Abstract P206: Automated Imaging Reveals a Switch Between Reversible and Irreversible Cardiac Myocyte Hypertrophy

Abstract

Cardiac hypertrophy increases risk for heart failure, arrhythmia, and sudden death. Little is known about the specific signaling pathways that distinguish reversible forms of hypertrophy from irreversible forms which lead to heart failure. This project utilizes a novel high-throughput cell phenotype imaging and analysis protocol to study the reversibility of cardiac myocyte hypertrophy in a scalable cell culture model. Primary cardiac myocytes were transfected with GFP plasmid driven under a cardiac specific troponin T promoter. Five by five mosaic images (∼100 cells) within each well of a 96-well plate were recorded with an automated XYZ stage and focus. Post-processing algorithms automatically background corrected, segmented cell edges, quantified cell phenotypes, and tracked cells between measurements. This platform therefore has the ability to track changes in area and shape of hundreds of individual cells over a time period of about a week. Cell shape changes after washout of a dose response to the hypertrophic agonist phenylephrine (PE) showed that hypertrophy was reversible at low but not high levels of alpha-adrenergic signaling: a cellular “toggle switch”. Specialized alpha-adrenergic receptor (αAR) antagonists were used to study a potential mechanism for this biphasic response. Prazosin, an αAR antagonist that could act at the sarcolemma and also be transported inside the cell, reversed PE-induced hypertrophy. In contrast, CGP-12177a, an αAR antagonist that acts at the sarcolemma, did not reverse PE-induced hypertrophy. These experimental results and a computational model support the hypothesis that cellular uptake of PE and activity at nuclear alpha-adrenergic receptors may explain the biphasic response in the reversibility of PE-induced hypertrophy.