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KCI등재 학술저널

Ca<SUP>2+</SUP>-induced Ca<SUP>2+</SUP> Release from Internal Stores in INS-1 Rat Insulinoma Cells

Ca<SUP>2+</SUP>-induced Ca<SUP>2+</SUP> Release from Internal Stores in INS-1 Rat Insulinoma Cells

The secretion of insulin from pancreatic &#1346;-cells is triggered by the influx of Ca<sup>2+</sup> through voltage-dependent Ca<sup>2+</sup> channels. The resulting elevation of intracellular calcium ([Ca<sup>2+</sup>]<sub>i</sub>) triggers additional Ca<sup>2+</sup> release from internal stores. Less well understood are the mechanisms involved in Ca<sup>2+</sup> mobilization from internal stores after activation of Ca<sup>2+</sup> influx. The mobilization process is known as calcium-induced calcium release (CICR). In this study, our goal was to investigate the existence of and the role of caffeine-sensitive ryanodine receptors (RyRs) in a rat pancreatic &#1346;-cell line, INS-1 cells. To measure cytosolic and stored Ca<sup>2+</sup>, respectively, cultured INS-1 cells were loaded with fura-2/AM or furaptra/AM. [Ca<sup>2+</sup>]<sub>i</sub> was repetitively increased by caffeine stimulation in normal Ca<sup>2+</sup> buffer. However, peak [Ca<sup>2+</sup>]<sub>i</sub> was only observed after the first caffeine stimulation in Ca<sup>2+</sup> free buffer and this increase was markedly blocked by ruthenium red, a RyR blocker. KCl-induced elevations in [Ca<sup>2+</sup>]<sub>i </sub>were reduced by pretreatment with ruthenium red, as well as by depletion of internal Ca<sup>2+</sup> stores using cyclopiazonic acid (CPA) or caffeine. Caffeine-induced Ca<sup>2+ </sup>mobilization ceased after the internal stores were depleted by carbamylcholine (CCh) or CPA. In permeabilized INS-1 cells, Ca<sup>2+</sup> release from internal stores was activated by caffeine, Ca<sup>2+</sup>, or ryanodine. Furthermore, ruthenium red completely blocked the CICR response in permeabilized cells. RyRs were widely distributed throughout the intracellular compartment of INS-1 cells. These results suggest that caffeine-sensitive RyRs exist and modulate the CICR response from internal stores in INS-1 pancreatic &#1346;-cells.

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