Synthesis of [³H]2-Amino-4-phosphonobutyric acid and characterization of its binding to rat brain membranes: a selective ligand for the chloride/calcium-dependent class of l-glutamate binding sites

[³H]2-amino-4-phosphonobutyric acid was synthesized by the conjugate addition of 1-lithio-2-trimethylsilyethyne to diethyl ethynylphosphate followed by catalytic tritiation and hydrolysis. Radiolabelled 2-amino-4-phosphonobutyric acid binds to a distinct class of l-glutamate binding sites and does not exhibit appreciable binding to sites not displaced by l-glutamate. The binding affinity (K_d = 5.1 ± 0.4 μM) and pharmacological profile correspond to those values obtained from physiological studies of 2-amino-4-phosphonobutyric acid inhibition of synaptic transmission, and to those values obtained in [³H]l-glutamate binding assays. [³H]2-amino-4-phosphonobutyric acid does not exhibit significant binding to the Cl^-/Ca²⁺-independent l-glutamate binding site(s), nor to the Na⁺-dependent l-glutamate binding site (up to 50 mM Na⁺). These data provide further evidence that the physiological action of 2-amino-4-phosphonobutyric acid is mediated by the previously described Cl^-/Ca²⁺-dependent l glutamate binding sites, and provides an assay system which is optimal for the study of these sites.