A novel Ca2+-induced Ca2+ release mechanism mediated by neither inositol trisphosphate nor ryanodine receptors

Members of both major families of intracellular Ca²⁺ channels, ryanodine and inositol 1,4,5-trisphosphate (IP₃) receptors, are stimulated by substantial increases in cytosolic free Ca²⁺ concentration ([Ca²⁺]_c). They thereby mediate Ca²⁺-induced Ca²⁺ release (CICR), which allows amplification and regenerative propagation of intracellular Ca²⁺ signals. In permeabilized hepatocytes, increasing [Ca²⁺]_c to 10μM stimulated release of 30±1% of the intracellular stores within 60s; the EC₅₀ occurred with a free [Ca²⁺] of 170±29nM. This CICR was abolished at 2°C. The same fraction of the stores was released by CICR before and after depletion of the IP₃-sensitive stores, and CICR was not blocked by antagonists of IP₃ receptors. Ryanodine, Ruthenium Red and tetracaine affected neither the Ca²⁺ content of the stores nor the CICR response. Sr²⁺ and Ba²⁺ (EC₅₀ = 166nM and 28μM respectively) mimicked the effects of increased [Ca²⁺] on the intracellular stores, but Ni²⁺ blocked the passive leak of Ca²⁺ without blocking CICR. In rapid superfusion experiments, maximal concentrations of IP₃ or Ca²⁺ stimulated Ca²⁺ release within 80ms. The response to IP₃ was complete within 2s, but CICR continued for tens of seconds despite a slow [half-time (t_1/2) = 3.54±0.07s] partial inactivation. CICR reversed rapidly (t_1/2 = 529±17ms) and completely when the [Ca²⁺] was reduced. We conclude that hepatocytes express a novel temperature-sensitive, ATP-independent CICR mechanism that is reversibly activated by modest increases in [Ca²⁺], and does not require IP₃ or ryanodine receptors or reversal of the sarcoplasmic/endoplasmic-reticulum Ca²⁺-ATPase. This mechanism may both regulate the Ca²⁺ content of the intracellular stores of unstimulated cells and allow even small intracellular Ca²⁺ signals to be amplified by CICR.