Photorelease of inositol 1,4,5-trisphosphate (InsP3) from a caged precursor was used to study characteristics of Ca(2+)-activated Cl- currents activated in Xenopus oocytes by the InsP3-Ca2+ signaling pathway. Photolysis flashes shorter than a threshold duration evoked no response, but the current amplitude then grew about linearly as the flash duration was further lengthened. Currents directly evoked by photorelease of Ca2+ from a caged precursor grew linearly with increasing flash duration and showed a small threshold before they were activated. However, the major part of the threshold of InsP3-evoked responses appears to arise because a certain concentration of InsP3 (estimated to be approximately 60 nM) is required to evoke Ca2+ liberation. Subthreshold conditioning flashes potentiated responses to subsequent flashes, and the potentiation increased linearly with increasing conditioning flash duration before abruptly declining. The potentiation decayed exponentially with a time constant of approximately 17 s with increasing interflash interval. Currents evoked by photoreleased InsP3 began after a latency that shortened from 10 s or longer to 100 ms as the photolysis intensity was increased. This dose dependence of the latency could be quantitatively explained by the time required for the InsP3 concentration to rise above threshold. Intracellular injection of heparin (a competitive antagonist at the InsP3 receptor) increased the threshold for InsP3 action, as did increased temperature. We conclude that several characteristics of InsP3-evoked responses, including their dose dependence, latency, and facilitation with paired stimuli, arise because a distinct threshold level of InsP3 is required to evoke release of Ca2+ from intracellular stores.