Characterization and solubilization of the sulphonylurea receptor in rat brain

Abstract

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 K d of 0.58 and 17nM, and β max of 123 and 392 fmol/mg protein, respectively. When brain microsomes were solubilized with CHAPS, high affinity sites were retained with K d and β 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 IC 50, at 5 nM, 25 μM and 130 μM, 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 β-cell receptor. However, inhibition of the binding by nucleotides is not identical, possibly reflecting differences in the nucleotide-binding subunit.