optical-imaging-of-isolated-murine-ventricular-myocytes

Optical Imaging of Isolated Murine Ventricular Myocytes

The ability to isolate adult cardiac myocytes has permitted researchers to study a variety of cardiac pathologies at the single cell level. While advances ...

UNIVERSITY HOSPITALS CLEVELAND MEDICAL CENTER

Mutations in the human cardiac motor protein beta-myosin heavy chain (βMHC) have been long recognized as a cause of familial hypertrophic cardiomyopathy. Recently, mutations (P830L and A1004S) in the less abundant but faster isoform alpha-myosin heavy chain (αMHC) have been linked to dilated cardiomyopathy (DCM). In this study, we sought to determine the cellular contractile phenotype associated with these point mutations. Ventricular myocytes were isolated from 2 month male Sprague Dawley rats. Cells were cultured in M199 media and infected with recombinant adenovirus containing the P830L or the A1004S mutant human αMHC at a MOI of 500 for 18 h. Uninfected cells (UI), human βMHC (MOI 500, 18 h), and human αMHC (MOI 500, 18 h) were used as controls. Cells were loaded with fura-2 (1 μM, 15 min) after 48 h. Sarcomere shortening and calcium transients were recorded in CO buffered M199 media (36°±1 C) with and without 10 nM isoproterenol (Iso). The A1004S mutation resulted in decreased peak sarcomere shortening while P830L demonstrated near normal shortening kinetics at baseline. In the presence of Iso, the A1004S sarcomere shortening was identical to the βMHC shortening while the P830L was identical to the αMHC control. All experimental groups had identical calcium transients. Despite a shared association with DCM, the P830L and A1004S αMHC mutations alter myocyte contractility in completely different ways while at the same preserving peak intracellular calcium.

Eric Devaney

Pediatric Cardiac and Thoracic Surgery

UNIVERSITY HOSPITALS CLEVELAND MEDICAL CENTER

Optical Imaging of Isolated Murine Ventricular Myocytes

Eric Devaney

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UNIVERSITY HOSPITALS CLEVELAND MEDICAL CENTER

Optical Imaging of Isolated Murine Ventricular Myocytes

Pediatric Cardiac and Thoracic Surgery