The role of dye affinity in optical measurements of Cai(2+) transients in cardiac muscle

Previous experiments in cultures of neonatal rat myocytes demonstrated that the shape of Cai(2+) transients measured using high-affinity Ca(2+)-sensitive dyes may be misrepresented. The purpose of this study was to examine the role of dye affinity in Cai(2+) measurements in intact adult cardiac tiss...

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Published in:American journal of physiology. Heart and circulatory physiology 2014-07, Vol.307 (1), p.H73
Main Authors: Kong, Wei, Fast, Vladimir G
Format: Article
Language:English
Subjects:
Animals
Binding Sites
Calcium - metabolism
Calcium Signaling - physiology
Fluorescent Dyes - pharmacokinetics
Heart Ventricles - metabolism
In Vitro Techniques
Membrane Potentials - physiology
Pyridinium Compounds - pharmacokinetics
Reproducibility of Results
Sensitivity and Specificity
Swine
Voltage-Sensitive Dye Imaging - methods
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Fast, Vladimir G
description Previous experiments in cultures of neonatal rat myocytes demonstrated that the shape of Cai(2+) transients measured using high-affinity Ca(2+)-sensitive dyes may be misrepresented. The purpose of this study was to examine the role of dye affinity in Cai(2+) measurements in intact adult cardiac tissue by comparing optical recordings obtained with high- and low-affinity dyes. Experiments were carried out in porcine left ventricular (LV) wedge preparations stained locally by intramural injection via microcapillaries (diameter = 150 μm) with a low-affinity Ca(2+)-sensitive dye Fluo-4FF or Fluo-2LA (nominal Kd, ~7-10 μmol/l), high-affinity dye Rhod-2 (Kd = 0.57 μmol/l), and Fluo-4 or Fluo-2MA (Kd, ~0.4 μmol/l); in addition, tissue was stained with transmembrane potential (Vm)-sensitive dye RH-237. Optical recordings of Vm and Cai(2+) were made using optical fibers (diameter = 325 μm) glued with the microcapillaries. The durations of Cai(2+) transients measured at 50% level of recovery (CaD50) using high-affinity Fluo-4/Fluo-2MA dyes were up to ~81% longer than those measured with low-affinity Fluo-4FF/Fluo-2LA at long pacing cycle lengths (CL). In Fluo-4/Fluo-2MA measurements at long CLs, Cai(2+) transients often (~50% of cases) exhibited slow upstroke rise and extended plateau. In Rhod-2 measurements, CaD50 was moderately longer (up to ~35%) than in Fluo-4FF recordings, but Cai(2+) transient shapes were similar. In all series of measurements, mean action potential duration values were not significantly different (P > 0.05). The delays between Vm and Cai(2+) upstrokes were comparable for low- and high-affinity dyes (P > 0.05). In conclusion, measurements of Cai(2+) transient in ventricular myocardium are strongly affected by the affinity of Ca(2+) dyes. The high-affinity dyes may overestimate the duration and alter the shape of Cai(2+) transients.
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The purpose of this study was to examine the role of dye affinity in Cai(2+) measurements in intact adult cardiac tissue by comparing optical recordings obtained with high- and low-affinity dyes. Experiments were carried out in porcine left ventricular (LV) wedge preparations stained locally by intramural injection via microcapillaries (diameter = 150 μm) with a low-affinity Ca(2+)-sensitive dye Fluo-4FF or Fluo-2LA (nominal Kd, ~7-10 μmol/l), high-affinity dye Rhod-2 (Kd = 0.57 μmol/l), and Fluo-4 or Fluo-2MA (Kd, ~0.4 μmol/l); in addition, tissue was stained with transmembrane potential (Vm)-sensitive dye RH-237. Optical recordings of Vm and Cai(2+) were made using optical fibers (diameter = 325 μm) glued with the microcapillaries. The durations of Cai(2+) transients measured at 50% level of recovery (CaD50) using high-affinity Fluo-4/Fluo-2MA dyes were up to ~81% longer than those measured with low-affinity Fluo-4FF/Fluo-2LA at long pacing cycle lengths (CL). In Fluo-4/Fluo-2MA measurements at long CLs, Cai(2+) transients often (~50% of cases) exhibited slow upstroke rise and extended plateau. In Rhod-2 measurements, CaD50 was moderately longer (up to ~35%) than in Fluo-4FF recordings, but Cai(2+) transient shapes were similar. In all series of measurements, mean action potential duration values were not significantly different (P &gt; 0.05). The delays between Vm and Cai(2+) upstrokes were comparable for low- and high-affinity dyes (P &gt; 0.05). In conclusion, measurements of Cai(2+) transient in ventricular myocardium are strongly affected by the affinity of Ca(2+) dyes. The high-affinity dyes may overestimate the duration and alter the shape of Cai(2+) transients.