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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Bibliographic Details
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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Summary: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.
ISSN:1522-1539
DOI:10.1152/ajpheart.00751.2013