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International Journal of Molecular... Jun 2023Though microscopy is most often intended as a technique for providing qualitative assessment of cellular and subcellular properties, when coupled with other instruments... (Review)
Review
Though microscopy is most often intended as a technique for providing qualitative assessment of cellular and subcellular properties, when coupled with other instruments such as wavelength selectors, lasers, photoelectric devices and computers, it can perform a wide variety of quantitative measurements, which are demanding in establishing relationships between the properties and structures of biological material in all their spatial and temporal complexities. These combinations of instruments are a powerful approach to improve non-destructive investigations of cellular and subcellular properties (both physical and chemical) at a macromolecular scale resolution. Since many subcellular compartments in living cells are characterized by structurally organized molecules, this review deals with three advanced microscopy techniques well-suited for these kind of investigations, i.e., microspectrophotometry (MSP), super-resolution localization microscopy (SRLM) and holotomographic microscopy (HTM). These techniques can achieve an insight view into the role intracellular molecular organizations such as photoreceptive and photosynthetic structures and lipid bodies play in many cellular processes as well as their biophysical properties. Microspectrophotometry uses a set-up based on the combination of a wide-field microscope and a polychromator, which allows the measurement of spectroscopic features such as absorption spectra. Super resolution localization microscopy combines dedicated optics and sophisticated software algorithms to overcome the diffraction limit of light and allow the visualization of subcellular structures and dynamics in greater detail with respect to conventional optical microscopy. Holotomographic microscopy combines holography and tomography techniques into a single microscopy set-up, and allows 3D reconstruction by means of the phase separation of biomolecule condensates. This review is organized in sections, which for each technique describe some general aspects, a peculiar theoretical aspect, a specific experimental configuration and examples of applications (fish and algae photoreceptors, single labeled proteins and endocellular aggregates of lipids).
Topics: Animals; Microscopy, Fluorescence; Proteins; Optics and Photonics; Biophysics; Holography
PubMed: 37373120
DOI: 10.3390/ijms24129973 -
International Journal of Molecular... Apr 2023The review briefly describes various types of infrared (IR) and Raman spectroscopy methods. At the beginning of the review, the basic concepts of biological methods of... (Review)
Review
The review briefly describes various types of infrared (IR) and Raman spectroscopy methods. At the beginning of the review, the basic concepts of biological methods of environmental monitoring, namely bioanalytical and biomonitoring methods, are briefly considered. The main part of the review describes the basic principles and concepts of vibration spectroscopy and microspectrophotometry, in particular IR spectroscopy, mid- and near-IR spectroscopy, IR microspectroscopy, Raman spectroscopy, resonance Raman spectroscopy, Surface-enhanced Raman spectroscopy, and Raman microscopy. Examples of the use of various methods of vibration spectroscopy for the study of biological samples, especially in the context of environmental monitoring, are given. Based on the described results, the authors conclude that the near-IR spectroscopy-based methods are the most convenient for environmental studies, and the relevance of the use of IR and Raman spectroscopy in environmental monitoring will increase with time.
Topics: Vibration; Biological Monitoring; Spectrophotometry, Infrared; Spectrum Analysis, Raman; Spectroscopy, Near-Infrared; Spectroscopy, Fourier Transform Infrared
PubMed: 37108111
DOI: 10.3390/ijms24086947 -
Handbook of Experimental Pharmacology 2021Neuromodulators are critical regulators of the brain's signaling processes, and thus they are popular pharmacological targets for psychoactive therapies. It is clear... (Review)
Review
Neuromodulators are critical regulators of the brain's signaling processes, and thus they are popular pharmacological targets for psychoactive therapies. It is clear that monoamine uptake mechanisms are complicated and subject to multiple uptake mechanisms. Uptake 1 describes uptake of the monoamine via its designated transporter (SERT for serotonin, NET for norepinephrine, and DAT for dopamine), whereas Uptake 2 details multiple transporter types on neurons and glia taking up different types of modulators, not necessarily specific to the monoamine. While Uptake 1 processes have been well-studied over the past few decades, Uptake 2 mechanisms have remained more difficult to study because of the limitations in methods that have the sensitivity and spatiotemporal resolution to look at the subtleties in uptake profiles. In this chapter we review the different experimental approaches that have yielded important information about Uptake 2 mechanisms in vivo. The techniques (scintillation microspectrophotometry, microdialysis, chronoamperometry, and voltammetry) are described in detail, and pivotal studies associated with each method are highlighted. It is clear from these reviewed works that Uptake 2 processes are critical to consider to advance our understanding of the brain and develop effective neuropsychiatric therapies.
