[Ca²⁺]_i dependence of TRPM2 single-channel gating parameters is well described by the MWC model. (A–C) Steady-state (mean ± SEM) open probabilities (P_o; A), opening rates (k_CO; B), and closing rates (k_OC; C) of single TRPM2 channels at ≥4.4 µM Ca²⁺ are plotted (circles) as a function of [Ca²⁺]_i. In C, diamonds represent closing rates at ≤4.4 µM Ca²⁺ determined from macroscopic current decay time courses. Pipette [Ca²⁺] was either ∼4 µM (closed circles) or 1 mM (open circles and dotted lines). Solid black lines illustrate the fit of the data at low extracellular Ca²⁺ (closed circles) by the scheme in D. (D) MWC model with four Ca²⁺-binding sites (1 site/subunit). Squares represent closed and circles represent open channel subunits; filled black symbols represent subunits with Ca²⁺ bound. Solid and dotted arrow indicate the pathways of channel closure upon rapid removal of intracellular Ca²⁺ in the absence and presence, respectively, of extracellular Ca²⁺.