Control of IP₃-mediated Ca²⁺ puffs in Xenopus laevis oocytes by the Ca²⁺-binding protein parvalbumin

1. Elementary events of Ca²⁺ release (Ca²⁺ puffs) can be elicited from discrete clusters of inositol 1,4,5 trisphosphate receptors (IP₃Rs) at low concentrations of IP₃. Ca²⁺ puffs have rarely been observed unless elicited by either hormone treatment or introduction of IP₃ into the cell. However, cells appear to have sufficient concentrations of IP₃ (0.1-3.0 μM) to induce Ca²⁺ release under resting conditions. 2. Here, we investigated Ca²⁺ puff activity in non-stimulated Xenopus oocytes using confocal microscopy. The fluorescent Ca²⁺ dye indicators Calcium Green 1 and Oregon Green 488 BAPTA-2 were injected into oocytes to monitor basal Ca²⁺ activity. 3. In this preparation, injection or overexpression of parvalbumin, an EF-hand Ca²⁺-binding protein (CaBP), induced Ca²⁺ puffs in resting Xenopus oocytes. This activity was inhibited by heparin, an IP₃R channel blocker, and by mutation of the Ca²⁺-binding sites in parvalbumin. 4. Ca²⁺ puff activity was also evoked by injection of low concentrations of the Ca²⁺ chelator EGTA, but not by calbindin D_28k, another member of the EF-hand CaBP superfamily. 5. BAPTA and the Ca²⁺ indicator dye Oregon Green 488 BAPTA-1 evoked Ca²⁺ puff activity, while the dextran conjugate of Oregon Green 488 BAPTA-1 did not. These data indicate that a Ca²⁺ buffer must be mobile in order to increase Ca²⁺ puff activity. 6. Together, the data indicate that some IP₃Rs spontaneously release Ca²⁺ under resting concentrations of IP₃. These elementary Ca²⁺ events appear to be below the level of detection of current imaging techniques. We suggest that parvalbumin evokes Ca²⁺ puffs by coordinating the activity of elementary IP₃R channel openings. 7. We conclude that Ca²⁺ release can be evoked not only by hormone-induced increases in IP₃, but also by expression of mobile cytosolic CaBPs under resting concentrations of IP₃.