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

Although membrane sites from brain, labelled with [3H]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 degrees C) both led to decreases in the apparent specific binding of [3H]glutamate to membranes from the brain of the rat in the presence of chloride. Furthermore, only 15% of the [3H]glutamate, bound at 37 degrees C, was dissociable when the membranes were then cooled to 4 degrees C. The binding of [3H]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 5 mM glutamate increased the apparent binding of [3H]glutamate and abolished the stimulatory effect of Phe-Glu. These findings are consistent with the hypothesis that chloride-dependent association of [3H]glutamate with membranes from brain reflects, in part, a sequestration process, which may be driven by glutamate exchange.