Figure 3.

(A) Representative whole-cell currents mediated by GluA2Q_flip receptors expressed in HEK-293S cells in the absence (left) and presence (right) of BDZ-f, obtained by flow measurement. The concentrations of glutamate and BDZ-f are 3 mM and 2 µM, respectively. The inhibition ratio (A/A_I) for the pair is ~1.61. The whole-cell current was recorded at −60 mV, pH 7.4, and 22 °C. (B) Effect of BDZ-f on the whole-cell current amplitude of GluA2Q_flip receptors obtained from the flow measurement. K_I of 3.8 ± 0.4 µM was determined by using eq 6 for the closed-channel state (100 µM glutamate, ●); K̅_I of 5.4 ± 0.8 µM was obtained for the open-channel state (3 mM glutamate, ○). (C) The effect of BDZ-f on the channel desensitization rates for the closed-channel (lower, determined at 100 µM glutamate) and open-channel (upper, determined at 3 mM glutamate) states of GluA2Q_flip. (D) A representative whole-cell current mediated by GluA2Q_flip receptors at 100 µM of glutamate shows that the total current amplitude (A_tot) equals 425 pA with the amplitude of non-desensitizing phase (A_non-des) remaining at 61 pA (until glutamate was removed), and the desensitizing phase being 364 pA. The fraction of the non-desensitizing phase is 14%. (E) Effect of BDZ-f on the two components of the whole-cell current of the closed-channel state of GluA2Q_flip receptors. In the left panel, the amplitude of the desensitized phase is plotted against inhibitor concentration, and a K_I of 3.2 ± 0.3 µM was determined. In the right panel, a K_I of 6.9 ± 2.3 µM was determined for the non-desensitizing phase. (F) Effect of BDZ-f on the two components of the whole-cell current of the open-channel conformation of GluA2Q_flip. In the left panel, the amplitude of the desensitized phase is plotted against inhibitor concentration, and a K̅_I of 5.4 ± 0.3 µM was determined. We did not estimate an inhibition constant for the data on the right panel due to large experimental error.