Topics: Biological Transport; Dopamine; Dopamine Plasma Membrane Transport Proteins; Humans; Norepinephrine Plasma Membrane Transport Proteins; Serotonin Plasma Membrane Transport Proteins
PubMed: 34196807
DOI: 10.1007/164_2021_452 -
Theriogenology Jul 2020In the last years, an increasing interest has emerged on the development of new non-invasive methods for the assessment of oocyte quality in order to improve outcomes of... (Review)
Review
In the last years, an increasing interest has emerged on the development of new non-invasive methods for the assessment of oocyte quality in order to improve outcomes of assisted reproductive technologies (ARTs) either in medical or veterinary fields. Raman microspectroscopy (RMS) has been proposed as a promising tool for the examination of the mammalian female gamete and identification of markers of its developmental competence. This technique provides a unique spectral fingerprint indicative of molecular composition of the cell and allows probing subcellular compartments. Studies have been carried out analysing by RMS fixed or living oocytes derived from different animal models. RMS imaging has been successfully applied to discriminate the biochemical changes of the global molecular architecture of mouse oocytes at different stages of maturation and those occurring in different conditions of maturation and oocyte aging. RMS can also detect modifications of specific structural components, including the oocyte zona pellucida and F-actin subcortical cytoskeleton in fresh sheep oocytes and those underwent to vitrification procedures. Finally, the recent application of Coherent anti-Stokes Raman scattering (CARS) microscopy for examination of oocyte lipid component will be briefly discussed. CARS overcomes some limits of RMS providing vibrational and spectral information with higher sensitivity, spatial resolution which is ideal to study living oocytes. This review summarizes the research on RMS approaches for oocyte evaluation showing the high potential use, current limitations and new improvements.
Topics: Animals; Biomarkers; Cryopreservation; Female; Mammals; Microspectrophotometry; Ovum; Spectrum Analysis, Raman
PubMed: 32088036
DOI: 10.1016/j.theriogenology.2020.01.059 -
Faraday Discussions Oct 2020Peacock feathers feature a rich gamut of colours, created by a most sophisticated structural colouration mechanism. The feather barbules contain biophotonic structures...
Peacock feathers feature a rich gamut of colours, created by a most sophisticated structural colouration mechanism. The feather barbules contain biophotonic structures consisting of two-dimensionally-ordered lattices of cylindrical melanosomes and air channels embedded in keratin. Here, we study the reflectance characteristics of the various peacock tail feather colours by applying bifurcated-probe- and micro-spectrophotometry and imaging scatterometry. We compare the experimental results with published anatomical SEM and TEM data, using a transfer-matrix based effective-medium multilayer model that includes the number and diameter of the melanosome rodlets and air channels, the lattice spacing and the keratin cortex thickness, together with the recently determined wavelength-dependence of the refractive indices of keratin and melanin. Slight variations in the parameter values cause substantial changes in the spectral position and shape of the reflectance bands. We find that the number of layers crucially determines the number of peaks in the reflectance spectra. For a small number of melanosome layers, the reflectance band shape is particularly sensitive to the properties of the uppermost layer, which provides a simple mechanism for tuning the feather colours.
Topics: Animals; Birds; Color; Feathers; Microscopy, Electron, Scanning; Microscopy, Electron, Transmission; Optics and Photonics; Spectrophotometry; Tail
PubMed: 32720960
DOI: 10.1039/d0fd00033g -
The Journal of Experimental Biology Nov 2019Snapping shrimp engage in heterospecific behavioral associations in which their partners, such as goby fish, help them avoid predators. It has been argued that snapping...
