Combined system for high-time-resolution dual-excitation fluorescence photometry and fluorescence imaging of calcium transients in single normal and diseased skeletal muscle fibers

1994 ◽  
Author(s):  
Dietmar Uttenweiler ◽  
Reinhold Wojciechowski ◽  
Makoto Makabe ◽  
Claudia Veigel ◽  
Rainer H. Fink
Keyword(s):  
Skeletal Muscle ◽  
Fluorescence Imaging ◽  
Time Resolution ◽  
Muscle Fibers ◽  
Calcium Transients ◽  
High Time ◽  
High Time Resolution ◽  
Combined System ◽  
Skeletal Muscle Fibers

scholarly journals Arsenazo III and antipyrylazo III calcium transients in single skeletal muscle fibers.

1982 ◽  
Vol 79 (4) ◽  
pp. 679-707 ◽  
Author(s):  
P Palade ◽  
J Vergara
Keyword(s):  
Skeletal Muscle ◽  
Voltage Clamp ◽  
Spectral Characteristics ◽  
Muscle Fibers ◽  
Calcium Transients ◽  
Arsenazo Iii ◽  
Tension Development ◽  
Skeletal Muscle Fibers ◽  
Concentration Changes

The metallochrome calcium indicators arsenazo III and antipyrylazo III have been introduced individually into cut single frog skeletal muscle fibers from which calcium transients have been elicited either by action potential stimulation or by voltage-clamp pulses of up to 50 ms in duration. Calcium transients recorded with both dyes at selected wavelengths have similar characteristics when elicited by action potentials. Longer voltage-clamp pulse stimulation reveals differences in the late phases of the optical signals obtained with the two dyes. The effects of different tension blocking methods on Ca transients were compared experimentally. Internal application of EGTA at concentrations up to 3 mM was demonstrated to be efficient in blocking movement artifacts without affecting Ca transients. Higher EGTA concentrations affect the Ca signals' characteristics. Differential effects of internally applied EGTA on tension development as opposed to calcium transients suggest that diffusion with binding from Ca++ release sites to filament overlap sites may be significant. The spectral characteristics of the absorbance transients recorded with arsenazo III suggest that in situ recorded signals cannot be easily interpreted in terms of Ca concentration changes. A more exhaustic knowledge of the dye chemistry and/or in situ complications in the use of the dye will be necessary.

scholarly journals Imaging of calcium transients in skeletal muscle fibers

1991 ◽  
Vol 59 (1) ◽  
pp. 12-24 ◽  
Author(s):  
J. Vergara ◽  
M. DiFranco ◽  
D. Compagnon ◽  
B.A. Suarez-Isla
Keyword(s):  
Skeletal Muscle ◽  
Muscle Fibers ◽  
Calcium Transients ◽  
Skeletal Muscle Fibers

scholarly journals High Time Resolution Analysis of Voltage-Dependent and Voltage-Independent Calcium Sparks in Frog Skeletal Muscle Fibers

2020 ◽  
Vol 11 ◽  
Author(s):  
Henrietta Cserne Szappanos ◽  
János Vincze ◽  
Dóra Bodnár ◽  
Beatrix Dienes ◽  
Martin F. Schneider ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Time Resolution ◽  
Ryanodine Receptors ◽  
High Time ◽  
Fiber Axis ◽  
High Time Resolution ◽  
Frog Skeletal Muscle ◽  
Calcium Sparks ◽  
Caffeine Treatment ◽  
Voltage Dependent

In amphibian skeletal muscle calcium (Ca2+) sparks occur both as voltage-dependent and voltage-independent ligand-activated release events. However, whether their properties and their origin show similarities are still in debate. Elevated K+, constant Cl– content solutions were used to initiate small depolarizations of the resting membrane potential to activate dihydropyridine receptors (DHPR) and caffeine to open ryanodine receptors (RyR) on intact fibers. The properties of Ca2+ sparks observed under control conditions were compared to those measured on depolarized cells and those after caffeine treatment. Calcium sparks were recorded on intact frog skeletal muscle fibers using high time resolution confocal microscopy (x-y scan: 30 Hz). Sparks were elicited by 1 mmol/l caffeine or subthreshold depolarization to different membrane potentials. Both treatments increased the frequency of sparks and altered their morphology. Images were analyzed by custom-made computer programs. Both the amplitude (in ΔF/F0; 0.259 ± 0.001 vs. 0.164 ± 0.001; n = 24942 and 43326, respectively; mean ± SE, p < 0.001) and the full width at half maximum (FWHM, in μm; parallel with fiber axis: 2.34 ± 0.01 vs. 1.92 ± 0.01, p < 0.001; perpendicular to fiber axis: 2.08 ± 0.01 vs. 1.68 ± 0.01, p < 0.001) of sparks was significantly greater after caffeine treatment than on depolarized cells. 9.8% of the sparks detected on depolarized fibers and about one third of the caffeine activated sparks (29.7%) overlapped with another one on the previous frame on x-y scans. Centre of overlapping sparks travelled significantly longer distances between consecutive frames after caffeine treatment then after depolarization (in μm; 1.66 ± 0.01 vs. 0.95 ± 0.01, p < 0.001). Our results suggest that the two types of ryanodine receptors, the junctional RyRs controlled by DHPRs and the parajunctional RyRs are activated independently, using alternate ways, with the possibility of cooperation between neighboring release channels.

scholarly journals Elevated nuclear Foxo1 suppresses excitability of skeletal muscle fibers

