Both the histamine H-1-receptor (H1R) and H-2-receptor (H2R) exhibit pronounced species selectivity in their pharmacological properties; i.e., bulky agonists possess higher potencies and efficacies at guinea pig (gp) than at the corresponding human (h) receptor isoforms. In this study, we examined the effects of N-G-acylated imidazolylpropylguanidines substituted with a single phenyl or cyclohexyl substituent on H1R and H2R species isoforms expressed in Sf9 insect cells. N-1-(3-Cyclohexyl-butanoyl)N-2-[3-(1H-imidazol-4-yl) propyl]guanidine (UR-AK57) turned out to be the most potent hH(2)R agonist identified so far (EC50 of 23 nM in the GTPase assay at the hH(2)R-G(s alpha) fusion protein expressed in Sf9 insect cells). UR-AK57 was almost a full-hH(2)R agonist and only slightly less potent and efficacious than at gpH(2)R-G(s alpha). Several N-G-acylated imidazolylpropylguanidines showed similar potency at hH(2)R and gpH(2)R. Most unexpectedly, UR-AK57 exhibited moderately strong partial hH(1)R agonism with a potency similar to that of histamine, whereas at gpH(1)R, UR-AK57 was only a very weak partial agonist. Structure/activity relationship studies revealed that both the alkanoyl chain connecting the aromatic or alicyclic substituent with the guanidine moiety and the nature of the carbocycle (cyclohexyl versus phenyl ring) critically determine the pharmacological properties of this class of compounds. Collectively, our data show that gpH(1)R and gpH(2)R do not necessarily exhibit preference for bulky agonists compared with hH(1)R and hH(2)R, respectively, and that UR-AK57 is a promising starting point for the development of both potent and efficacious hH(1)R and hH(2)R agonists.