NAD(P)H Redox In Rat Heart:  Fluorescence Imaging And Spectroscopy

Kevin E. Conley, Clyde H. Barlow, Jeffrey J. Kelly, Dawn A. Rorvik and Martin J. Kushmerick

University of Washington Medical Center, Seattle, WA 98195
Barlow Scientific, Inc., Olympia, WA 98502

Conley, K.E., Barlow, C.H., Kelly, J.J., Rorvik, D.A. and Kushmerick, M.J.  (1992)  NAD(P)H redox in rat heart:  Fluorescence imaging and spectroscopy.  Biophysical Journal  61(2 part 2), A178.  Surface fluorescence imaging and spectroscopy of NAD(P)H in isolated perfused rat hearts were compared over a physiological range of work output and oxygen consumption at 25° C. Images were obtained throughout the cardiac cycle with a pixel resolution of 85 x 70 µm from fluorescence emission at 420-630 nm after a 330-380 nm excitation flash. Fluorescence spectra were obtained with a fiber optic cable and a photodiode array over 400-600 nm. An in vivo NAD(P)H fluorescence spectrum was obtained by subtracting spectra taken under control and maximal NAD(P)H reduction (5 mM pentobarbital in the perfusate). The in vivo spectrum fell predominantly (>90%) within the wavelenghts used for imaging. Comparison of the in vivo and solution NAD(P)H spectra showed close correspondence except: i) a red shift in the spectral peak between the in vivo (472 nm) and solution (465 nm) spectra and ii) a small flavoprotein shoulder at ~560 nm which contributed negligibly to the fluorescence signal. Images obtained before, during and after pacing showed a reversible heterogeneity of NAD(P)H intensity. This heterogeneity resulted from both increases and decreases in NAD(P)H intensity over the heart surface with pacing. Thus, fluorescence imaging makes possible more rigorous measurement of the direction and magnitude of NAD(P)H changes than spatially averaged spectroscopy.

Supported by NIH AR 36281, HL 37798, HL 43473, AHA 901369 and U.W. Dept. of Radiology.

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Last modified: 01/08/2000