scholarly journals Spike Generators and Cell Signaling in the Human Auditory Nerve: An Ultrastructural, Super-Resolution, and Gene Hybridization Study

2021 ◽  
Vol 15 ◽  
Author(s):  
Wei Liu ◽  
Maria Luque ◽  
Hao Li ◽  
Anneliese Schrott-Fischer ◽  
Rudolf Glueckert ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Auditory Nerve ◽  
Spiral Ganglion ◽  
Super Resolution ◽  
Nerve Fibers ◽  
The Body ◽  
Structured Illumination ◽  
Structured Illumination Microscopy ◽  
Prosthetic Devices ◽  
Complex Sounds

Background: The human auditory nerve contains 30,000 nerve fibers (NFs) that relay complex speech information to the brain with spectacular acuity. How speech is coded and influenced by various conditions is not known. It is also uncertain whether human nerve signaling involves exclusive proteins and gene manifestations compared with that of other species. Such information is difficult to determine due to the vulnerable, “esoteric,” and encapsulated human ear surrounded by the hardest bone in the body. We collected human inner ear material for nanoscale visualization combining transmission electron microscopy (TEM), super-resolution structured illumination microscopy (SR-SIM), and RNA-scope analysis for the first time. Our aim was to gain information about the molecular instruments in human auditory nerve processing and deviations, and ways to perform electric modeling of prosthetic devices.Material and Methods: Human tissue was collected during trans-cochlear procedures to remove petro-clival meningioma after ethical permission. Cochlear neurons were processed for electron microscopy, confocal microscopy (CM), SR-SIM, and high-sensitive in situ hybridization for labeling single mRNA transcripts to detect ion channel and transporter proteins associated with nerve signal initiation and conductance.Results: Transport proteins and RNA transcripts were localized at the subcellular level. Hemi-nodal proteins were identified beneath the inner hair cells (IHCs). Voltage-gated ion channels (VGICs) were expressed in the spiral ganglion (SG) and axonal initial segments (AISs). Nodes of Ranvier (NR) expressed Nav1.6 proteins, and encoding genes critical for inter-cellular coupling were disclosed.Discussion: Our results suggest that initial spike generators are located beneath the IHCs in humans. The first NRs appear at different places. Additional spike generators and transcellular communication may boost, sharpen, and synchronize afferent signals by cell clusters at different frequency bands. These instruments may be essential for the filtering of complex sounds and may be challenged by various pathological conditions.

scholarly journals Chlamydomonas FAP70 is a component of the previously uncharacterized ciliary central apparatus projection C2a

2021 ◽  
Author(s):  
Yuqing Hou ◽  
Lei Zhao ◽  
Tomohiro Kubo ◽  
Xi Cheng ◽  
Nathan McNeill ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Mass Spectrometry ◽  
Chlamydomonas Reinhardtii ◽  
Model Organism ◽  
Super Resolution ◽  
Structured Illumination ◽  
Structured Illumination Microscopy ◽  
Cryo Electron Microscopy ◽  
N Terminus ◽  
Central Apparatus

Cilia are essential organelles required for cell signaling and motility. Nearly all motile cilia have a “9+2” axoneme composed of 9 outer doublet microtubules plus 2 central microtubules; the central microtubules together with their projections is termed the central apparatus (CA). In Chlamydomonas reinhardtii, a model organism for studying cilia, 30 proteins are known CA components, and ∼36 more are predicted to be CA proteins. Among the candidate CA proteins is the highly conserved FAP70, which also has been reported to be associated with the doublet microtubules. Here we determined by super-resolution structured illumination microscopy that FAP70 is located exclusively in the CA, and show by cryo-electron microscopy that its N-terminus is located at the base of the CA's C2a projection. We also found that fap70-1 mutant axonemes lack most of the C2a projection. Mass spectrometry revealed that fap70-1 axonemes lack not only FAP70 but two other conserved candidate CA proteins, FAP65 and FAP147. Finally, FAP65 and FAP147 co-immunoprecipitated with HA-tagged FAP70. Taken together, these data identify FAP70, FAP65, and FAP147 as the first defining components of the C2a projection.

