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. 2005 Sep 23;89(6):3960–3975. doi: 10.1529/biophysj.105.060731

FIGURE 1.

FIGURE 1

Inhibitory activity of Lqh venom is state-dependent. (A) Representative traces from excised, inside-out macropatch experiments of WT-CFTR are shown before (top) and during application of either 0.1 mg/mL Lqh-pf venom (middle) or 200 μM DPC (bottom); Vm = −80 mV. In each case either Lqh-pf venom or DPC was applied to closed channels for ∼30 s before reopening by MgATP. (B) Representative traces from excised, inside-out macropatch experiments of Flag-cut-ΔR-CFTR are shown with (bottom) and without application of 0.1 mg/mL Lqh-pf venom (top); Vm = −80 mV. (C) Representative traces from excised, inside-out macropatch experiments of WT-CFTR are shown during application of either Lqh-pf venom (top) or DPC (bottom) to activated channels. Lqh-pf venom or DPC were applied to open channels for 30 s; Vm = −80 mV. Bath solution contained 1 mM MgATP, no MgATP, 1 mM MgATP plus either 0.1 mg/mL Lqh-pf venom or 200 μM DPC, or venom or DPC in the absence of ATP. WT-CFTR channels were phosphorylated before the beginning of the experiment. (D) Summary of the effects of either Lqh-pf venom (0.1 mg/mL) or DPC (200 μM) on WT-CFTR steady-state current in the presence of 1 mM MgATP. Mean steady-state current was determined by averaging the current over the final 15–20 s in each condition. (Circles represent individual experiments, some are shown to overlap. Triangles and error bars indicate mean ± SE of n = 5–14 observations for each condition. The asterisks indicate that Lqh venom activity is significantly different when applied to closed versus open CFTR (*p ≤ 0.001).) (E) Effect of application of either 0.1 mg/mL Lqh-pf venom or 200 μM DPC to WT-CFTR while activated by 1 mM MgATP. Arrow indicates the timing of the rapid solution switch from normal bath solution with 1 mM MgATP to bath solution containing MgATP and either venom or DPC. The membrane potential was set to 0 mV and then stepped to −80 mV for 150 ms and repeated once per second for 30 s to measure the on-rate of CFTR inhibition by either DPC or Lqh-pf venom. Mean fractional block ± SE (n = 14 with Lqh-pf venom, n = 5 with DPC) is plotted as a function of time.