We report the preparation of the four possible stereoisomers of 4-methylglutamic acid via diastereoselective alkylation or chemical resolution. Radioligand binding studies showed these single isomers exhibit variable selectivity for kainate (KA) receptors depending on their stereochemical configuration. Among them, the (2S,4R)-isomer has exceptional selectivity for the KA receptor with an IC50 for inhibition of [3H]KA binding of ~35 nM, comparable to kainic acid itself (~11 nM). The threo isomers (2S,4S and 2R,4R) are moderately potent inhibitors of [3H]KA binding (IC50s of 0.31 and 0.33 μM, respectively) with marked selectivity for KA over AMPA and NMDA receptors (IC50s >10 μM). The (2R,4S)-isomer is more than 20-fold less potent (IC50 ~820 nM) but maintains selectivity for KA. Introduction of a methyl group at the 4-position of glutamic acid enhances selectivity for the KA receptor subtype. The (2S,4R)-isomer is the most selective KA receptor ligand described to date without significant cross reactivity with AMPA or NMDA receptors. While additional electrophysiological and functional studies are required to determine whether these compounds possess agonist or antagonist properties, they will undoubtedly be valuable for elucidating the physiological and pharmacological functions of the KA receptor subtype of glutamate receptors.