Snapping shrimp engage in heterospecific behavioral associations in which their partners, such as goby fish, help them avoid predators. It has been argued that snapping shrimp engage in these partnerships because their vision is impaired by their orbital hood, an extension of their carapace that covers their eyes. To examine this idea, we assessed the visual abilities of snapping shrimp. We found the big claw snapping shrimp, , has spatial vision provided by compound eyes with reflecting superposition optics. These eyes view the world through an orbital hood that is 80-90% as transparent as seawater across visible wavelengths (400-700 nm). Through electroretinography and microspectrophotometry, we found the eyes of have a temporal sampling rate of >40 Hz and have at least two spectral classes of photoreceptors (λ=500 and 519 nm). From the results of optomotor behavioral experiments, we estimate the eyes of provide spatial vision with an angular resolution of ∼8 deg. We conclude that snapping shrimp have competent visual systems, suggesting the function and evolution of their behavioral associations should be re-assessed and that these animals may communicate visually with conspecifics and heterospecific partners.
Topics: Animals; Decapoda; Electroretinography; Female; Male; Microspectrophotometry; Vision, Ocular; Visual Perception
PubMed: 31624099
DOI: 10.1242/jeb.209015 -
Molecular Biology and Evolution Mar 2020The diversity of color vision systems found in extant vertebrates suggests that different evolutionary selection pressures have driven specializations in photoreceptor...
The diversity of color vision systems found in extant vertebrates suggests that different evolutionary selection pressures have driven specializations in photoreceptor complement and visual pigment spectral tuning appropriate for an animal's behavior, habitat, and life history. Aquatic vertebrates in particular show high variability in chromatic vision and have become important models for understanding the role of color vision in prey detection, predator avoidance, and social interactions. In this study, we examined the capacity for chromatic vision in elasmobranch fishes, a group that have received relatively little attention to date. We used microspectrophotometry to measure the spectral absorbance of the visual pigments in the outer segments of individual photoreceptors from several ray and shark species, and we sequenced the opsin mRNAs obtained from the retinas of the same species, as well as from additional elasmobranch species. We reveal the phylogenetically widespread occurrence of dichromatic color vision in rays based on two cone opsins, RH2 and LWS. We also confirm that all shark species studied to date appear to be cone monochromats but report that in different species the single cone opsin may be of either the LWS or the RH2 class. From this, we infer that cone monochromacy in sharks has evolved independently on multiple occasions. Together with earlier discoveries in secondarily aquatic marine mammals, this suggests that cone-based color vision may be of little use for large marine predators, such as sharks, pinnipeds, and cetaceans.
Topics: Animals; Color Vision; Fish Proteins; Gene Expression Profiling; Microspectrophotometry; Opsins; Phylogeny; Retina; Retinal Cone Photoreceptor Cells; Sequence Analysis, RNA; Sharks; Skates, Fish
PubMed: 31770430
DOI: 10.1093/molbev/msz269 -
Langmuir : the ACS Journal of Surfaces... Nov 2023Aspirin has been used for broad therapeutic treatment, including secondary prevention of cardiovascular disease associated with increased cholesterol levels. Aspirin and...
