Probing of cardiomyocyte metabolism by spectrally resolved lifetime detection of NAD(P)H fluorescence

NAD(P)H, crucial in effective management of cellular oxidative metabolism and the principal electron donors for enzymatic reactions, is a major source of autofluorescence induced in cardiac cells following excitation by UV light. Spectrally-resolved time-correlated single photon counting was used to simultaneously measure the fluorescence spectra and fluorescence lifetimes of NAD(P)H, following excitation by a pulsed picosecond 375 nm laser diode. Spectra, as well as fluorescence lifetimes of NADH and NADPH molecules were investigated in solution at different concentrations Effects of their respective dehydrogenation by lipoamide dehydrogenase (LipDH) or glutathione reductase (GR) were also questioned. NAD(P)H autofluorescence recorded in vitro was compared to the one measured in freshly isolated cardiac cells. We observed a good comparability between NAD(P)H parameters recorded in solution and in cells.