Urinary and Brain β -carboline-3-carboxylates as Potent Inhibitors of Brain Benzodiazepine Receptors

Benzodiazepines probably exert their anxiolytic, hypnotic, and anticonvulsant effects by interacting with brain-specific high-affinity benzodiazepine receptors. In searching for possible endogenous ligands for these receptors, we have purified a compound 107-fold from human urine by extractions, treatment with hot ethanol, and column chromatography. The compound was identified as β -carboline-3-carboxylic acid ethyl ester (IIc) by mass spectrometry, NMR spectrometry, and synthesis; IIc was also isolated from brain tissues (20 ng/g) by similar procedures. Very small concentrations of IIc displaced [3H]diazepam completely from specific cerebral receptors, but not from liver and kidney binding sites; the concentration causing 50% inhibition of specific [3H]diazepam binding (IC50) was 4-7 nM compared to ca. 5 nM for the potent benzodiazepine lorazepam. Specific binding sites for quinuclidinyl benzilate, naloxone, spiroperidol, serotonin, muscimol, and WB 4101 were not affected by IIc. In contrast to benzodiazepines, IIc exhibits ``mixed type'' competitive inhibition of forebrain benzodiazepine receptors (negative cooperativity). We surmise that an endogenous ligand for benzodiazepine receptors may be a derivative of β -carboline-3-carboxylic acid.