1. The effects of the IP3-receptor antagonist 2-aminoethyldiphenyl borate (2-APB) on the Ca2+ release-activated Ca2+ current (ICRAC) in Jurkat human T cells, DT40 chicken B cells and rat basophilic leukaemia (RBL) cells were examined. 2. 2-APB elicited both stimulatory and inhibitory effects on Ca2+ influx through CRAC channels. At concentrations of 1-5 μM, 2-APB enhanced Ca2+ entry in intact cells and increased ICRAC amplitude by up to fivefold. At levels ≥ 10 μM, 2-APB caused a transient enhancement of ICRAC followed by inhibition. 3. 2-APB altered the kinetics of fast Ca2+-dependent inactivation of ICRAC. At concentrations of 1-5 μM, 2-APB increased the rate of fast inactivation. In contrast, 2-APB at higher concentrations (≥ 10 μM) reduced or completely blocked inactivation. 4. 2-APB inhibited Ca2+ efflux from mitochondria. 5. 2-APB inhibited ICRAC more potently when applied extracellularly than intracellularly. Furthermore, increased protonation of 2-APB at low pH did not affect potentiation or inhibition. Thus, 2-APB may have an extracellular site of action. 6. Neither ICRAC activation by passive store depletion nor the effects of 2-APB were altered by intracellular dialysis with 500 μg ml-1 heparin. 7. ICRAC is present in wild-type as well as mutant DT40 B cells lacking all three IP3 receptor isoforms. 2-APB also potentiates and inhibits ICRAC in both cell types, indicating that 2-APB exerts its effects independently of IP3 receptors. 8. Our results show that CRAC channel activation does not require physical interaction with IP3 receptors as proposed in the conformational coupling model. Potentiation of ICRAC by 2-APB may be a useful diagnostic feature for positive identification of putative CRAC channel genes, and provides a novel tool for exploring the physiological functions of store-operated channels.
|Original language||English (US)|
|Number of pages||17|
|Journal||Journal of physiology|
|State||Published - Oct 1 2001|
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