Spatial and temporal characterization of structure and function of contracting single cardiomyocytes in the three-dimensional nano- to micro-domain

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The heart is composed of billions of cardiomyocytes, the functional units of the myocardium. Each cell is a highly compartmentalised. This compartmentalisation is necessary for proper cell function, and disturbances of the inner architecture are a tell-tale sign of all chronic cardiac diseases. But even in the healthy heart, sub-cellular compartments are deformed with every heartbeat. While it is astonishing how robust cardiac cell function is in this context, it is evident that we know very little about the nature of three-dimensional nano-deformations, their roles in cell activity, and – thus – their potential relevance for disease or therapy. One of the membranous compartments essential for cardiomyocyte function are so-called T-tubules (T-tub) – extracellular fluid-filled membrane invaginations that traverse the entire cell, and help to coordinate electro-mechanical function. Our pilot data shows that, as cardiomyocytes contract and relax, the shape and orientation of T-tub change drastically. I suggest that this cyclic deformation of T-tub is critical to their function, supporting T-tub content mixing and driving exchange with fluid in the bulk extracellular space. This could play a role in maintaining spatially-uniform ion concentration across the T-tub network, relevant for normal electrical activation and mechanical function. I further propose that mechanical deformation is transmitted to neighbouring cell organelles, such as the sarcoplasmic reticulum, mitochondria, the contractile lattice, and microtubules, again with important implications for signalling processes that depend on coordinated, topologically defined interaction spaces. As a consequence, pathological changes in structures such as T-tub disturb beat-by-beat auto-regulatory dynamics, contributing to organ level dysfunction and disease progression.In the proposed project, I will use state-of-the-art high-speed, nano-resolution 3D imaging of murine, rabbit, and human cells from healthy and diseased hearts, enabled by combining advanced optogenetic and high-pressure freezing approaches, to link sub-cellular structure to live-cell function. By taking advantage of a unique combination of cutting-edge techniques to study normal and pathological behaviour in multiple species, I expect to uncover the functional impact of beat-by-beat alterations of sub-cellular component architecture, to reveal novel mechanisms of cardiac autoregulation that may be targeted for treatment or prevention of cardiac disease, and to provide a representative ‘construction drawing for the architecture’ of the functional units of heart.