Modulation of ligands binding to the benzodiazepine receptor by an endogenous inhibitor (GABA-modulin) and bicuculline.

We have prepared and purified an endogenous protein (brain neuropeptide) by extraction with 0.025 N acetic acid and precipitation with a saturated ammonium sulfate solution followed by column chromatography. This protein was isolated and partially purified (approx. 500 times). When added to Triton X-100 treated synaptic membrane preparation obtained from rat CNS, this modulates specific high affinity GABA binding site without affecting low affinity site. This protein was called GABA-modulin (GM). GM like bicuculline (competitive GABA antagonist) modulates the affinity of benzodiazepine receptor ligands. But the effect of bicuculline is more important than that of GM. Bicuculline increases or decreases respectively the binding of 3H-beta-CCM or 3H-flunitrazepam by approximately 60%. In the same conditions, at a concentration of 12 micrograms/ml GM increases or decreases respectively the binding of 3H-beta-CCM or 3H-flunitrazepam by approximately 35% and no more modulation was obtained even the concentration of GM was increased. Bicuculline and GM doesn't modify the specific binding of 3H-Ro 15-1788 to the BZD receptor. It is concluded that GM exhibit a lower ability than bicuculline to modulate the binding of the BZD receptor ligands because GM interacts only with the GABA high affinity binding site without affecting the low affinity binding site. These observations suggest that the two GABA binding sites probably interact with the BZD receptor.