FIGURE 6.

Activation by internal free Ca²⁺ of K_Ca3.1 recorded in standard and fluoride-free internal solution. (A) CHO cells expressing K_Ca3.1 were captured with almost 50% success rate for R_Seal ≥ 1 GΩ with standard and in fluoride-free internal solution (left). If a QC cutoff of R_Seal ≥ 0.25 GΩ was used, the success rate was increased in both conditions (right). Similar success rates were also achieved with multi-hole (4X) chips (gray). (B) Raw current traces from example CHO cell expressing K_Ca3.1 recorded in both standard and fluoride-free internal solution, activated by 1 μM free internal Ca²⁺ and blocked by 1 mM Ba²⁺ (external). (C) Success rates after applying initial R_Seal (>0.25 GΩ) and current QC plot of I_Peak values at –120/60 mV for standard and fluoride-free solutions after internal application of 1 μM free Ca²⁺. (D) IT plot during application of internal free Ca²⁺ with 0.3, 1, and 3 μM free internal Ca²⁺ and subsequent block by external Ba²⁺ (1 mM) at –120/60 mV. (E) Current values at –120 or 60 mV in increasing concentrations of internal free Ca²⁺ were normalized to maximum peak amplitude and fit using a Hill equation which shows a slightly increased EC₅₀ in standard internal solutions for both voltages. Shown are mean ± S.E.M. (F) Current values at –120 or 60 mV in increasing concentrations of external Ba²⁺ were normalized to maximum peak amplitude and fit using a Hill equation which showed no difference in the IC₅₀ at –120 mV between standard internal (170 ± 8 μM, n = 2 chips) and fluoride-free internal (158 ± 15 μM, n = 2 chips) or 60 mV, although the IC₅₀ was higher at 60 vs. –120 mV for both groups (540 ± 105 μM, n = 2 chips in standard external and 728 ± 161 μM, n = 2 chips in fluoride-free internal solution). IC₅₀ values are given as mean of 2 chips ± S.D.