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Abstract

Bioengineering

A Fluorescent Intravital Imaging Approach to Study Load-Induced Calcium Signaling Dynamics in Mouse Osteocytes

Published: February 24th, 2023

DOI:

10.3791/64366

1Meinig School of Biomedical Engineering, Cornell University, 2Sibley School of Mechanical and Aerospace Engineering, Cornell University, 3Department of Biomedical Engineering, The City College of New York, 4Department of Neuroscience, Albert Einstein College of Medicine, 5Department of Orthopaedic Surgery, Albert Einstein College of Medicine, 6Department of Molecular Pharmacology, Albert Einstein College of Medicine

Abstract

Bone tissue is exquisitely sensitive to differences in mechanical load magnitude. Osteocytes, dendritic cells that form a syncytium throughout the bone, are responsible for the mechanosensory function of bone tissue. Studies employing histology, mathematical modeling, cell culture, and ex vivo bone organ cultures have greatly advanced the understanding of osteocyte mechanobiology. However, the fundamental question of how osteocytes respond to and encode mechanical information at the molecular level in vivo is not well understood. Intracellular calcium concentration fluctuations in osteocytes offer a useful target for learning more about acute bone mechanotransduction mechanisms. Here, we report a method for studying osteocyte mechanobiology in vivo, combining a mouse strain with a fluorescently genetically encoded calcium indicator expressed in osteocytes with an in vivo loading and imaging system to directly detect osteocyte calcium levels during loading. This is achieved with a three-point bending device that can deliver well-defined mechanical loads to the third metatarsal of living mice while simultaneously monitoring fluorescently indicated calcium responses of osteocytes using two-photon microscopy. This technique allows for direct in vivo observation of osteocyte calcium signaling events in response to whole bone loading and is useful in the endeavor to reveal mechanisms in osteocyte mechanobiology.

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Keywords Fluorescent Intravital Imaging

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