Ca2+- and volume-sensitive chloride currents are differentially regulated by agonists and store-operated Ca2+ entry

Using patch-clamp and calcium imaging techniques, we characterized the effects of ATP and histamine on human keratinocytes. In the HaCaT cell line, both receptor agonists induced a transient elevation of [Cd2+](i) in a Ca2+-free medium followed by a secondary [Ca2+], rise upon Ca2+ readmission due to store-operated calcium entry (SOCE). In voltage-clamped cells, agonists activated two kinetically distinct currents, which showed differing voltage dependences and were identified as Ca2+-activated (I-Cl(Ca)) and volume-regulated (I-Cl,I-swell) chloride currents. NPPB and DIDS more efficiently inhibited I-Cl(Ca) and I-Cl,I-swell, respectively. Cell swelling caused by hypotonic solution invariably activated I-Cl,I-swell while regulatory volume decrease occurred in intact cells, as was found in flow cytometry experiments. The PLC inhibitor U-73122 blocked both agonist- and cell swelling-induced I-Cl,I-swell while its inactive analogue U-73343 had no effect. I-Cl(Ca) Could be activated by cytoplasmic calcium increase due to thapsigargin (TG)-induced SOCE as well as by buffering [Ca2+](i) in the pipette solution at 500 nM. In contrast, I-Cl,I-swell could be directly activated by 1-oleoyl-2-acetyl-sn-glycerol (OAG), a cell-permeable DAG analogue, but neither by InsP(3), infusion nor by the cytoplasmic calcium increase. PKC also had no role in its regulation. Agonists, OAG, and cell in a nonarlditive manner, Suggesting their convergence on a common pathway. I-Cl,I-swell and I-Cl(Ca) and swelling induced I-Cl(Ca) showed only a limited overlap (i.e., simultaneous activation), although various maneuvers were able to induce these currents sequentially in the same cell. TG-induced SOCE strongly potentiated I-Cl(Ca), but abolished I-Cl,I-swell, thereby providing a clue for this paradox. Thus, we have established for the first time using a keratinocyte model that I-Cl,I-swell can be physiologically activated under isotonic conditions by receptors coupled to the phosphoinositide pathway. These results also suggest a novel function for SOCE, which can operate as a "selection" switch between closely localized channels.