Synthesis of the Nanomolar Photoaffinity GABAB Receptor Ligand CGP 71872 Reveals Diversity in the Tissue Distribution of GABAB Receptor Forms Original Research Article

A radioiodinated probe, [125I]-CGP 71872, containing an azido group that can be photoactivated, was synthesized and used to characterize GABAB receptors. Photoaffinity labeling experiments using crude membranes prepared from rat brain revealed two predominant ligand binding species at ∼130 and ∼100 kDa believed to represent the long (GABABR1a) and short (GABABR1b) forms of the receptor. Indeed, these ligand binding proteins were immunoprecipitated using a GABAB receptor-specific antibody confirming the receptor specificity of the photoaffinity probe. Most convincingly, [125I]-CGP 71872 binding was competitively inhibited in a dose-dependent manner by cold CGP 71872, GABA, saclofen, (−)-baclofen, (+)-baclofen and (l)-glutamic acid with a rank order and stereospecificity characteristic of the GABAB receptor. Photoaffinity labeling experiments revealed that the recombinant GABABR2 receptor does not bind [125I]-CGP 71872, providing surprising and direct evidence that CGP 71872 is a GABABR1 selective antagonist. Photoaffinity labeling experiments using rat tissues showed that both GABABR1a and GABABR1b are co-expressed in the brain, spinal cord, stomach and testis, but only the short GABABR1b receptor form was detected in kidney and liver whereas the long GABABR1a form was selectively expressed in the adrenal gland, pituitary, spleen and prostate. We report herein the synthesis and biochemical characterization of the nanomolar affinity [125I]-CGP 71872 and CGP 71872 GABABR1 ligands, and differential tissue expression of the long GABABR1a and short GABABR1b receptor forms in rat and dog.