The diazepam binding inhibitor’s modulation of the GABA-A receptor is subunit-dependent

First synthesized in the 1950s, benzodiazepines are widely prescribed drugs that exert their anxiolytic, sedative and anticonvulsant actions by binding to GABA-A receptors, the main inhibitory ligand-gated ion channel in the brain. Scientists have long theorized that there exists an endogenous benzodiazepine, or endozepine, in the brain. While there is indirect evidence suggesting a peptide, the diazepam binding inhibitor, is capable of modulating the GABA-A receptor, direct evidence of the modulatory effects of the diazepam binding inhibitor is limited. Here we take a reductionist approach to understand how purified diazepam binding inhibitor interacts with and affects GABA-A receptor activity. We used two-electrode voltage clamp electrophysiology to study how the effects of diazepam binding inhibitor vary with GABA-A receptor subunit composition, and found that GABA-evoked currents from α3-containing GABA-A receptors are weakly inhibited by the diazepam binding inhibitor, while currents from α5-containing receptors are positively modulated. We also used in silico protein-protein docking to visualize potential diazepam binding inhibitor/GABA-A receptor interactions that revealed diazepam binding inhibitor bound at the benzodiazepine α/γ binding site interface, which provides a structural framework for understanding diazepam binding inhibitor effects on GABA-A receptors. Our results provide novel insights into mechanisms underlying how the diazepam binding inhibitor modulates GABA-mediated inhibition in the brain.