2013 ◽  
Vol 305 (6) ◽  
pp. C643-C653 ◽  
Author(s):  
Erick O. Hernández-Ochoa ◽  
Tova Neustadt Schachter ◽  
Martin F. Schneider
Keyword(s):  
Skeletal Muscle ◽  
Electrical Stimulation ◽  
Muscle Atrophy ◽  
Action Potentials ◽  
Muscle Fibers ◽  
Calcium Transients ◽  
Skeletal Muscle Atrophy ◽  
Nuclear Activity ◽  
Ec Coupling ◽  
Skeletal Muscle Fibers

Forkhead box O 1 (Foxo1) controls the expression of proteins that carry out processes leading to skeletal muscle atrophy, making Foxo1 of therapeutic interest in conditions of muscle wasting. The transcription of Foxo1-regulated proteins is dependent on the translocation of Foxo1 to the nucleus, which can be repressed by insulin-like growth factor-1 (IGF-1) treatment. The role of Foxo1 in muscle atrophy has been explored at length, but whether Foxo1 nuclear activity affects skeletal muscle excitation-contraction (EC) coupling has not yet been examined. Here, we use cultured adult mouse skeletal muscle fibers to investigate the effects of Foxo1 overexpression on EC coupling. Fibers expressing Foxo1-green fluorescent protein (GFP) exhibit an inability to contract, impaired propagation of action potentials, and ablation of calcium transients in response to electrical stimulation compared with fibers expressing GFP alone. Evaluation of the transverse (T)-tubule system morphology, the membranous system involved in the radial propagation of the action potential, revealed an intact T-tubule network in fibers overexpressing Foxo1-GFP. Interestingly, long-term IGF-1 treatment of Foxo1-GFP fibers, which maintains Foxo1-GFP outside the nucleus, prevented the loss of normal calcium transients, indicating that Foxo1 translocation and the atrogenes it regulates affect the expression of proteins involved in the generation and/or propagation of action potentials. A reduction in the sodium channel Nav1.4 expression in fibers overexpressing Foxo1-GFP was also observed in the absence of IGF-1. We conclude that increased nuclear activity of Foxo1 prevents the normal muscle responses to electrical stimulation and that this indicates a novel capability of Foxo1 to disable the functional activity of skeletal muscle.

High time resolution fluorescence imaging with a CCD camera

1991 ◽  
Vol 36 (2-3) ◽  
pp. 253-261 ◽  
Author(s):  
Nechama Lasser-Ross ◽  
Hiroyoshi Miyakawa ◽  
Varda Lev-Ram ◽  
Steven R. Young ◽  
William N. Ross
Keyword(s):  
Fluorescence Imaging ◽  
Time Resolution ◽  
Ccd Camera ◽  
High Time ◽  
High Time Resolution

scholarly journals Calcium transients in asymmetrically activated skeletal muscle fibers

1981 ◽  
Vol 36 (3) ◽  
pp. 491-507 ◽  
Author(s):  
G. Trube ◽  
J.R. Lopez ◽  
S.R. Taylor
Keyword(s):  
Skeletal Muscle ◽  
Muscle Fibers ◽  
Calcium Transients ◽  
Skeletal Muscle Fibers

scholarly journals Time Course of Individual Ca2+ Sparks in Frog Skeletal Muscle Recorded at High Time Resolution

1999 ◽  
Vol 113 (2) ◽  
pp. 187-198 ◽  
Author(s):  
Alain Lacampagne ◽  
Christopher W. Ward ◽  
Michael G. Klein ◽  
Martin F. Schneider
Keyword(s):  
Skeletal Muscle ◽  
Time Resolution ◽  
Rise Time ◽  
Laser Scanning ◽  
Time Course ◽  
High Time ◽  
High Time Resolution ◽  
Frog Skeletal Muscle ◽  
Resolution System ◽  
Constant Rate

Discrete Ca2+ release events (Ca2+ “sparks”) were recorded in cut segments of single frog skeletal muscle fibers using a video-rate laser-scanning confocal microscope operating in line-scan mode (63 μs per line). Fibers loaded with the Ca2+ indicator fluo-3 were voltage clamped at a holding potential of 0 mV, briefly reprimed at −90 mV, and then strongly depolarized with a large test pulse to activate any reprimed voltage sensors. Using this high time resolution system, it was possible to record individual Ca2+ sparks at ∼30-fold higher time resolution than previously attained. The resulting new experimental data provides a means of characterizing the time course of fluorescence during the brief (a few milliseconds) rising phase of a spark, which was not possible with the previously used 1.5–2 ms per line confocal systems. Analysis of the time course of individual identified events indicates that fluorescence begins to rise rather abruptly at the start of the spark, continues to rise at a slightly decreasing rate to a relatively sharp peak, and then declines along a quasi-exponential time course. The mean rise time of 198 sparks was 4.7 ± 0.1 ms, and there was no correlation between rise time and peak amplitude. Average sparks constructed by temporally and spatially superimposing and summing groups of individual sparks having similar rise times gave a lower noise representation of the sparks, consistent with the time course of individual events. In theory, the rising phase of a spark provides a lower bound estimation of the time that Ca2+ ions are being released by the sarcoplasmic reticulum Ca2+ channel(s) generating the spark. The observed time course of fluorescence suggests that the Ca2+ release underlying a spark could continue at a fairly constant rate throughout the rising phase of the spark, and then stop rather abruptly at the time of the peak.

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