Expression and modulation of the intermediate-conductance Ca 2+-activated K+ channel in glioblastoma GL-15 cells

We report here the expression and properties of the intermediate- conductance Ca²⁺-activated K⁺ (IK_Ca) channel in the GL-15 human glioblastoma cell line. Macroscopic IK_Ca 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, IC ₅₀=257 nM), TRAM-34 (IC₅₀=55 nM), and charybdotoxin (CTX, IC₅₀=10.3 nM). RT-PCR analysis demonstrated the expression of mRNA encoding the IK_Ca 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 IK_Ca 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 μM, for 5 days) virtually suppressed the IK _Ca 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 IK_Ca 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 down-regulation of the IK_Ca current was not accompanied by changes in the ERK activity, as assessed by immunoblot analysis. Semiquantitative RT-PCR analysis demonstrated a ∼35% reduction of the IK_Ca channel mRNA resulting from ERK inhibition and a ∼50% reduction with time in culture.