IN-VITRO BINDING-PROPERTIES IN RAT-BRAIN OF [H-3]RO-25-6981, A POTENTAND SELECTIVE ANTAGONIST OF NMDA RECEPTORS CONTAINING NR2B SUBUNITS

The in vitro binding of a new subtype-selective NMDA receptor antagoni st, [H-3]Ro 25-6981, to rat brain membranes and sections was character ized. The compound bound to a single site on the membranes with a K-D of 3 nM and a B-max of 1.6 pmol/mg of protein. Specific binding, defin ed with a new NR2B-specific antagonist, Ro 04-5595 methoxy-7-hydroxy-1 ,2,3,4-tetrahydroisoquinoline], at 10 mu M, was fully inhibited by sev eral compounds with the following rank order of affinities-Ro 25-6981 > CP-101,606 > Ro 04-5595 = ifenprodil much greater than eliprodil > h aloperidol > spermine > spermidine > MgCl2 > CaCl2-and partially inhib ited by competitive glutamate recognition site antagonists. A high den sity of binding sites was detected, radioautographically, in several l ayers of the cerebral cortex, in the hippocampus, dentate gyrus, tuber culum olfactorium, caudate putamen, medium densities in the globus pal lidus, thalamus, spinal cord dorsal horn, and motoneurons, whereas the cerebellum, pens, and medulla were, with a few exceptions, e.g., locu s coeruleus, poorly labeled. Overall, the distribution of [H-3] RO 25- 6981 binding sites correlated well with that of NR2B (but not NR2A) tr anscripts, revealed by in situ hybridization histochemistry. The high affinity of [H-3]Ro 25-6981 for NR2B-containing receptors renders this compound the ligand of choice to study the regulation of NR2B-contain ing receptor expression.