Using cAMP Sensors to Study Cardiac Nanodomains

Figures

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  Figure 1. Evolution of cardiac cAMP detection from bulk biochemical analysis to targeted real-time imaging in living cardiomyocytes. In all panels, cAMP molecules are represented by golden spheres; fluorophore molecules are represented by coloured spheres, with the colours indicating their emission spectra. Exposure to light with the excitation spectrum of a fluorophore is represented by a monochrome lightning bolt; emission of fluorescent light is represented by a halo around the fluorophore. Fluorescence transfer is represented by a lightning bolt that originates from the fluorescence donor, which matches the donor in colour and points towards the fluorescence acceptor. EC50 for cAMP is indicated for all cAMP detection systems, as reported in the literature [10,11,12,13].

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  Figure 2. Scales of cAMP signalling domains in adult cardiomyocytes. Continued development of cAMP detection methods improved the resolution of cAMP signal measurement within adult cardiomyocytes. (a) Initially, whole-cell and fractionation assays across a 100 μm long adult cardiomyocyte were performed. (b) Later, differences in cAMP concentrations were measured along the sarcolemmal crest and T tubules, which are spread across the membrane roughly every 2 μm. (c) Targeted sensors showed differential cAMP signalling between cytosol and mitochondrial matrix, which can be located under 1 μm apart from each other in cardiomyocytes. (d) Even differences between mitochondrial outer membrane and matrix, or sarcoplasmic reticulum and T tubules, in cardiomyocytes ca. 100 nm apart, can be detected. (e) Future sensors might be able to quantify signalling events within single signalosomes that are few nanometres apart, such as the protein complexes of the sarcomere.