Fig. 4. Time course of the blockade of Na⁺ currents by Cn11 and determination of K_m. (A) Under control conditions, the current amplitude decreases steadily by approximately 10 % during the first 160 s, and this is taken as a normal rundown (open circles). Subsequent curves were obtained after superfusion with the toxin at various concentrations: filled squares, 50 nmol l^–1; open squares, 100 nmol l^–1; open diamonds, 200 nmol l^–1; filled diamonds, 500 nmol l^–1. Normalized current (I_m) was obtained as I/I_m. After 160 s, the preparation was superfused with the external solution. The current amplitude decreased in a concentration-dependent manner, with no apparent recovery. (B) The percentage of Na⁺ current blockade plotted against Cn11 concentration, 70 s after application of the toxin, corrected for 5 % rundown. The points were fitted to a Boltzman’s equation, giving a K_m of approximately 320 nmol l^–1.