Characterization of quisqualate-type l-glutamate receptors in the retina

In the vertebrate retina excitatory transmission seems to be mediated mainly by excitatory amino acids; glutamate and/or aspartate are the most viable candidates to subserve this function. Postsynaptic receptors for N-methyl- d-aspartate (NMDA), kainate (KA), quisqualate (QA) and 2-amino-4-phosphonobutyric acid have been electrophysiologically identified. In this work we have tried to identify and characterize QA receptors through the binding of the most specific analogue available for this receptor: [ 3H]α-amino-3-hydroxy-5-methylisoxazole-4-propionic acid ([ 3H]AMPA). AMPA binding to retinal membranes was sodium- and temperature-independent, with optimum pH at 6–7. Ligand-receptor interaction was reversible and saturable. Pharmacologically, glutamate analogues were more active displacers than NMDA analogues: AMPA> (RS)-3-hydroxy-4,5,6,7-tetrahydro-isoxazolo-(5,4-C)-pyridine-7-car☐ylic acid= l-Glu=QA ; with IC 50 in the low μM range. Glutamic acid diethylester was uneffective while KA and cis-2,3-piperidine dicar☐ylate were potent inhibitors of binding. Binding was stereospecific, l-isomers being more effective displacers than d-forms. Subcellular distribution showed binding concentrated in the inner plexiform layer (IPL), but also present in the outer plexi-form layer (OPL). Kinetics of [ 3H]AMPA binding showed a high affinity k B = 1–2 μM in membranes from complete retina, IPL and OPL, with binding sites concentrated in P 2 ( B max = 16.2pmol/mgprotein ). Our results provide biochemical evidence for the presence and distribution of physiologically relevant QA receptors in the chick retina which is an agreement with previous physiological findings.