Intracellular Ca2+ fluctuation through multiple Ca2+ regulatory proteins is inevitable for initiating apoptosis in adult cardiac myocytes

Abstract Adult cardiac myocytes show a dynamic and unique process during execution of apoptosis; they exhibit beating just before and at the early phase of apoptosis (Am J Pathol. 2001), suggesting important roles of intracellular Ca 2+ concentration ([Ca 2+ ]i) and Ca 2+ regulatory proteins for initiating apoptosis in those cells. In the present study, we examined the regulatory mechanisms of Ca 2+ regulatory proteins for initiating apoptosis in adult cardiac myocytes, which was induced by activation of β-adrenergic receptor by isoproterenol (10 −5 M). Apoptosis was evident, as documented by activated caspase-3, DNA fragmentation based on in situ nick end-labeling (TUNEL) and DNA ladder pattern, and apoptotic ultrastructure. These apoptotic features were found significantly inhibited by L-type Ca 2+ channel antagonist (nifedipine), SERCA antagonist (thapsigargin), and ryanodine receptor antagonist (ryanodine) in a dose-dependent manner. According to serial observation under a real-time videomicroscope, those antagonists inhibited not only apoptotic morphological change but also the pre-apoptotic beating. These findings suggest an important role of a fluctuation of [Ca 2+ ]i - i.e., not only an increase in [Ca 2+ ]i through L-type Ca 2+ channel or ryanodine receptor but also a decrease in it through SERCA, for initiating cardiac myocyte apoptosis, implying an exquisite regulation of initiating apoptosis in adult cardiac myocytes by multiple Ca 2+ regulatory proteins.