Volume-activated, chloride currents contribute to the resting conductance and invasive migration of human glioma cells

We used an in vitro model for glioma cell invasion (transwell migration assay) and patch-clamp techniques to investigate the role of volume-activated Cl- currents (I-Cl,I-Vol) in glioma cell invasion. Hypotonic solutions (approximate to 230 mOsm) activated outwardly rectifying currents that reversed near the equilibrium potential for Cl- ions (E-Cl). These currents (I-Cl,I-Vol) were sensitive to several known Cl- channel inhibitors, including DIDS, tamoxifen, and 5-nitro-2-(3-phenylpropylamino)-benzoate (NPPB). The IC50 for NPPB inhibition of I-Cl,I-Vol was 21 mum. Under isotonic, conditions, NPPB (165 mum) blocked inward currents (at -40 mV) and increased input resistance In both standard whole-cell recordings and amphotericin perforated-patch recordings. Reducing [Cl-](circle) under isotonic conditions positively shifted the reversal potential of whole-cell currents. These findings suggest a significant resting Cl- conductance in glioma cells. Under isotonic and hypotonic conditions, Cl- channels displayed voltage- and time-dependent inactivation and had an I->Cl- permeability. To assess the potential role of these channels in cell migration, we studied the chemotactic migration of glioma cells toward laminin or vitronectin in a Boyden chamber containing transwell filters with 8 mum pores. Inhibition of I-Cl,I-Vol with NPPB reduced chemotactic migration in a dose-dependent fashion with an IC50 of 27 mum. Time-lapse video microscopy during patch-clamp recordings revealed visible changes in cell shape and/or movement that accompanied spontaneous activation of I-Cl,I-Vol, suggesting that I-Cl,I-Vol is activated during cell movement, consistent with the effects of NPPB in migration assays. We propose that I-Cl,I-Vol contributes to cell shape and volume changes required for glioma cell migration through brain tissue.