Kinetically distinguishable AMPA receptors in rat hippocampus are associated with the loss of glutamate-sensitive conformational transitions

We describe a stopped-flow method to study alpha-amino-7-hydroxy-5-methyl-4 -isoxazole propionate (AMPA)-kainate receptor-mediated Na+ ion flux through native membranes. Resealed plasmalemma vesicles and nerve endings from the rat hippocampus were mixed rapidly with a membrane impermeant form of the fluorescence indicator, sodium binding benzofurane oxazole and the changes in fluorescence intensity in response to various [Glu] on the time scale of 0.04 ms-10 s were monitored at a sampling rate of 6.55 kHz. Inhibitors lik e ouabain (1 mM) and 5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5, 1 0-imine maleate (dizocilpine, 50 mu M) enhanced Na+ ion translocation under low-[Na+] and physiological conditions, respectively. Dependence of AMPA-k ainate receptor kinetics on [Glu] was described in a model of channel activ ation by faster and slower desensitizing receptors. The model accounted for almost all of the Na+ ion flux activity in the 30 mu M-10 mM range of [Glu ]. We found that the values of the initial rate constant for Na+ ion influx , J(A), and rate constant for desensitization, alpha, for the faster desens itizing receptor were dependent on data sampling rate, whereas the initial rate constant for Na+ ion flux through the slower desensitizing receptor, J (B), varied much less with the sampling rate. These phenomena can be descri bed by (1) a fractal model of short-lived AMPA-kainate receptor channel wit h many closely spaced states (fractal dimension similar to 1.8) and (2) a m odel of long-lived AMPA-kainate receptor channel with two discrete states. (C) 1999 Elsevier Science Ltd. All rights reserved.