Sub-Millisecond Time Resolved Imaging of Voltage Changes in Excitable Cells and Tissues Using Multiple Site Optical Recording of Transmembrane Voltage (Msortv) and Molecular Probes

1997 ◽  
Vol 3 (S2) ◽  
pp. 803-804
Author(s):  
B.M. Salzberg ◽  
A.L. Obaid
Keyword(s):  
Membrane Potential ◽  
Conformational Changes ◽  
Molecular Probes ◽  
Optical Recording ◽  
Membrane Voltage ◽  
Excitable Cells ◽  
Time Resolved ◽  
Molecular Weights ◽  
Transmembrane Voltage ◽  
Time Resolved Imaging

Molecular indicators of membrane potential may be used to obtain sub-millisecond time resolved images of transient changes in membrane voltage in a variety of biological systems. These probes are small amphipathic molecules having molecular weights of 400-500, and dimensions on the order of 10 Angstroms, which bind to, but do not cross cell membranes, and change either their absorbance or fluorescence in response to membrane voltage. These extrinsic optical signals depend linearly upon membrane potential, and the best of the dyes respond to a step change in voltage in less than 1.5 μsec at room temperature. The salient properties of fast potentiometric probes will be discussed, and the fidelity of optical recordings to transmembrane voltage changes will be considered.Since voltage changes in excitable cells take place on a time scale that is determined by the kinetics of conformational changes in membrane proteins, and by membrane electrical time constants, these changes tend to be very rapid, and resolving them requires imaging systems that are frequently orders of magnitude faster than usual video rates.

scholarly journals The RCK1 domain of the human BKCa channel transduces Ca2+ binding into structural rearrangements

2010 ◽  
Vol 136 (2) ◽  
pp. 189-202 ◽  
Author(s):  
Taleh Yusifov ◽  
Anoosh D. Javaherian ◽  
Antonios Pantazis ◽  
Chris S. Gandhi ◽  
Riccardo Olcese
Keyword(s):  
Steady State ◽  
Membrane Potential ◽  
Molecular Mechanism ◽  
Conformational Changes ◽  
Bkca Channel ◽  
Structural Rearrangements ◽  
Bkca Channels ◽  
Time Resolved ◽  
High Affinity ◽  
Cytosolic Domains

Large-conductance voltage- and Ca2+-activated K+ (BKCa) channels play a fundamental role in cellular function by integrating information from their voltage and Ca2+ sensors to control membrane potential and Ca2+ homeostasis. The molecular mechanism of Ca2+-dependent regulation of BKCa channels is unknown, but likely relies on the operation of two cytosolic domains, regulator of K+ conductance (RCK)1 and RCK2. Using solution-based investigations, we demonstrate that the purified BKCa RCK1 domain adopts an α/β fold, binds Ca2+, and assembles into an octameric superstructure similar to prokaryotic RCK domains. Results from steady-state and time-resolved spectroscopy reveal Ca2+-induced conformational changes in physiologically relevant [Ca2+]. The neutralization of residues known to be involved in high-affinity Ca2+ sensing (D362 and D367) prevented Ca2+-induced structural transitions in RCK1 but did not abolish Ca2+ binding. We provide evidence that the RCK1 domain is a high-affinity Ca2+ sensor that transduces Ca2+ binding into structural rearrangements, likely representing elementary steps in the Ca2+-dependent activation of human BKCa channels.

Fluorescent potentiometric dyes for membrane-potential imaging

1994 ◽  
Vol 52 ◽  
pp. 166-167
Author(s):  
Leslie M. Loew
Keyword(s):  
Membrane Potential ◽  
Single Cells ◽  
Quantitative Imaging ◽  
Optical Recording ◽  
Biological Processes ◽  
Major Focus ◽  
The Past ◽  
Excitable Systems ◽  
Major Application ◽  
The Impact

A major application of potentiometric dyes has been the multisite optical recording of electrical activity in excitable systems. After being championed by L.B. Cohen and his colleagues for the past 20 years, the impact of this technology is rapidly being felt and is spreading to an increasing number of neuroscience laboratories. A second class of experiments involves using dyes to image membrane potential distributions in single cells by digital imaging microscopy - a major focus of this lab. These studies usually do not require the temporal resolution of multisite optical recording, being primarily focussed on slow cell biological processes, and therefore can achieve much higher spatial resolution. We have developed 2 methods for quantitative imaging of membrane potential. One method uses dual wavelength imaging of membrane-staining dyes and the other uses quantitative 3D imaging of a fluorescent lipophilic cation; the dyes used in each case were synthesized for this purpose in this laboratory.

