Ca²⁺ entry through the T-type Ca²⁺ channels causes apoptosis via the mitochondrial pathway

Abstract

Background: It is postulated that T-type Ca²⁺ channels play important roles not only in physiological condition of various organs and tissues, but also during the progression of various diseases. In this study, we tested the hypothesis that Ca²⁺ entry through the T-type Ca²⁺ channels causes apoptosis using recombinant Ca_V3.2 (α1H) T-type Ca²⁺ channels expressed in HEK293 (HEK-α1H) cells. Methods and Results: Cultured HEK293 cells and HEK-α1H cells were incubated for 12 hours in serum-free medium containing different concentrations of Ca²⁺ (1.8-7.2 mM). To determine the degree of apoptosis, mitochondrial transmembrane potential (ΔΨ_m) and activities of caspase-3, -8, -9 were measured quantitatively. Intracellular free Ca²⁺ measured by flow cytometry using Fluo-3 was elevated depending on Ca²⁺ concentration in the medium ([Ca²⁺]o) in HEK-α1H cells but not in HEK293 cells. In HEK-α1H cells, apoptosis in terms of loss of ΔΨ_m and activation of caspase-3 and -9 was observed at [Ca²⁺]o of 5.4 mM or more, while caspase-8 was not activated. In contrast, apoptosis was not induced at any [Ca²⁺]o in HEK293 cells. Conclusion: We demonstrated that Ca²⁺ entry through the T-type Ca²⁺ channels causes apoptosis via the mitochondrial pathway. Thus, T-type Ca²⁺ channels may be therapeutic targets for certain pathophysiological conditions related to apoptosis. [J Physiol Sci. 2006;56 Suppl:S126]