Figure 4.

Influence of the combined cGMP-analogues treatment on retinal CNG channels. (A,B) Relative current amplitudes of rod and cone CNG channels measured under steady-state conditions, in the presence of either cGMP + inhibitor (Rp-8-Br-PET-cGMPS, 50 µM, green) or cGMP + inhibitor (50 µM) + activator (8-pCPT-cGMP, 0.2 µM, 0.1 µM, and 0.05 µM, red). The measured currents were related to the activation triggered by 5 µM and 100 µM cGMP for cone and rod channels, respectively. (C,D) Concentration–activation relationships for rod (left) and cone (right) CNG channels, measured at −35 mV, in the presence of cGMP (black) and cGMP + inhibitor (50 µM) + activator (0.1 µM) (red). The experimental data points, representing means of several measurements, were fitted with Equation (1) (for EC₅₀, H, and n, see Table S1). cGMP levels < 10 µM were considered physiological conditions, while cGMP levels > 10 µM represent “RP-like”-conditions (gray arrows). (E,F) Superimposition of representative activation- and deactivation- time courses following a concentration jump from 0 µM cGMP to 100 µM cGMP + 50 µM inhibitor + 0.1 µM activator for rod CNG channels (left) and to 5 µM cGMP + 50 µM inhibitor + 0.1 µM activator for cone (right). The currents triggered by cGMP (black) and by cGMP-analogues co-application (red) were normalized to the level observed in (C,D). The respective time courses were quantified by a mono-exponential fit yielding the activation and deactivation time constants (τ_act and τ_deact, respectively, Equation (3), green curves). The inset in (E) shows the magnified effect of the cGMP-analogues combination on the rod CNG-channel activity (red). (G) Activation and deactivation time constants (τ_act, τ_deact) for rod and cone CNG channels (n = 4–9). Statistical significance (****) was estimated with the Student’s t-test. “ns” means not statistically significant. The gray symbols represent individual measurements.