Modulation of histamine H(3) receptor function by monovalent ions

Abstract

Monovalent ions differently affect ligand binding to G protein-coupled receptors (GPCRs) by as yet poorly defined mechanisms. In particular, NaCl often decreases the affinity of agonists but increases it for antagonists. We examined the effect of various monovalent ions on human histamine H(3) receptor (hH(3)R), co-expressed with mammalian G proteins (Galpha(i1), Galpha(i2), Galpha(i3) or Galpha(o1), and beta(1)gamma(2) dimers, respectively) in Sf9 insect cell membranes, with respect to agonist binding and G protein activation. NaCl (100mM) had no effect on affinity of the agonist [(3)H]N(alpha)-methylhistamine ([(3)H]NAMH). In steady-state GTPase assays, the endogenous agonist histamine had a lower potency and the inverse agonist thioperamide had a higher potency, when NaCl (100mM) was present. Monovalent ions reduced H(3)R-regulated signalling in the order of efficacy Li(+) approximately Na(+) approximately K(+)<Cl(-)<Br(-)<I(-). NaCl had a stronger effect on basal hH3R-signalling when Galpha(i3) was co-expressed. Asp80(2.50), a putative interaction site for Na(+), was mutated to Asn80(2.50) (D2.50N-hH(3)R). Strikingly, the mutation was unable to activate Galpha(i3) at all. The effects can be explained by a model, where (i) monovalent ions as well as a charge-neutralizing mutation of Asp80(2.50) generally reduce the interaction of hH(3)R with G proteins, (ii) monovalent anions increase the affinity of G proteins for GDP and thus, indirectly affect their interaction with hH(3)R and, (iii) Asp80(2.50) is a key residue for hH(3)R/Galpha(i3)-protein activation. The latter result suggests that hH(3)R/G protein-coupling interfaces may differ even between closely related subunits.