Expression and modulation of the intermediate-conductance Ca2+-activated K+ channel in glioblastoma GL-15 cells

We report here the expression and properties of the intermediate-conductance Ca2+-activated K+ (IKCa) channel in the GL-15 human glioblastoma cell line. Macroscopic IKCa currents on GL-15 cells displayed a mean amplitude of 7.2 +/- 0.8 pA/pF at 0 mV, at day 1 after plating. The current was inhibited by clotrimazole (CTL, IC50=257 nM), TRAM-34 (IC50=55 nM), and charybdotoxin (CTX, IC50= 10.3 nM). RT-PCR analysis demonstrated the expression of mRNA encoding the IKCa channel in GL-15 cells. Unitary currents recorded using the inside-out configuration had a conductance of 25 pS, a K-D for Ca2+ of 188 nM at -100 mV, and no voltage dependence. We tested whether the IKCa channel expression in GL-15 cells could be the result of an increased ERK activity. Inhibition of the ERK pathway with the MEK antagonist PD98059 (25 mu M, for 5 days) virtually suppressed the IKCa current in GL-15 cells. PD98059 treatment also increased the length of cellular processes and up-regulated the astrocytic differentiative marker GFAP. A significant reduction of the IKCa current amplitude was also observed with time in culture, with mean currents of 7.17 +/- 0.75 pA/pF at 1-2 days, and 3.11 +/- 1.35 pA/pF at 5-6 days after plating. This time-dependent downregulation of the IKCa current was not accompanied by changes in the ERK activity, as assessed by immunoblot analysis. Semiquantitative RT-PCR analysis demonstrated a similar to 35% reduction of the IKCa channel mRNA resulting from ERK inhibition and a similar to 50% reduction with time in culture. Copyright (c) 2006 S. Karger AG, Basel.