ISSN:
1432-2013
Keywords:
Key words Caged calcium
;
Calcium indicators
;
Flash photolysis
;
Fluorescence transients
;
DM-nitrophen
;
Kinetic modeling
Source:
Springer Online Journal Archives 1860-2000
Topics:
Medicine
Notes:
Abstract We describe a high temporal resolution confocal spot microfluorimetry setup which makes possible the detection of fluorescence transients elicited by Ca2+ indicators in response to large (50–200 μM), short duration (〈100 ns), free [Ca2+] transients generated by laser flash photolysis of DM-nitrophen (DM-n; caged Ca2+). The equilibrium and kinetic properties of the commercially available indicators Fluo-3, Rhod-2, CalciumOrange-5N (COr-5N) and CalciumGreen-2 (CGr-2) were determined experimentally. The data reveal that COr-5N displays simple, fast response kinetics while, in contrast, Fluo-3, Rhod-2 and CGr-2 are characterized by significantly slower kinetic properties. These latter indicators may be unsuitable for tracking Ca2+ signaling events lasting only a few milliseconds. A model which accurately predicts the time course of fluorescence transients in response to rapid free [Ca2+] changes was developed. Experimental data and model predictions concur only when the association rate constant of DM-n is approximately 20 times faster than previously reported. This work establishes a quantitative theoretical framework for the study of fast Ca2+ signaling events and the use of flash photolysis in cells and model systems.
Type of Medium:
Electronic Resource
URL:
http://dx.doi.org/10.1007/s004240050444
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