Synthesis of the nanomolar photoaffinity GABA(B) receptor ligand CGP 71872reveals diversity in the tissue distribution of GABA(B) receptor forms

A radioiodinated probe, [I-125]-CGP 71872, containing an azido group that c an be photoactivated, was synthesized and used to characterize GABA(B) rece ptors. Photoaffinity labeling experiments using crude membranes prepared fr om rat brain revealed two predominant ligand binding species at similar to 130 and similar to 100 kDa believed to represent the long (GABA(B)R1a) and short (GABA(B)R1b) forms of the receptor. Indeed, these ligand binding prot eins were immunoprecipitated using a GABA(B) receptor-specific antibody con firming the receptor specificity of the photoaffinity probe. Most convincin gly, [I-125]-CGP 71872 binding was competitively inhibited in a dose-depend ent manner by cold CGP 71872, GABA, saclofen, (-)-baclofen, (+)-baclofen an d (L)-glutamic acid with a rank order and stereospecificity characteristic of the GABA(B) receptor. Photoaffinity labeling experiments revealed that t he recombinant GABA(B)R2 receptor does not bind; [I-125]-CGP 71872, providi ng surprising and direct evidence that CGP 71872 is a GABA(B)R1 selective a ntagonist. Photoaffinity labeling experiments using rat tissues showed that both GABA(B)R1a and GABA(B)R1b are co-expressed in the brain, spinal cord, stomach and testis, but only the short GABA(B)R1b receptor form was detect ed in kidney and liver whereas the long GABA(B)R1a form was selectively exp ressed in the adrenal gland, pituitary, spleen and prostate. We report here in the synthesis and biochemical characterization of the nanomolar affinity [I-125]-CGP 71872 and CGP 71872 GABA(B)R1 ligands, and differential tissue expression of the long GABA(B)R1a and short GABA(B)R1b receptor forms in r at and dog. (C) 1999 Elsevier Science Ltd. All rights reserved.