Microfluorometric monitoring of NAD redox state in isolated perfused renal tubules.

We report the successful application of microfluorometric techniques to monitor the redox state of mitochondrial nicotinamide adenine dinucleotide (NAD) in conjunction with measurements of transepithelial transport in intact isolated perfused tubules from the rabbit kidney. The determination of the redox state of NAD yields important information on the delivery of substrate to, and the energy demands on, the tubular mitochondria. These studies required only minor modifications of the basic perfusion apparatus. With this technique we detected net reductions of mitochondrial NAD upon the addition of metabolic substrates (succinate), the inhibition of active ion transport (ouabain), and the inhibition of mitochondrial electron flow (cyanide). Uncoupling of oxidative phosphorylation with 1799 caused an oxidation of NAD. These changes in NAD redox state are predictable from previous studies in isolated mitochondria, intact tissues, and from experiments on cortical tubule suspensions from the rabbit cortex.