cGMP activation of wild-type CNGA1 channels. (A and B) Macroscopic current traces recorded in symmetric 130 mM Na⁺ from inside-out patches containing CNGA1 channels in the presence of 2 mM (A) or 30 µM (B) of intracellular cGMP. Currents were elicited by stepping from the −80-mV holding potential to voltages between −200 and 200 mV in 10-mV increments. Only traces every 20 mV are shown for clarity. Dotted line indicates zero current level. (C) I-V curves (mean ± SEM; n = 3–6) in the presence of 0.01 mM (filled inverted triangles), 0.03 mM (open triangles), 0.1 mM (filled circles), and 2 mM (open squares) cGMP, each normalized to the current at 200 mV. (D) Corresponding normalized G-V curves (mean ± SEM; n = 3–6). (E) Fraction of maximal current (I/I_max; mean ± SEM; n = 7) plotted against cGMP concentration for −200 mV (open circles) and 200 mV (filled circles). Black dashed curves are Hill equation fits with a common Hill coefficient (n) yielding EC₅₀ = 66 ± 2 µM at −200 mV, EC₅₀ = 53 ± 1 µM at 200 mV, and n = 1.48 ± 0.03. Red dashed curves are fits of Eq. 1 to both datasets simultaneously, with a single adjustable K_L and all other parameters fixed (n = 1.48; K₁ = 0.62, K₂ = 1.04, and Z_K2 = 0.25), which yield K_L = 96 ± 2 µM at −200 mV and K_L = 182 ± 5 µM at 200 mV.