We describe a stopped-flow method to study α-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 like ouabain (1 mM) and 5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine maleate (dizocilpine, 50 μM) enhanced Na+ ion translocation under low-[Na+] and physiological conditions, respectively. Dependence of AMPA-kainate receptor kinetics on [Glu] was described in a model of channel activation by faster and slower desensitizing receptors. The model accounted for almost all of the Na+ ion flux activity in the 30 μM-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, α, for the faster desensitizing 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 described by (1) a fractal model of short-lived AMPA-kainate receptor channel with many closely spaced states (fractal dimension ~1.8) and (2) a model of long-lived AMPA-kainate receptor channel with two discrete states. Copyright (C) 1999 Elsevier Science Ltd.
- Fractal kinetics
- Hippocampal AMPA-kainate receptors
- Stopped-flow method
- Transmembrane NA ion flux
ASJC Scopus subject areas
- Cellular and Molecular Neuroscience
- Cell Biology