Dynamic process of apoptosis in adult rat cardiomyocytes analyzed using 48-hour videomicroscopy and electron microscopy - Beating and rate are associated with the apoptotic process

Dynamic process of apoptosis has not been elucidated in adult rat cardiomyo cytes. Soluble Fas ligand (0.1 mug/ml) in the presence of actinomycin D (0- 05 mug/ml) induced apoptosis in cultured adult rat cardiomyocytes, as docum ented by activated caspase-3, DNA fragmentation, and apoptotic ultrastructu re. In the present model, we observed 60 adult cardiomyocytes with a normal rod shape under a real-time videomicroscope continuously for 48 hours. Sev enteen cells (28%) were unchanged and 7 cells (12%) showed oncosis (so-call ed necrosis) in which no beating was evident. In the remaining 36 cells (ap optosis, 60%), a slow beating (17 +/- 3/min) was initiated 16 +/- I hours l ater. Approximately I hour later, the rod cells showed long-axial shortenin g as bone- or club-like, or square-shaped, accompanied with faster beating rates (35 +/- 7/min). In 29 cells (type Al and A2), marked shrinkage occurr ed; the cellular shape became almost completely round with a smooth surface and the beating ceased 3.0 +/- 0.4 hours later. Then, smooth budding appea red 0.6 +/- 0.2 hours later. Apoptotic bodies were found in 8 cells 10 +/- 4 hours later (type Al, 13%) but not in 21 cells (type A2, 35%). in the oth er 7 cells (type A3, 12%), the cell surface became rough 8 +/- 3 hours late r and the beating ceased. Maximal beating rate was greatest in type Al (72 +/- 26/min) and greater in type A2 (29 +/- 5/min) than in type A3 (10 +/- 2 /min). Electron microscopy confirmed apoptotic ultrastructure even in the c ardiomyocytes with bone-, club-like, or square shapes, suggesting that type A3 as well as Al and A2 is also under apoptotic process. A caspase inhibit or, zVAD.fmk, blocked beating, apoptotic morphology, and DNA fragmentation, indicating these depended on caspase activation. in the caspase-dependent apoptotic process of cultured adult cardiomyocytes, beating and the followi ng deformity of the cellular edges were the initial signs and the rate of b eating was related with the subsequent three different processes of apoptos is.