Resolution, configurational assignment, and enantiopharmacology of 2-amino-3-[3-hydroxy-5-(2-methyl-2H-tetrazol-5-yl)isoxazol-4-yl]propionic acid, a potent GluR3-and GluR4-preferring AMPA receptor agonist

We have previously shown that (RS)-2-amino-3-[3-hydroxy-5-(2-methyl-2H-tetr azol-5-yl) isoxazol-4-yl] propionic acid (2-Me-Tet-AMPA) is a selective ago nist at (RS) -2-amino-3- (3-hydroxy-5-methylisoxazol-4-yl) propionic acid ( AM PA) receptors, markedly more potent than AMPA itself, whereas the isomer ic compound 1-Me-Tet-AMPA is essentially inactive. We here report the enant iopharmacology of 2-Me-Tet-AMPA in radioligand binding and cortical wedge e lectrophysiological assay systems, and using cloned AMPA (GluR1-4) and kain ic acid (KA) (GluR5, 6, and KA2) receptor subtypes expressed in Xenopus ooc ytes. 2-Me-Tet-AMPA was resolved using preparative chiral HPLC. Zwitterion (-)-2-Me-Tet-AMPA was assigned the (R)-configuration based on an X-ray crys tallographic analysis supported by the elution order of (-)- and (+)-2-Me-T et-AMPA using four different chiral HPLC columns and by circular dichroism spectra. None of the compounds tested showed detectable affinity for N-meth yl-D-aspartic acid (NMDA) receptor sites, and (R)-2-Me-Tet-AMPA was essenti ally inactive in all of the test systems used. Whereas (S)-2-Me-Tet-AMPA sh owed low affinity (IC50 = 11 muM) in the [H-3]KA binding assay, it was sign ificantly more potent (IC50 = 0.009 muM) than AMPA (IC50 = 0.039 muM) in th e [H-3]AMPA binding assay, and in agreement with these findings, (S)-2-Me-T et-AMPA (EC50 = 0.11 muM) was markedly more potent than AMPA (EC50 = 3.5 mu M) in the electrophysiological cortical wedge model. In contrast to AMPA, w hich showed comparable potencies (EC50 = 1.3-3.5 muM) at receptors formed b y the AMPA receptor subunits (GluR1-4) in Xenopus oocytes, more potent effe cts and a substantially higher degree of subunit selectivity were observed for (S)-2-Me-Tet-AMPA: GluR1o (EC50 = 0.16 muM), GluR1o/GluR2i (EC50 = 0.12 muM), GluR3o (EC50 = 0.014 muM) and GluR4o (EC50 = 0.009 muM). At the KA-p referring receptors GluR5 and GluR6/KA2, (S)-2-Me-Tet-AMPA showed much weak er agonist effects (EC50 = 8.1 and 15.3 muM, respectively). It is concluded that (S)2-Me-Tet-AMPA is a subunit-selective and highly potent AMPA recept or agonist and a potentially useful tool for studies of physiological AMPA receptor subtypes. (C) 2000 Wiley-Liss, Inc.