Aspirin has been used for broad therapeutic treatment, including secondary prevention of cardiovascular disease associated with increased cholesterol levels. Aspirin and other nonsteroidal anti-inflammatory drugs have been shown to interact with lipid membranes and change their biophysical properties. In this study, mixed lipid model bilayers made from 1-palmitoyl-2-oleoyl--glycero-3-phosphatidylcholine (POPC) or 1,2-dioleoyl--glycero-3-phosphatidylcholine (DOPC) comprising varying concentrations of cholesterol (10:1, 4:1, and 1:1 mole ratio of lipid:chol), prepared by the droplet interface bilayer method, were used to examine the effects of aspirin at various pH on transbilayer water permeability. The presence of aspirin increases the water permeability of POPC bilayers in a concentration-dependent manner, with a greater magnitude of increase at pH 3 compared to pH 7. In the presence of cholesterol, aspirin is similarly shown to increase water permeability; however, the extent of the increase depends on both the concentration of cholesterol and the pH, with the least pronounced enhancement in water permeability at high cholesterol levels at pH 7. A fusion of data from differential scanning calorimetry, confocal Raman microspectrophotometry, and interfacial tensiometric measurements demonstrates that aspirin can promote significant thermal, structural, and interfacial property perturbations in the mixed-lipid POPC or DOPC membranes containing cholesterol, indicating a disordering effect on the lipid membranes. Our findings suggest that aspirin fluidizes phosphocholine membranes in both cholesterol-free and cholesterol-enriched states and that the overall effect is greater when aspirin is in a neutral state. These results confer a deeper comprehension of the divergent effects of aspirin on biological membranes having heterogeneous compositions, under varying physiological pH and different cholesterol compositions, with implications for a better understanding of the gastrointestinal toxicity induced by the long term use of this important nonsteroidal anti-inflammatory molecule.
Topics: Aspirin; Phosphatidylcholines; Cholesterol; Lipid Bilayers; Water; Anti-Inflammatory Agents; Hydrogen-Ion Concentration
PubMed: 37939382
DOI: 10.1021/acs.langmuir.3c02242 -
Journal of the Royal Society, Interface May 2023Optical transparency is rare in terrestrial organisms, and often originates through loss of pigmentation and reduction in scattering. The coloured wings of some...
Optical transparency is rare in terrestrial organisms, and often originates through loss of pigmentation and reduction in scattering. The coloured wings of some butterflies and moths have repeatedly evolved transparency, offering examples of how they function optically and biologically. Because pigments are primarily localized in the scales that cover a colourless wing membrane, transparency has often evolved through the complete loss of scales or radical modification of their shape. Whereas bristle-like scales have been well documented in glasswing butterflies, other scale modifications resulting in transparency remain understudied. The butterfly achieves transparency while retaining its scales and exhibiting blue/cyan transparent zones. Here, we investigate the mechanism of wing transparency in by light microscopy, focused ion beam milling, microspectrophotometry and optical modelling. We show that transparency is achieved via loss of pigments and vertical orientation in normal paddle-like scales. These alterations are combined with an anti-reflective nipple array on portions of the wing membrane being more exposed to light. The blueish coloration of the transparent regions is due to the properties of the wing membrane, and local scale nanostructures. We show that scale retention in the transparent patches might be explained by these perpendicular scales having hydrophobic properties.
Topics: Animals; Butterflies; Wings, Animal; Pigmentation; Microscopy, Electron, Scanning; Vision, Ocular
PubMed: 37254701
DOI: 10.1098/rsif.2023.0135 -
The New Phytologist Mar 2021Hydrangea sepals exhibit a wide range of colors, from red, through purple, to blue; the purple color is a color mosaic. However, all of these colors are derived from the...
Hydrangea sepals exhibit a wide range of colors, from red, through purple, to blue; the purple color is a color mosaic. However, all of these colors are derived from the same components: simple anthocyanins, 3-O-glycosyldelphinidins, three co-pigment components, acylquinic acids and aluminum ions (Al ). We show the color mosaic is a result of graded differences in intravacuolar factors. In order to clarify the mechanisms of mosaic color, we performed single-cell analyses of vacuolar pH, and anthocyanin, co-pigment and Al content. From the sepals, a protoplast mixture of various colors was obtained. The cell color was evaluated by microspectrophotometry and vacuolar pH then was recorded by using a pH microelectrode. The organic and Al contents were quantified by micro-HPLC. We found that the bluer the cell, the greater the ratio of 5-O-acylquinic acids and Al to anthocyanins. Furthermore, reproducing experiments were conducted by mixing the components under various pH condition; all the colors could be reproduced in the various mixing conditions. Based on the above, we provide experimental evidence for cell color variation in hydrangea. Our study demonstrates the expression of phenotypic differences without any direct genomic control.
Topics: Aluminum; Anthocyanins; Color; Flowers; Hydrangea; Single-Cell Analysis
PubMed: 33220077
DOI: 10.1111/nph.17099