Study of polymer chain dynamics in solution by time-resolved spectrofluorometry

1989 ◽  
Vol 54 (11) ◽  
pp. 3011-3024 ◽  
Author(s):  
Vlastimil Fidler ◽  
Stefan Vajda ◽  
Zuzana Limpouchová ◽  
Jiří Dvořák ◽  
Karel Procházka ◽  
...  
Keyword(s):  
Aqueous Solutions ◽  
Conformational Changes ◽  
Methacrylic Acid ◽  
Correlation Time ◽  
General Level ◽  
Polymeric Chain ◽  
Relative Molecular Mass ◽  
Chain Dynamics ◽  
Time Resolved ◽  
Covalently Bonded

The methodology of polarization time-resolved fluorometry and interpretation of its results are outlined at a general level, and the measurement on and use of facilities of the Edinburgh Instruments Model 299T apparatus are discussed in detail. The dynamics of conformational changes in chains of poly(methacrylic acid) containing covalently bonded dansyl labels are studied in aqueous solutions at various pH. It is shown that at pH > 6, the shorter effective rational correlation time τr < 2 ns corresponds to the rotation of the free dansyl label about bonds by which it is attached to the polymeric chain; at pH < 4 the longer effective rational correlation time τr = 20-26 ns corresponds to the rotation of the compact spherical formation constituted by a part of the collapsed polymeric chain in which the label is fixed and whose relative molecular mass is approx. 15 000-20 000.

scholarly journals Giant enhancement of THz-frequency optical nonlinearity by phonon polariton in ionic crystals

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Yao Lu ◽  
Qi Zhang ◽  
Qiang Wu ◽  
Zhigang Chen ◽  
Xueming Liu ◽  
...  
Keyword(s):  
Near Infrared ◽  
Optical Nonlinearity ◽  
Ionic Crystals ◽  
Nonlinear Optical Susceptibility ◽  
Time Resolved ◽  
Light Coupling ◽  
Tremendous Progress ◽  
Fundamental Research ◽  
Ionic Contribution ◽  
Time Resolved Imaging

AbstractThe field of nonlinear optics has grown substantially in past decades, leading to tremendous progress in fundamental research and revolutionized applications. Traditionally, the optical nonlinearity for a light wave at frequencies beyond near-infrared is observed with very high peak intensity, as in most materials only the electronic nonlinearity dominates while ionic contribution is negligible. However, it was shown that the ionic contribution to nonlinearity can be much larger than the electronic one in microwave experiments. In the terahertz (THz) regime, phonon polariton may assist to substantially trigger the ionic nonlinearity of the crystals, so as to enhance even more the nonlinear optical susceptibility. Here, we experimentally demonstrate a giant second-order optical nonlinearity at THz frequency, orders of magnitude higher than that in the visible and microwave regimes. Different from previous work, the phonon-light coupling is achieved under a phase-matching setting, and the dynamic process of nonlinear THz generation is directly observed in a thin-film waveguide using a time-resolved imaging technique. Furthermore, a nonlinear modification to the Huang equations is proposed to explain the observed nonlinearity enhancement. This work brings about an effective approach to achieve high nonlinearity in ionic crystals, promising for applications in THz nonlinear technologies.

scholarly journals Time-resolved detection of SDS-induced conformational changes in α-synuclein by a micro-stopped-flow system

RSC Advances ◽  
2021 ◽  
Vol 11 (2) ◽  
pp. 1086-1097
Author(s):  
Shunki Takaramoto ◽  
Yusuke Nakasone ◽  
Kei Sadakane ◽  
Shinsaku Maruta ◽  
Masahide Terazima
Keyword(s):  
Conformational Changes ◽  
Flow System ◽  
Stopped Flow ◽  
Time Resolved ◽  
Sds Micelles ◽  
Conformation Changes ◽  
System P

