Identifying the Ca++ signalling sources activating chloride currents in Xenopus oocytes using ionomycin and thapsigargin

The calcium ionophore, ionomycin (IM), and the sarcoplasmic/endoplasmic reticulum (SER) calcium pump inhibitor, thapsigargin (TG), were used to study the roles of Ca++ from different sources in regulating Ca++-dependent Cl- currents in Xenopus oocytes. The Ca++-dependent Cl- currents, Ic, were measured in voltage-clamped oocytes (Vc = -60 mV). In the presence of extracellular Ca++, both TG (0.1 to 10 muM) and IM (0.1 to 10 muM) induce release of Ca++ from SER and activated capacitative Ca++ entry (CCE) across the plasma membrane leading to activation of both "fast" and "slow" Cl- currents. The fast Ic was produced by Ca++ release from SER while Ca++ entry across the plasma membrane activated the slow Ic. Intracellular application of the calcium buffer, BAPTA, blocked activation of the slow Ic due to Ca++ entry via CCE pathways, but not via IM-mediated movement across the plasma membrane. It is concluded that predominantly Ca++ release from stores regulates a fast Ic while Ca++ entry through CCE pathways regulates a slow Ic. Further, the CCE and slow Ic pathways must be located in spatially separated compartments since BAPTA can effectively abolish the effects of Ca++ entry via the CCE pathway, but not by the IM-mediated entry pathway.