Title Page Probing ligand-specific histamine H 1 -and H 2 -receptor conformations with N G -acylated imidazolylpropylguanidines Running Title Page Running title: Ligand-specific histamine receptor conformations

Abstract

G sαS , fusion protein of the guinea pig histamine H 2 -receptor and the short splice variant of G sα ; GTPγS, guanosine 5'-[γ-thio]triphosphate; hH 1 R, human histamine H 1 -receptor; hH 2 R, human histamine H 2 -receptor; hH 2 R-G sαS , fusion protein of the human histamine H 2 -receptor and the short splice variant of G sα ; HIS, histamine; IMP, impromidine; RGS protein, regulator of G-protein signaling; rH 2 R, rat histamine H 2 -receptor; TM, transmembrane domain. Section: Cellular & Molecular JPET #97923 3 Abstract Impromidine (IMP) and arpromidine (ARP)-derived guanidines are more potent and efficacious guinea pig (gp) histamine H 2 -receptor (gpH 2 R)-than human (h) H 2 R agonists and histamine H 1 -receptor (H 1 R) antagonists with preference for hH 1 R relative to gpH 1 R. We examined N G -acylated imidazolylpropylguanidines (AIPGs) which are less basic than guanidines at hH 2 R, gpH 2 R, rat H 2 R (rH 2 R), hH 1 R and gpH 1 R expressed in Sf9 cells as probes for ligand-specific receptor conformations. AIPGs were similarly potent H 2 R agonists as the corresponding guanidines IMP and ARP, respectively. Exchange of pyridyl in ARP against phenyl increased AIPG potency ten-fold, yielding the most potent agonists at the hH 2 R-G sα fusion protein and gpH 2 R-G sα identified so far. Some AIPGs were similarly potent and efficacious at hH 2 R-G sα and gpH 2 R-G sα . AIPGs stabilized the ternary complex in hH 2 R-G sα and gpH 2 R-G sα differently than the corresponding guanidines. Guanidines, AIPGs and small H 2 R agonists exhibited distinct agonist properties at hH 2 R, gpH 2 R and rH 2 R measuring adenylyl cyclase activity. In contrast to ARP and IMP, AIPGs were partial H 1 R agonists exhibiting higher efficacies at hH 1 R than at gpH 1 R. This is remarkable since so far, all bulky H 1 R agonists exhibited higher efficacies at gpH 1 R than at hH 1 R. Collectively, our data suggest that AIPGs stabilize different active conformations in hH 2 R, gpH 2 R and rH 2 R than guanidines and that in contrast to guanidines, AIPGs are capable of stabilizing a partially active state of hH 1 R. JPET #97923