title: Effect of ouabain on metabolic oxidative state in living cardiomyocytes evaluated by time-resolved spectroscopy of endogenous NAD(P)H fluorescence.
abstract: Time-resolved spectrometry of endogenous nicotinamide dinucleotide phosphate [NAD(P)H] fluorescence is a useful method to evaluate metabolic oxidative state in living cells. Ouabain is a well-known pharmaceutical drug used in the treatment of cardiovascular disease, the effects of which on myocardial metabolism were recently demonstrated. Mechanisms implicated in these actions are still poorly understood. We investigate the effect of ouabain on the metabolic oxidative state of living cardiac cells identified by time-resolved fluorescence spectroscopy of mitochondrial NAD(P)H. Spectral unmixing is used to resolve individual NAD(P)H fluorescence components. Ouabain decreased the integral intensity of NAD(P)H fluorescence, leading to a reduced component amplitudes ratio corresponding to a change in metabolic state. We also noted that lactate/pyruvate, affecting the cytosolic NADH gradient, increased the effect of ouabain on the component amplitudes ratio. Cell oxidation levels, evaluated as the percentage of oxidized NAD(P)H, decreased exponentially with rising concentrations of the cardiac glycoside. Ouabain also stimulated the mitochondrial NADH production. Our study sheds a new light on the role that ouabain plays in the regulation of metabolic state, and presents perspective on a noninvasive, pharmaceutical approach for testing the effect of drugs on the mitochondrial metabolism by means of time-resolved fluorescence spectroscopy in living cells.
authors.affiliation: Department of Biophotonics, International Laser Center, 84104 Bratislava, Slovakia. alzbeta.chorvatova@ilc.sk
authors.forename: Alzbeta
authors.initials: A
authors.lastname: Chorvatova
authors.forename: Fathia
authors.initials: F
authors.lastname: Elzwiei
authors.forename: Anton
authors.initials: A
authors.lastname: Mateasik
authors.forename: Dusan
authors.initials: D
authors.lastname: Chorvat
mesh_terms: D000818:Animals; D002316:Cardiotonic Agents / Q000494:pharmacology; D005260:Female; D019344:Lactic Acid / Q000378:metabolism; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism; D032383:Myocytes, Cardiac / Q000187:drug effects* / Q000378:metabolism*; D009243:NAD / Q000378:metabolism*; D009249:NADP / Q000378:metabolism; D010042:Ouabain / Q000494:pharmacology*; D010084:Oxidation-Reduction / Q000187:drug effects; D019289:Pyruvic Acid / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D013050:Spectrometry, Fluorescence / Q000379:methods*
chemical_list: D002316:Cardiotonic Agents; D009243:NAD; D019344:Lactic Acid; D009249:NADP; D010042:Ouabain; D019289:Pyruvic Acid