Characterization of the pharmacology and regional distribution of (S)‐[3H]‐5‐fluorowillardiine binding in rat brain

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Abstract
1 This study examined the binding of the new radioligand (S)-[3H]-5-fluorowillardiine to rat brain synaptic membranes. Specific binding represented greater than 80% of the total binding and was increased by 10% in the presence of 100 mM potassium thiocyanate (KSCN). 2 In the absence of KSCN, (S)-[3H]-5-fluorowillardiine identified two binding sites with KD1 = 22.5nM, Bmax1= 1.4 pmol mg-1 protein and KD2=1.5 μM, Bmax2= 10.8 pmol mg−1 protein. In the presence of 100 mM KSCN the affinities of both the binding sites were increased, yielding values of KD1 = 6.9nM and KD2 = 0.4 μm KSCN was without effect on the Bmax values. 3 (S)-[3H]-5-fluorowillardiine binding was displaced by non-NMDA receptor ligands with the rank order of potency: 2,3-dihydroxy-6-nitro-7-sulphamoyl-benzo(F)quinoxaline (NBQX) > domoate > (S)-α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) ≅ L-glutamate > 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) > kainate >> (R)-5-fluorowillardiine. In contrast, both N-methyl-D-aspartate (NMDA) and the metabotropic glutamate receptor agonist, (lS,3R)-l-aminocyclopentane-l,3-dicarboxylic acid (ACPD) were inactive. 4 By use of quantitative autoradiography the regional distribution of (S)-[3H]-5-fluorowillardiine binding in rat brain was assessed. The highest levels of binding were in the dentate gyrus and the CA1 region of the hippocampus. Lower levels of binding were detected in the cerebral cortex, olfactory system, lateral septum, caudate putamen and nucleus accumbens. 5 We conclude that the pharmacological profile and regional distribution of (S)-[3H]-5-fluorowillardiine binding is consistent with its specific interaction with AMPA receptors. The advantages of high percentage specific binding and recognition of the two binding sites, in the absence of KSCN, suggest that (S)-[3H]-5-fluorowillardiine may be the ligand of choice for the future study of AMPA receptors.