The action of two cyclobutane derivatives of L-glutamic acid on the high affinity uptake of L-glutamic acid was investigated using a preparation of synaptosomes from rat cerebral cortex. cis-1-Aminocyclobutane-1,3-dicarboxylic acid (also known as trans-2,4-methanoglutamic acid) potently inhibited L-glutamic acid uptake (IC50 30 microM), whereas trans-1-aminocyclobutane-1,3-dicarboxylic acid (also known as cis-2,4-methanoglutamic acid), a potent N-methyl-D-aspartate (NMDA) agonist, was inactive. Analysis of the kinetics of L-glutamic acid uptake in the presence and absence of cis-1-aminocyclobutane-1,3-dicarboxylic acid (CACB) suggests that it may act as a competitive inhibitor (Ki 8 microM). CACB may be substrate for the L-glutamic acid high-affinity uptake carrier since preincubation of CACB with the synaptosomal preparation increased its potency in inhibiting L-glutamic acid uptake. The conformationally restricted structure of CACB may be indicative of the conformations of L-glutamic acid that interact with the high affinity uptake carrier.