scholarly journals Superaufgelöste Mikroskopie: Pilze unter Beobachtung

BIOspektrum ◽  
2021 ◽  
Vol 27 (4) ◽  
pp. 380-382
Author(s):  
Sebastian Sputh ◽  
Sabine Panzer ◽  
Christian Stigloher ◽  
Ulrich Terpitz
Keyword(s):  
Electron Microscopy ◽  
Membrane Protein ◽  
Fluorescence Imaging ◽  
Filamentous Fungus ◽  
Light And Electron Microscopy ◽  
Super Resolution ◽  
Diffraction Limit ◽  
Fusarium Fujikuroi ◽  
Structured Illumination ◽  
Structured Illumination Microscopy

AbstractThe diffraction limit of light confines fluorescence imaging of subcellular structures in fungi. Different super-resolution methods are available for the analysis of fungi that we briefly discuss. We exploit the filamentous fungus Fusarium fujikuroi expressing a YFP-labeled membrane protein showing the benefit of correlative light- and electron microscopy (CLEM), that combines structured illumination microscopy (SIM) and scanning election microscopy (SEM).

scholarly journals Two-dimensional TIRF-SIM–traction force microscopy (2D TIRF-SIM-TFM)

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Liliana Barbieri ◽  
Huw Colin-York ◽  
Kseniya Korobchevskaya ◽  
Di Li ◽  
Deanna L. Wolfson ◽  
...  
Keyword(s):  
Resolution Enhancement ◽  
Traction Force ◽  
Super Resolution ◽  
Traction Force Microscopy ◽  
Two Dimensional ◽  
Structured Illumination ◽  
Structured Illumination Microscopy ◽  
Force Microscopy ◽  
Health And Disease ◽  
Spatio Temporal

AbstractQuantifying small, rapidly evolving forces generated by cells is a major challenge for the understanding of biomechanics and mechanobiology in health and disease. Traction force microscopy remains one of the most broadly applied force probing technologies but typically restricts itself to slow events over seconds and micron-scale displacements. Here, we improve >2-fold spatially and >10-fold temporally the resolution of planar cellular force probing compared to its related conventional modalities by combining fast two-dimensional total internal reflection fluorescence super-resolution structured illumination microscopy and traction force microscopy. This live-cell 2D TIRF-SIM-TFM methodology offers a combination of spatio-temporal resolution enhancement relevant to forces on the nano- and sub-second scales, opening up new aspects of mechanobiology to analysis.

scholarly journals Double moiré localized plasmon structured illumination microscopy

Nanophotonics ◽  
2020 ◽  
Vol 0 (0) ◽  
Author(s):  
Ruslan Röhrich ◽  
A. Femius Koenderink
Keyword(s):  
Near Field ◽  
Super Resolution ◽  
Reconstruction Algorithm ◽  
Structured Illumination ◽  
Structured Illumination Microscopy ◽  
Experimental Realization ◽  
Spatial Frequencies ◽  
Wide Range ◽  
Localized Plasmon ◽  
Plasmonic Grating

AbstractStructured illumination microscopy (SIM) is a well-established fluorescence imaging technique, which can increase spatial resolution by up to a factor of two. This article reports on a new way to extend the capabilities of structured illumination microscopy, by combining ideas from the fields of illumination engineering and nanophotonics. In this technique, plasmonic arrays of hexagonal symmetry are illuminated by two obliquely incident beams originating from a single laser. The resulting interference between the light grating and plasmonic grating creates a wide range of spatial frequencies above the microscope passband, while still preserving the spatial frequencies of regular SIM. To systematically investigate this technique and to contrast it with regular SIM and localized plasmon SIM, we implement a rigorous simulation procedure, which simulates the near-field illumination of the plasmonic grating and uses it in the subsequent forward imaging model. The inverse problem, of obtaining a super-resolution (SR) image from multiple low-resolution images, is solved using a numerical reconstruction algorithm while the obtained resolution is quantitatively assessed. The results point at the possibility of resolution enhancements beyond regular SIM, which rapidly vanishes with the height above the grating. In an initial experimental realization, the existence of the expected spatial frequencies is shown and the performance of compatible reconstruction approaches is compared. Finally, we discuss the obstacles of experimental implementations that would need to be overcome for artifact-free SR imaging.

