Characteristics of chloride-dependent incorporation of glutamate into brain membranes argue against a receptor binding site

Abstract Although membrane sites from brain, labelled with [ 3 H]glutamate (Glu) under sodium-free conditions, are thought to represent excitatory receptors, certain anomalous characteristics of the kinetics of apparent binding raised the question of whether transport might contribute to this process, prompting a closer examination of it. Hyperosmolar media and low incubation temperatures (4°C) both led to decreases in the apparent specific binding of [ 3 H]glutamate to membranes from the brain of the rat in the presence of chloride. Furthermore, only 15% of the [ 3 H]glutamate, bound at 37°C, was dissociable when the membranes were then cooled to 4°C. The binding of [ 3 H]glutamate was increased in the presence of certain dipeptides such as l -phenylalanyl- l -glutamate (Phe-Glu); and the binding augmented by the presence of Phe-Glu, was also sensitive to temperature and osmolarity of the incubation buffer. Sonication of membranes in 5mM glutamate increased the apparent binding of [ 3 H]glutamate and abolished the stimulatory effect of Phe-Glu. These findings are consistent with the hypothesis that chloride-dependent association of [ 3 H]glutamate with membranes from brain reflects, in part, a sequestration process, which may be driven by glutamate exchange.