeriformes; Photoreceptor Cells, Vertebrate; Retinal Cone Photoreceptor Cells; Rhodopsin
PubMed: 35655413
DOI: 10.1111/jfb.15128 -
Molecular Biology and Evolution Apr 2024Visual systems adapt to different light environments through several avenues including optical changes to the eye and neurological changes in how light signals are...
Visual systems adapt to different light environments through several avenues including optical changes to the eye and neurological changes in how light signals are processed and interpreted. Spectral sensitivity can evolve via changes to visual pigments housed in the retinal photoreceptors through gene duplication and loss, differential and coexpression, and sequence evolution. Frogs provide an excellent, yet understudied, system for visual evolution research due to their diversity of ecologies (including biphasic aquatic-terrestrial life cycles) that we hypothesize imposed different selective pressures leading to adaptive evolution of the visual system, notably the opsins that encode the protein component of the visual pigments responsible for the first step in visual perception. Here, we analyze the diversity and evolution of visual opsin genes from 93 new eye transcriptomes plus published data for a combined dataset spanning 122 frog species and 34 families. We find that most species express the four visual opsins previously identified in frogs but show evidence for gene loss in two lineages. Further, we present evidence of positive selection in three opsins and shifts in selective pressures associated with differences in habitat and life history, but not activity pattern. We identify substantial novel variation in the visual opsins and, using microspectrophotometry, find highly variable spectral sensitivities, expanding known ranges for all frog visual pigments. Mutations at spectral-tuning sites only partially account for this variation, suggesting that frogs have used tuning pathways that are unique among vertebrates. These results support the hypothesis of adaptive evolution in photoreceptor physiology across the frog tree of life in response to varying environmental and ecological factors and further our growing understanding of vertebrate visual evolution.
Topics: Humans; Animals; Opsins; Retinal Pigments; Anura; Gene Duplication; Microspectrophotometry
PubMed: 38573520
DOI: 10.1093/molbev/msae049 -
Zoological Letters Nov 2020The dorsal wings of male Sasakia charonda butterflies display a striking blue iridescent coloration, which is accentuated by white, orange-yellow and red spots, as well...
The dorsal wings of male Sasakia charonda butterflies display a striking blue iridescent coloration, which is accentuated by white, orange-yellow and red spots, as well as by brown margins. The ventral wings also have a variegated, but more subdued, pattern. We investigated the optical basis of the various colors of intact wings as well as isolated wing scales by applying light and electron microscopy, imaging scatterometry and (micro)spectrophotometry. The prominent blue iridescence is due to scales with tightly packed, multilayered ridges that contain melanin pigment. The scales in the brown wing margins also contain melanin. Pigments extracted from the orange-yellow and red spots indicate the presence of 3-OH-kynurenine and ommochrome pigment. The scales in the white spots also have multilayered ridges but lack pigment. The lower lamina of the scales plays a so-far undervalued but often crucial role. Its thin-film properties color the majority of the ventral wing scales, which are unpigmented and have large windows. The lower lamina acting as a thin-film reflector generally contributes to the reflectance of the various scale types.
PubMed: 33292721
DOI: 10.1186/s40851-020-00164-6 -
Comparative Biochemistry and... Dec 2019Changes in visual pigments were studied in two marine fish species, the masked greenling Hexagrammos octogrammus and the prickleback Pholidapus dybowskii. A...
Changes in visual pigments were studied in two marine fish species, the masked greenling Hexagrammos octogrammus and the prickleback Pholidapus dybowskii. A microspectrophotometric (MSP) analysis showed that the rods and cones of the fish collected from the natural marine environment in summer or kept in a tank at a high illumination level predominantly contained porphyropsins based on chromophore A2. As a result, λ of the double cones significantly shifted to longer wavelengths, reaching 625 and 609 nm, respectively. After several weeks of dark adaptation, the spectra of all the photoreceptor types shifted to shorter wavelengths, as the A1:A2 ratio switched to A1. The MSP data from the fish kept under controlled light conditions were confirmed by chromatography (HPLC), which showed that the changes in the chromophore ratio were reversible and independent of the water temperature. After the preliminary deep dark adaptation, the first noticeable shift in the pigment ratio from A1 to A2 occurred within two weeks of exposure to bright light. A novel finding in this study was a reverse polarity of A1/A2 changes, unlike the case in most other fish species, where A2 chromophore predominated after the dark exposure. This demonstration of the unusual phenomenon of visual pigment transformation suggests a modification or a new way for the activation of specific biochemical mechanisms of A1:A2 conversion at both high and low illumination levels.
Topics: Animals; Aquatic Organisms; Dark Adaptation; Fishes; Light; Microspectrophotometry; Retinal Cone Photoreceptor Cells; Retinal Pigments; Retinal Rod Photoreceptor Cells; Seasons; Temperature
PubMed: 31499167
DOI: 10.1016/j.cbpa.2019.110560