High Extracellular Ca2+ Stimulates Ca2+-Activated Cl− Currents in Frog Parathyroid Cells through the Mediation of Arachidonic Acid Cascade

Abstract

Elevation of extracellular Ca2+ concentration induces intracellular Ca2+ signaling in parathyroid cells. The response is due to stimulation of the phospholipase C/Ca2+ pathways, but the direct mechanism responsible for the rise of intracellular Ca2+ concentration has remained elusive. Here, we describe the electrophysiological property associated with intracellular Ca2+ signaling in frog parathyroid cells and show that Ca2+ -activated Cl2 channels are activated by intracellular Ca2+ increase through an inositol 1,4,5-trisphophate (IP3)-independent pathway. High extracellular Ca2+ induced an outwardly-rectifying conductance in a dose-dependent manner (EC50,6 mM). The conductance was composed of an instantaneous time independent component and a slowly activating time-dependent component and displayed a deactivating inward tail current. Extracellular Ca2+ -induced and Ca2+ dialysis-induced currents reversed at the equilibrium potential of Cl2 and were inhibited by niflumic acid (a specific blocker of Ca2+ -activated Cl2 channel). Gramicidin-perforated whole-cell recording displayed the shift of the reversal potential in extracellular Ca2+ -induced current, suggesting the change of intracellular Cl2 concentration in a few minutes. Extracellular Ca2+ -induced currents displayed a moderate dependency on guanosine triphosphate (GTP). All blockers for phospholipase C, diacylglycerol (DAG) lipase, monoacylglycerol (MAG) lipase and lipoxygenase inhibited extracellular Ca2+ -induced current. IP3 dialysis failed to induce conductance increase, but 2- arachidonoylglycerol (2-AG), arachidonic acid and 12S-hydroperoxy-5Z,8Z,10E,14Z-eicosatetraenoic acid (12(S)-HPETE) dialysis increased the conductance identical to extracellular Ca2+ -induced conductance. These results indicate that high extracellular Ca2+ raises intracellular Ca2+ concentration through the DAG lipase/lipoxygenase pathway, resulting in the activation of Cl2 conductance.