The luminal Ca<sup>2+</sup> chelator, TPEN, inhibits NAADP-induced Ca<sup>2+</sup> release
Morgan AJ., Parrington J., Galione A.
The regulation of Ca 2+ release by luminal Ca 2+ has been well studied for the ryanodine and IP 3 receptors but has been less clear for the NAADP-regulated channel. In view of conflicting reports, we have re-examined the issue by manipulating luminal Ca 2+ with the membrane-permeant, low affinity Ca 2+ buffer, TPEN, and monitoring NAADP-induced Ca 2+ release in sea urchin egg homogenate. NAADP-induced Ca 2+ release was almost entirely blocked by TPEN (IC 50 17-25μM) which suppressed the maximal extent of Ca 2+ release without altering NAADP sensitivity. In contrast, Ca 2+ release via IP 3 receptors was 3- to 30-fold less sensitive to TPEN whereas that evoked by ionomycin was essentially unaffected. The effect of TPEN on NAADP-induced Ca 2+ release was not due to an increase in the luminal pH or chelation of trace metals since it could not be mimicked by NH 4 Cl or phenanthroline. The fact that TPEN had no effect upon ionophore-induced Ca 2+ release also argued against a substantial reduction in the driving force for Ca 2+ efflux. We propose that, in the sea urchin egg, luminal Ca 2+ is important for gating native NAADP-regulated two-pore channels. © 2012 Elsevier Ltd.