Modulation of histamine H3 receptor function by monovalent ions

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 H3 receptor (hH3R), co-expressed with mammalian G proteins (Gαi1, Gαi2, Gαi3 or Gαo1, and β1γ2 dimers, respectively) in Sf9 insect cell membranes, with respect to agonist binding and G protein activation. NaCl (100 mM) had no effect on affinity of the agonist [3H]Nα-methylhistamine ([3H]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 (100 mM) was present. Monovalent ions reduced H3R-regulated signalling in the order of efficacy Li+ ∼ Na+ ∼ K+ < Cl− < Br− < I−. NaCl had a stronger effect on basal hH3R-signalling when Gαi3 was co-expressed. Asp802.50, a putative interaction site for Na+, was mutated to Asn802.50 (D2.50N-hH3R). Strikingly, the mutation was unable to activate Gαi3 at all. The effects can be explained by a model, where (i) monovalent ions as well as a charge-neutralizing mutation of Asp802.50 generally reduce the interaction of hH3R with G proteins, (ii) monovalent anions increase the affinity of G proteins for GDP and thus, indirectly affect their interaction with hH3R and, (iii) Asp802.50 is a key residue for hH3R/Gαi3-protein activation. The latter result suggests that hH3R/G protein-coupling interfaces may differ even between closely related subunits.