Autoradiographic characterization of the non-N-methyl-D-aspartate binding sites in human cerebellum using the antagonist [3H]6-cyano-7-nitroquinoxaline-2,3-dione.

Using quantitative autoradiography, we have characterized the binding properties of the non-N-methyl-D-aspartate (NMDA) glutamate receptor antagonist [3H]6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) in adult human cerebellum. Saturation experiments revealed [3H]CNQX binding to a single class of sites with similar affinity in the molecular and granule cell layer (Kd = 89.0 +/- 6.4 and 83.3 +/- 9.9 nM, respectively). The maximum number of [3H]CNQX binding sites was much higher in the molecular compared to the granule cell layer (Bmax = 16.2 +/- 1.1 and 2.8 +/- 0.5 pmol/mg protein, respectively). Inhibition experiments were performed in order to examine the pharmacological profile of [3H]CNQX binding in the molecular layer. [3H]CNQX labeled sites with high affinity for both non-NMDA agonists, (RS)-alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) and kainate. Dose-response curves for inhibition of [3H]CNQX by AMPA and kainate were biphasic. The potency of AMPA for displacement of [3H]CNQX binding (Ki: 2.8 +/- 0.8 nM and 12.5 +/- 0.8 microM) was 4- to 6-fold greater than the corresponding potency of kainate (Ki: 18.1 +/- 5.7 nM and 48.7 +/- 9.3 microM). In conclusion, the pharmacological analysis of [3H]CNQX binding in the human cerebellar molecular layer reflects the existence of multiple binding sites of the non-NMDA receptor that have different affinities for both AMPA and kainate.