Simultaneous measurements of mitochondrial NADH and Ca2+ during increased work in intact rat heart trabeculae

The main goal of this study is to investigate the role of mitochondrial [Ca2+], [Ca2+](m), in the possible up-regulation of the NADH production rate during increased workload. Such up-regulation is necessary to support increased flux through the electron transport chain and increased ATP synthesis rates. Intact cardiac trabeculae were loaded with Rhod-2(AM), and [Ca2+](m), and mitochondrial [NADH] ([NADH](m)) were simultaneously measured during increased pacing frequency. It was found that 53% of Rhod-2 was localized in mitochondria. Increased pacing frequency caused a fast, followed by a slow rise of the Rhod-2 signal, which could be attributed to an abrupt increase in resting cytosolic [Ca2+], and a more gradual rise of [Ca2+](m), respectively. When the pacing frequency was increased from 0.25 to 2 Hz, the slow Rhod-2 component and the NADH signal increased by 18 and 11%, respectively. Based on a new calibration method, the 18% increase of the Rhod-2 signal was calculated to correspond to a 43% increase of [Ca2+](m). There was also a close temporal relationship between the rise (time constant similar to25 s) and fall (time constant similar to65 s) of [Ca2+](m), and [NADH](m) when the pacing frequency was increased and decreased, respectively, suggesting that increased workload and [Ca2+](c) cause increased [Ca2+](m) and consequently up-regulation of the NADH production rate.