A LIGAND-BINDING STUDY OF THE INTERACTIONS OF GUANINE-NUCLEOTIDES WITH NON-NMDA RECEPTORS

The interactions of guanine nucleotides, and particularly GTP, with th e pha-amino-3-hydroxy-5-methylisoxazole-4-propionate (AMPA) and [H-3]- kainate (KA) binding sites present on brain membranes was studied, usi ng the ligand binding methodology and Scatchard analysis. in order to establish the competitive/non competitive nature of the interaction an d determine whether guanine nucleotides, KA and AMPA share common bind ing sites. GTP was found to block [H-3]-AMPA and [H-3]-KA binding to r at cortical membranes with IC50 values of 0.4 mM and 1 mM respectively and the [H-3] KA-binding to chick cerebellar membranes with a IC50 va lue of 20 muM. Scatchard analysis of [H-3]-KA binding performed in the absence or presence of 1 mM GTP or 0.25 mM AMPA reveals that the high affinity [H-3]-KA binding component is not affected by GTP but blocke d in a non competitive fashion by AMPA while the low affinity [H-3]-KA binding component is not affected by AMPA but blocked by GTP. Scatcha rd analysis of [H-3]-KA binding to chick cerebellar membranes performe d in the absence or presence of 33 muM GTP reveals a single binding si te blocked in a competitive fashion by GTP. Scatchard analysis of [H-3 ]-AMPA binding performed in the absence or presence of 0.5 mM GTP or 3 0 muM KA reveals that the high affinity [H-3]-AMPA binding component i s affected in a non competitive fashion by both GTP and KA while the l ow affinity [H-3] AMPA binding component is affected in a competitive fashion by both GTP and KA. These results suggest that the proteins ca rrying the low affinity [H-3]-AMPA binding site in rat cortical membra nes and the [H-3]-KA binding sites in chick cerebellar membranes harbo ur in their extracellular domain a single binding site for the radioac tive ligand and GTP. The role of GTP as a brain endogenous glutamaterg ic ligand is discussed.