Evidence for inositol tetrakisphosphate-activated Ca2+ influx pathway refilling inositol trisphosphate-sensitive Ca2+ stores in hamster eggs.

To identify the Ca2+ influx pathway responsible for maintaining Ca2+ oscillations in hamster eggs, changes in intracellular Ca2+ concentration ([Ca2+]i) were recorded using the Fura-2 fluorescent imaging technique during iontophoretic injection of inositol phosphates under voltage clamp. Both inositol 1,4,5-trisphosphate (InsP3) and 1,3,4,5-tetrakisphosphate (InsP4) caused repetitive Ca2+ transients when injected continuously into eggs, although the latter was much less effective. These Ca2+ transients were inhibited by the monoclonal antibody 18A10 to the InsP3 receptor/Ca2+ channel. In Ca(2+)-free medium, InsP4-induced Ca2+ transients were absent or much less frequent than in normal medium. A small but persistent increase in [Ca2+]i during InsP4 injection was revealed when Ca2+ uptake into InsP3-sensitive Ca2+ stores was suppressed by thapsigargin. This Ca2+ rise is due to Ca2+ entry, but not Ca2+ release, because it was: (i) increased by raising the extracellular Ca2+ concentration and abolished in Ca(2+)-free medium; (ii) larger at more negative membrane potentials which provide greater electrical driving force for Ca2+ entry; and (iii) not affected by 18A10. A moderate dose of InsP3 did not cause substantial Ca2+ entry, as tested in thapsigargin- and 18A10-treated eggs. InsP4 facilitated the restoration of Ca2+ stores after Ca2+ releases induced by pulsatile InsP3 injections. Thus, we obtained evidence for a Ca2+ influx pathway activated by InsP4 which provides Ca2+ to refill InsP3-sensitive Ca2+ stores in intact cells.