t-DCA slows down the ATP-mediated activation of ensemble P2X4 currents and facilitates the spontaneous inactivation of the channel. (A) Representative continuous current trace recorded at a holding potential of −70 mV from an outside-out patch obtained from a control oocyte. Application of ATP (100 µM) is indicated by an open bar. (B and C) Representative continuous current traces recorded at a holding potential of −70 mV from outside-out patches obtained from an oocyte expressing P2X4. Application of ATP (100 µM) and t-DCA (250 µM) is indicated by open and filled bars, respectively. Linear fit of initial phase of ATP-activated currents and exponential decay fit of the subsequent spontaneous current decrease were calculated as described in Materials and methods and delineated in red. To quantify the responses, the current integral (Q, pC), the slope of the initial P2X4-mediated ensemble inward current increase (sact, pA/s), and the time constant of the spontaneous P2X4 ensemble current inactivation (τinact, s) were determined for each recording. (D–F) Summary of results from similar experiments as shown in B and C. Data are means ± SEM and individual data points (n = 9, N = 5). ***, *, Significantly different, P < 0.001 and P < 0.05, respectively (Student’s t test).