</description><identifier>EISSN: 1522-1539</identifier><identifier>DOI: 10.1152/ajpheart.00751.2013</identifier><identifier>PMID: 24791783</identifier><language>eng</language><publisher>United States</publisher><subject>Animals ; Binding Sites ; Calcium - metabolism ; Calcium Signaling - physiology ; Fluorescent Dyes - pharmacokinetics ; Heart Ventricles - metabolism ; In Vitro Techniques ; Membrane Potentials - physiology ; Pyridinium Compounds - pharmacokinetics ; Reproducibility of Results ; Sensitivity and Specificity ; Swine ; Voltage-Sensitive Dye Imaging - methods</subject><ispartof>American journal of physiology. 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Heart and circulatory physiology</title><addtitle>Am J Physiol Heart Circ Physiol</addtitle><description>Previous experiments in cultures of neonatal rat myocytes demonstrated that the shape of Cai(2+) transients measured using high-affinity Ca(2+)-sensitive dyes may be misrepresented. The purpose of this study was to examine the role of dye affinity in Cai(2+) measurements in intact adult cardiac tissue by comparing optical recordings obtained with high- and low-affinity dyes. Experiments were carried out in porcine left ventricular (LV) wedge preparations stained locally by intramural injection via microcapillaries (diameter = 150 μm) with a low-affinity Ca(2+)-sensitive dye Fluo-4FF or Fluo-2LA (nominal Kd, ~7-10 μmol/l), high-affinity dye Rhod-2 (Kd = 0.57 μmol/l), and Fluo-4 or Fluo-2MA (Kd, ~0.4 μmol/l); in addition, tissue was stained with transmembrane potential (Vm)-sensitive dye RH-237. Optical recordings of Vm and Cai(2+) were made using optical fibers (diameter = 325 μm) glued with the microcapillaries. The durations of Cai(2+) transients measured at 50% level of recovery (CaD50) using high-affinity Fluo-4/Fluo-2MA dyes were up to ~81% longer than those measured with low-affinity Fluo-4FF/Fluo-2LA at long pacing cycle lengths (CL). In Fluo-4/Fluo-2MA measurements at long CLs, Cai(2+) transients often (~50% of cases) exhibited slow upstroke rise and extended plateau. In Rhod-2 measurements, CaD50 was moderately longer (up to ~35%) than in Fluo-4FF recordings, but Cai(2+) transient shapes were similar. In all series of measurements, mean action potential duration values were not significantly different (P &gt; 0.05). The delays between Vm and Cai(2+) upstrokes were comparable for low- and high-affinity dyes (P &gt; 0.05). In conclusion, measurements of Cai(2+) transient in ventricular myocardium are strongly affected by the affinity of Ca(2+) dyes. 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The purpose of this study was to examine the role of dye affinity in Cai(2+) measurements in intact adult cardiac tissue by comparing optical recordings obtained with high- and low-affinity dyes. Experiments were carried out in porcine left ventricular (LV) wedge preparations stained locally by intramural injection via microcapillaries (diameter = 150 μm) with a low-affinity Ca(2+)-sensitive dye Fluo-4FF or Fluo-2LA (nominal Kd, ~7-10 μmol/l), high-affinity dye Rhod-2 (Kd = 0.57 μmol/l), and Fluo-4 or Fluo-2MA (Kd, ~0.4 μmol/l); in addition, tissue was stained with transmembrane potential (Vm)-sensitive dye RH-237. Optical recordings of Vm and Cai(2+) were made using optical fibers (diameter = 325 μm) glued with the microcapillaries. The durations of Cai(2+) transients measured at 50% level of recovery (CaD50) using high-affinity Fluo-4/Fluo-2MA dyes were up to ~81% longer than those measured with low-affinity Fluo-4FF/Fluo-2LA at long pacing cycle lengths (CL). In Fluo-4/Fluo-2MA measurements at long CLs, Cai(2+) transients often (~50% of cases) exhibited slow upstroke rise and extended plateau. In Rhod-2 measurements, CaD50 was moderately longer (up to ~35%) than in Fluo-4FF recordings, but Cai(2+) transient shapes were similar. In all series of measurements, mean action potential duration values were not significantly different (P &gt; 0.05). The delays between Vm and Cai(2+) upstrokes were comparable for low- and high-affinity dyes (P &gt; 0.05). In conclusion, measurements of Cai(2+) transient in ventricular myocardium are strongly affected by the affinity of Ca(2+) dyes. The high-affinity dyes may overestimate the duration and alter the shape of Cai(2+) transients.</abstract><cop>United States</cop><pmid>24791783</pmid><doi>10.1152/ajpheart.00751.2013</doi></addata></record>
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subjects Animals
Binding Sites
Calcium - metabolism
Calcium Signaling - physiology
Fluorescent Dyes - pharmacokinetics
Heart Ventricles - metabolism
In Vitro Techniques
Membrane Potentials - physiology
Pyridinium Compounds - pharmacokinetics
Reproducibility of Results
Sensitivity and Specificity
Swine
Voltage-Sensitive Dye Imaging - methods
title The role of dye affinity in optical measurements of Cai(2+) transients in cardiac muscle
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