To determine the PIP2 sensitivity, we examined the effect of applying different concentrations of diC8-PIP2 to GIRK2-containing liposomes (Fig. 2a). Note the progressively faster rate of quenching with higher diC8-PIP2 concentrations. Plotting the rate of K+ flux (1/tau) as a function of the diC8-PIP2 concentration reveals a concentration-dependent increase in the rate of quenching. The best-fit with the Hill equation indicates an apparent EC50 of 25.1 ± 3.3 μM and a Hill coefficient of 1.7 ± 0.1 (n = 10) (Fig. 2b). The EC50 with diC8-PIP2 is higher than for that for GIRK2 reconstituted in bilayers in the presence of Gβγ (~15 μM)34 and lower than that for GIRK1/GIRK4 channels expressed heterologously (~ 45 μM)39.
