Figure 4.

NSC culture conditions induce upregulation of K_Ca3.1 K⁺ currents in the plasma membrane of SMA-540 mouse glioma cells (A,B). Ionic composition with channel modulators of pipette and bath solution and applied voltage pulse protocol (A) used to record macroscopic on-cell (cell-attached) currents from an NSC-grown (10–14 d) SMA-540 cell. On-cell current tracings depicted in (B) were obtained before (left), during bath application of the K_Ca3.1 K⁺ channel opener 1-EBIO (200 µM) alone (middle) or in combination with the K_Ca3.1 inhibitor TRAM-34 (1 µM, right). For better readability, only current tracings elicited by voltage square pulses to −75, −50, −25, 0, +25, +50, and +75 mV are shown; red line in (B) indicates 0 pA (C). Dependence of macroscopic on-cell currents on clamp voltage in NSC- (left) and DMEM-grown (right) SMA-540 cells recorded before (control, open black circles), during bath application of 1-EBIO (closed red triangles) and co-application of 1-EBIO and Tram-34 (open red triangles). Data are means ± SE, n = 9; the inserts in the lower right corner of the plots show light micrographs of a SMA-540 spheroid (left) and a SMA-540 monolayer (right) during patch-clamp recording (D). Paired conductances as calculated for the inward currents from the data summarized in (C) in NSC- (left) and DMEM-grown (right) SMA-540 cells recorded successively with 1-EBIO and 1-EBIO/TRAM-34 in the bath solution. Given are individual values determined for that range of clamp voltage indicated in (C) by blue lines (E). Two-by-two contingency plots showing the number of NSC- and DMEM-grown SMA-540 cells with an on-cell inward conductance in 1-EBIO-containing bath solution (G_1-EBIO) of above (closed columns) and below (open columns) 0.1 nS (left) as well as with a TRAM-34-induced inward conductance decline (ΔG_TRAM-34) of above (closed columns) and below (open columns) 0.13 nS (right). Data are from (D), indicated p values refer to the difference between the culture conditions and were calculated with chi square test (F). Paired reversal potentials (V_revs, individual values) as given for the data summarized in (C) in NSC- (left) and DMEM-grown (right) SMA-540 cells recorded successively with 1-EBIO and 1-EBIO/TRAM-34 in the bath solution (G). Two-by-two contingency plots showing the number of NSC- and DMEM-grown cells with a V_rev in 1-EBIO-containig bath solution (V_{rev 1-EBIO)} of above (closed columns) and below (open columns) +40 mV (left) as well as with a TRAM-34-induced drop in V_rev (ΔV_{rev TRAM-34}) by above (closed columns) and below (open columns) 10 mV (right).