The glutamate uptake system in presynaptic vesicles: further characterization of structural requirements for inhibitors and substrates.

Noncyclic fluorine-substituted and cyclic analogs of glutamic acid were tested for their ability to inhibit glutamate uptake in isolated bovine presynaptic vesicles, in order to assess the specific structural requirements of the glutamate translocation system in the vesicle membrane. Cyclic analogs that permitted close interaction between the positive and negative charges of the glutamate molecule were effective inhibitors; maximum inhibitory potency was observed with L-trans-1-aminocyclopentane-1,3-dicarboxylic acid (L-t-ACPD), while D-t-ACPD was less active. Analogs with a larger or smaller ring (as in trans-1-aminocyclohexane-1,3-dicarboxylic acid or trans-1-aminocyclobutane-1,3-dicarboxylic acid) were also inhibitory, but somewhat less so. trans-ACPD was also taken up by the vesicles with a time course and ATP dependence similar to uptake of glutamate, and this uptake was inhibited by glutamate. The K(m) value for t-ACPD uptake was similar to its K(i) for inhibition of glutamate uptake, while its rate of uptake was lower than that of glutamate. Fluorine-substituted noncyclic analogs with substitutions at the 4-carbon were less effective than glutamic acid itself, although 4,4-difluoroglutamic acid was equal in activity to the unsubstituted compound. Inhibition by these derivatives appeared to be competitive in nature, and they probably were also transported by the vesicle uptake system.