Abstract
Traditional approaches for measuring cardiac myocyte hypertrophy have been of low throughput and subjective, limiting the scope of experimental studies designed to understand it. Here, we describe an automated image acquisition and analysis platform for studying the dynamics of cardiac myocyte hypertrophy in vitro. Image acquisition scripts record 5 × 5 mosaic images of fluorescent protein-labeled neonatal rat ventricular myocytes from each well of a 96-well plate using the microscope’s automated stage and focus. Image analysis algorithms automatically segment myocyte boundaries, track myocytes, and quantify changes in shape. We describe each step of the image acquisition and analysis algorithms and provide specific examples of how to implement them using Metamorph and CellProfiler software. With this system, shape dynamics of thousands of individual cardiac myocytes can be tracked for up to a week. This imaging platform was recently applied to study reversal of cardiac myocyte hypertrophy following withdrawal of the α-adrenergic agonist phenylephrine. Hypertrophy readily reversed at low but not high levels of α-adrenergic signaling, leading to identification of an intracellular population of α-adrenergic receptors responsible for this reversibility delay.
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Acknowledgements
This work is supported by the National Science Foundation (predoctoral fellowship to K.R., CAREER grant to J.S.) and the National Institutes of Health (grant HL094476).
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Ryall, K.A., Saucerman, J.J. (2015). Automated Microscopy of Cardiac Myocyte Hypertrophy: A Case Study on the Role of Intracellular α-Adrenergic Receptors. In: Allen, B., Hébert, T. (eds) Nuclear G-Protein Coupled Receptors. Methods in Molecular Biology, vol 1234. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-1755-6_11
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DOI: https://doi.org/10.1007/978-1-4939-1755-6_11
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Publisher Name: Humana Press, New York, NY
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Online ISBN: 978-1-4939-1755-6
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