Permeability of P2X₇R to NMDG⁺ depends on duration of agonist application. (A) Pattern of 100 μM BzATP-induced current and reversal potential in cells bathed in 10% Na⁺ and 90% NMDG⁺-containing KR buffer during the initial agonist application. (A, left) Current recording during a 40-s agonist application at −60 mV holding potential. (Middle) Time course of agonist-induced currents in cells under the ramp protocol. (Right) Positive shift in reversal potential observed during the initial 40-s application of BzATP; 0.485-s voltage ramps were delivered twice per second. (B) Patterns of BzATP-induced currents (left and middle) and shift in the reversal potential (right) in cells bathed in KR buffer, in which Na⁺ was completely substituted with NMDG⁺. (C) Time course of BzATP-induced P2X₇R current in cells bathed in NMDG⁺ buffer (left and middle) and the accompanied changes in the reversal potential (right). In this and all figures, only 15 out of 100 traces for the current voltage relationship with the equal time intervals are shown. Notice the lack of receptor deactivation in C compared with records shown in A (left) and B (left). (D) Increase in NMDG⁺ permeability occurs in cells bathed in physiological solution for agonist application of >∼10 s. Application of agonist was initially performed in cells bathed in KR buffer for 4 (left) and 40 s (middle) and was continued in NMDG⁺ medium (gray areas). Notice a difference in responses when replacement of KR buffer with NMDG⁺ buffer was performed during the phase I (left) and II (middle) of current growth. Right panel illustrates the mean ± SEM values of the peak currents reached after replacement of KR buffer with NMDG⁺ buffer in two time points. All experiments were performed in HEK293 cells.