Application of flow cytometry in the study of apoptosis in neonatal rat cardiomyocytes

• Published in: Methods and findings in experimental and clinical pharmacology Vol. 29; no. 10; p. 681
• Main Authors: Kniewald, H, Malcic, I, Radosevic, K, Gaurina Srcek, V, Slivac, I, Polancec, D, Matijasic, M, Kniewald, J, Kniewald, Z
• Format: Journal Article
• Language: English
• Published: Spain 01.12.2007
• Subjects: Analysis of Variance; Animals; Animals, Newborn; Apoptosis - drug effects; Benzimidazoles - chemistry; Carbocyanines - chemistry; Cell Proliferation - drug effects; Cell Survival - drug effects; Cells, Cultured; Dose-Response Relationship, Drug; Flow Cytometry - methods; Fluorescent Dyes - chemistry; Isoproterenol - pharmacology; Microscopy, Confocal - methods; Microscopy, Electron, Scanning; Myocytes, Cardiac - cytology; Myocytes, Cardiac - drug effects; Myocytes, Cardiac - ultrastructure; Norepinephrine - pharmacology; Rats
• ISSN: 0379-0355
• DOI: 10.1358/mf.2007.29.10.1147767

Depending on the concentration, catecholamines activate various intracellular signaling pathways and can induce apoptosis in cardiac myocytes. Although 5,5',6,6'-tetrachloro-1,1',3,3'-tetraethylbenzimidazolocarbocyanine iodide (JC-1) has been previously used to study mitochondria in intact cardiomyocytes, there have been no reports on the detection of apoptosis in neonatal cardiomyocytes in combination with flow cytometry and confocal microscopy. In our study, neonatal rat cardiomyocytes were exposed to norepinephrine (NE) and isoproterenol (ISO) in concentrations of 1 and 10 microM for 48 h. NE concentrations of 1 and 10 microM decreased the number of viable cardiomyocytes by 18% (*p < 0.05) and 24% (**p = 0.01), respectively. ISO in a concentration of 1 microM increased the number of viable cardiomyocytes by 13% while 10 microM decreased the number of viable cardiomyocytes by 43% (***p < 0.001). Apoptotic cells were detected by flow cytometry and confocal microscopy. NE in concentrations of 1 and 10 microM increased the percentage of apoptotic cells by 12.2% and 34.3%, respectively, while ISO alone in a concentration of 10 microM increased the percentage of apoptotic cells by 11.3%. The results demonstrated that these two methods are reliable and suitable for the detection and study of apoptosis in cultures of neonatal cardiomyocytes.