Dynamics of conformation changes of α-synuclein induced by the presence of SDS micelles are revealed using time-resolved diffusion, CD, and FRET measurements combined with a micro-stopped flow system.

scholarly journals Influence of Oxidative Stress on Time-Resolved Oxygen Detection by [Ru(Phen)3]2+ In Vivo and In Vitro

Molecules ◽  
2021 ◽  
Vol 26 (2) ◽  
pp. 485
Author(s):  
Veronika Huntosova ◽  
Denis Horvath ◽  
Robert Seliga ◽  
Georges Wagnieres
Keyword(s):  
Oxidative Stress ◽  
Tissue Oxygenation ◽  
Intact Cell ◽  
Molecular Probes ◽  
Stress Factors ◽  
Time Resolved ◽  
Invasive Approach ◽  
Oxygen Detection

Detection of tissue and cell oxygenation is of high importance in fundamental biological and in many medical applications, particularly for monitoring dysfunction in the early stages of cancer. Measurements of the luminescence lifetimes of molecular probes offer a very promising and non-invasive approach to estimate tissue and cell oxygenation in vivo and in vitro. We optimized the evaluation of oxygen detection in vivo by [Ru(Phen)3]2+ in the chicken embryo chorioallantoic membrane model. Its luminescence lifetimes measured in the CAM were analyzed through hierarchical clustering. The detection of the tissue oxygenation at the oxidative stress conditions is still challenging. We applied simultaneous time-resolved recording of the mitochondrial probe MitoTrackerTM OrangeCMTMRos fluorescence and [Ru(Phen)3]2+ phosphorescence imaging in the intact cell without affecting the sensitivities of these molecular probes. [Ru(Phen)3]2+ was demonstrated to be suitable for in vitro detection of oxygen under various stress factors that mimic oxidative stress: other molecular sensors, H2O2, and curcumin-mediated photodynamic therapy in glioma cancer cells. Low phototoxicities of the molecular probes were finally observed. Our study offers a high potential for the application and generalization of tissue oxygenation as an innovative approach based on the similarities between interdependent biological influences. It is particularly suitable for therapeutic approaches targeting metabolic alterations as well as oxygen, glucose, or lipid deprivation.

scholarly journals Expression of TASK and TREK, two-pore domain K+ channels, in human myometrium

Reproduction ◽  
2005 ◽  
Vol 129 (4) ◽  
pp. 525-530 ◽  
Author(s):  
Xilian Bai ◽  
George J Bugg ◽  
Susan L Greenwood ◽  
Jocelyn D Glazier ◽  
Colin P Sibley ◽  
...  
Keyword(s):  
Membrane Potential ◽  
Pregnant Women ◽  
Human Myometrium ◽  
Rt Pcr ◽  
Excitable Cells ◽  
K Channels ◽  
Myometrial Cells ◽  
Channel Proteins ◽  
A Site ◽  
Pore Domain

Two-pore domain K+channels are an emerging family of K+channels that may contribute to setting membrane potential in both electrically excitable and non-excitable cells and, as such, influence cellular function. The human uteroplacental unit contains both excitable (e.g. myometrial) and non-excitable cells, whose function depends upon the activity of K+channels. We have therefore investigated the expression of two members of this family, TWIK (two-pore domain weak inward rectifying K+channel)-related acid-sensitive K+channel (TASK) and TWIK-related K+channel (TREK) in human myometrium. Using RT-PCR the mRNA expression of TASK and TREK isoforms was examined in myometrial tissue from pregnant women. mRNAs encoding TASK1, 4 and 5 and TREK1 were detected whereas weak or no signals were observed for TASK2, TASK3 and TREK2. Western blotting for TASK1 gave two bands of approximately 44 and 65 kDa, whereas TREK1 gave bands of approximately 59 and 90 kDa in myometrium from pregnant women. TASK1 and TREK1 immunofluorescence was prominent in intracellular and plasmalemmal locations within myometrial cells. Therefore, we conclude that the human myometrium is a site of expression for the two-pore domain K+channel proteins TASK1 and TREK1.

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