scholarly journals Optical Microscopy and the Extracellular Matrix Structure: A Review

Cells ◽  
2021 ◽  
Vol 10 (7) ◽  
pp. 1760
Author(s):  
Joshua J. A. Poole ◽  
Leila B. Mostaço-Guidolin
Keyword(s):  
Extracellular Matrix ◽  
Confocal Microscopy ◽  
Laser Scanning ◽  
Biological Tissues ◽  
The Body ◽  
Stimulated Emission ◽  
Substantial Part ◽  
Fluorescence Lifetime Imaging ◽  
Structured Illumination ◽  
Structured Illumination Microscopy

Biological tissues are not uniquely composed of cells. A substantial part of their volume is extracellular space, which is primarily filled by an intricate network of macromolecules constituting the extracellular matrix (ECM). The ECM serves as the scaffolding for tissues and organs throughout the body, playing an essential role in their structural and functional integrity. Understanding the intimate interaction between the cells and their structural microenvironment is central to our understanding of the factors driving the formation of normal versus remodelled tissue, including the processes involved in chronic fibrotic diseases. The visualization of the ECM is a key factor to track such changes successfully. This review is focused on presenting several optical imaging microscopy modalities used to characterize different ECM components. In this review, we describe and provide examples of applications of a vast gamut of microscopy techniques, such as widefield fluorescence, total internal reflection fluorescence, laser scanning confocal microscopy, multipoint/slit confocal microscopy, two-photon excited fluorescence (TPEF), second and third harmonic generation (SHG, THG), coherent anti-Stokes Raman scattering (CARS), fluorescence lifetime imaging microscopy (FLIM), structured illumination microscopy (SIM), stimulated emission depletion microscopy (STED), ground-state depletion microscopy (GSD), and photoactivated localization microscopy (PALM/fPALM), as well as their main advantages, limitations.

scholarly journals High-fidelity structured illumination microscopy by point-spread-function engineering

2021 ◽  
Vol 10 (1) ◽  
Author(s):  
Gang Wen ◽  
Simin Li ◽  
Linbo Wang ◽  
Xiaohu Chen ◽  
Zhenglong Sun ◽  
...  
Keyword(s):  
Point Spread Function ◽  
Super Resolution ◽  
Reconstruction Algorithm ◽  
High Fidelity ◽  
Structured Illumination ◽  
Structured Illumination Microscopy ◽  
Imaging Tool ◽  
Point Spread ◽  
Spread Function ◽  
Point Spread Function Engineering

AbstractStructured illumination microscopy (SIM) has become a widely used tool for insight into biomedical challenges due to its rapid, long-term, and super-resolution (SR) imaging. However, artifacts that often appear in SIM images have long brought into question its fidelity, and might cause misinterpretation of biological structures. We present HiFi-SIM, a high-fidelity SIM reconstruction algorithm, by engineering the effective point spread function (PSF) into an ideal form. HiFi-SIM can effectively reduce commonly seen artifacts without loss of fine structures and improve the axial sectioning for samples with strong background. In particular, HiFi-SIM is not sensitive to the commonly used PSF and reconstruction parameters; hence, it lowers the requirements for dedicated PSF calibration and complicated parameter adjustment, thus promoting SIM as a daily imaging tool.

