Characterization and solubilization of the sulphonylurea receptor in rat brain

The binding characteristics of the sulphonylurea receptor were investigated using rat brain microsomes. Scatchard plots for binding of [3H]glibenclamide, a potent sulphonylurea which inhibits the ATP-sensitive K-channel, suggested the presence of both high and low affinity binding sites with Kd of 0.58 and 17 nM, and beta max of 123 and 392 fmol/mg protein, respectively. When brain microsomes were solubilized with CHAPS, high affinity sites were retained with Kd and beta max of 1.2 nM and 42.1 fmol/mg protein, respectively, whereas the low affinity sites disappeared. The specific binding was displaced by non-labelled glibenclamide, meglitinide, and tolbutamide with IC50 at 5 nM, 25 microM and 130 microM, respectively. ATP and GTP inhibited [3H]glibenclamide binding in a Mg-dependent manner whereas the inhibition by ADP and GDP was Mg-independent [3H]Glibenclamide binding to the solubilized receptor was similarly inhibited by those nucleotides. Diazoxide inhibited [3H]glibenclamide binding in the presence of MgATP, but after CHAPS-solubilization diazoxide failed to inhibit [3H]glibenclamide binding even with MgATP. These findings suggest the brain sulphonylurea receptor has similar features to the beta-cell receptor. However, inhibition of the binding by nucleotides is not identical, possibly reflecting differences in the nucleotide-binding subunit.