TFP, a CaM inhibitor, inhibits Ano1 in a voltage-dependent manner in whole-cell and excised inside-out patch recordings. (A) Effect of extracellular 10 µM TFP on whole-cell Ano1 current activated by 20 µM Ca²⁺. TFP inhibits the current, and the effect is partially reversed upon washout. (B) I-V relationship of Ano1 before and 1 min after extracellular application of TFP. (C) The mean normalized currents at 100 mV after 1-min exposure to TFP (n = 4). (D) Application of 10 µM TFP to the cytosolic side of an inside-out excised patch has no effect on Ano1 current. (E) A CaM-binding peptide, 10 µM CaMKII (281–309), applied to the cytosolic side of an inside-out patch causes no effect on Ano1 current in inside-out patch. (F) Quantification of the effect of TFP and CaMKII(281–309) on Ano1 current. The current decrease was quantified before and during exposure to compounds as I_1min/I_0min as described in Fig. 2 (n = 3). Error bars are ±SEM.