The FDTD Analysis of Near-Field Response for Microgroove Structure With Standing Wave Illumination for the Realization of Coherent Structured Illumination Microscopy

2021 ◽  
Author(s):  
Yizhao Guan ◽  
Hiromasa Kume ◽  
Shotaro Kadoya ◽  
Masaki Michihata ◽  
Satoru Takahashi
Keyword(s):  
Standing Wave ◽  
Incident Angle ◽  
Near Field ◽  
Super Resolution ◽  
Structured Illumination ◽  
Field Phase ◽  
Structured Illumination Microscopy ◽  
Depth Measurement ◽  
Field Response ◽  
Depth Dependency

Abstract Microstructures are widely used in the manufacture of functional surfaces. An optical-based super-resolution, non-invasive method is preferred for the inspection of surfaces with massive microstructures. The Structured Illumination Microscopy (SIM) uses standing-wave illumination to reach optical super-resolution. Recently, coherent SIM is being studied. It can obtain not only the super-resolved intensity distribution but also the phase and amplitude distribution of the sample surface beyond the diffraction limit. By analysis of the phase-depth dependency, the depth measurement for microgroove structures with coherent SIM is expected. FDTD analysis is applied for observing the near-field response of microgroove under the standing-wave illumination. The near-field phase shows depth dependency in this analysis. Moreover, the effects from microgroove width, the incident angle, and the relative position between the standing-wave peak and center of the microgroove are investigated. It is found the near-field phase change can measure depth until 200 nm (aspect ratio 1) with an error of up to 20.4 nm in the case that the microgroove width is smaller than half of the wavelength.

Coding of temporal parameters of complex sounds by frog auditory nerve fibers

1991 ◽  
Vol 65 (3) ◽  
pp. 424-445 ◽  
Author(s):  
A. S. Feng ◽  
J. C. Hall ◽  
S. Siddique
Keyword(s):  
Auditory Nerve ◽  
Transfer Functions ◽  
Nerve Fibers ◽  
Spike Count ◽  
Complex Sounds ◽  
Temporal Parameters ◽  
Low Pass ◽  
The Mean ◽  
Auditory Nerve Fibers ◽  
High Pass

1. Physiological recordings were made from single auditory fibers in the frog eighth nerve to determine quantitatively how the different behaviorally relevant temporal parameters (the signal rise-fall time, duration, and rate of amplitude modulation) of complex sounds are encoded in the auditory periphery. Individual temporal parameters were varied. Response functions (RFs) were constructed with respect to each of these parameters using each unit's best excitatory frequency (BF) as the carrier. 2. In response to a change in signal rise-fall time, auditory nerve fibers showed little change in the mean spike count or firing rate, i.e., all fibers displayed ALL-PASS RFrfts. But the transient components, particularly the early phasic component, of responses varied with rise-fall times; these components were more pronounced in the responses to stimuli with shorter rise-fall times. 3. In response to an increase in signal duration, auditory nerve fibers showed a corresponding increase in firing duration and thus in the mean spike count, giving rise to HIGH-PASS RFdurs. The shape of response curves differed among fibers; the difference appeared to be related to the fiber's temporal adaptation characteristic. When the firing rate was measured, all fibers displayed higher mean firing rates in response to shorter duration stimuli than they did to longer duration stimuli, thus giving rise to LOW-PASS response functions. 4. To determine the response transfer functions to modulation rate, pulsed (PAM) and sinusoidally (SAM) amplitude-modulated signals were used. These signals differed substantially in terms of their envelopes and how they varied with AM rate. Data were analyzed by 1) plotting spike counts against the AM rate to derive modulation transfer functions (MTFspks) and 2) plotting synchronization coefficients (SCs) against the AM rate to generate MTFscs. 5. In response to PAM stimuli, all fibers showed an increase in mean spike count with modulation frequency over the range examined, giving rise to HIGH-PASS MTFspks. 6. For SAM stimuli, the average energy and duty cycle are independent of AM rate. Most (79%) auditory fibers showed little selectivity for AM rate over a range of 5-400 Hz, giving rise to ALL-PASS MTFspks. The remaining auditory fibers displayed LOW-PASS MTFspks, i.e., there was a distinct decline in the mean spike count with increasing AM rate. 7. In response to PAM stimuli, most fibers showed good response synchrony at low AM rates but the SC declined with an increase in the AM rate (i.e., LOW-PASS MTFscs). The cut-off frequency was typically very high, averaging 90 pulses/s.(ABSTRACT TRUNCATED AT 400 WORDS)

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