Fig. 4.

CNQX dissociation kinetics reveal that two molecules of antagonist bind to the AMPA receptor. A, An outside-out patch was stepped from a saturating concentration of CNQX (20 μm) into a saturating concentration of kainate (1 mm). CNQX dissociation is the rate-limiting process for activation of this response. The early part of the response (box) is sigmoidal, which suggests a multistep process.B, An enlargement of the boxed area inA shows the response (dots) fitted with three different kinetic models incorporating one, two, or threeantagonist binding sites (lines). All three models gave a good fit late in the response when CNQX unbinding was complete (data not shown). However, the fits diverged in the rising phase. The optimally fitted transients are shown as dashed lines for the one- and three-antagonist site models and asolid line for the two-antagonist site model.C, Responses after a step from CNQX into kainate were recorded in the absence or presence of 100 μm CTZ and fitted with the three kinetic models. As shown in the SSE ratios in thehistogram, the two-antagonist-site model gave the best fit to the activation phase of the response in the absence (−CTZ; n = 7) and in the presence (+CTZ; n = 12) of CTZ.