Stimulation of human H1 and H2-histamine receptors (HRs) primarily activates signaling pathways to increase intracellular calcium [Ca2+]i and cyclic AMP (cAMP), respectively. Activation of H2-HR in human embryonic kidney (HEK) cells by histamine and dimaprit increases both cAMP formation and [Ca2+]i, as determined by cAMP-scintillation proximity assays and fluorescence imaging plate reader (FLIPR) assays. In HEK cells expressing relatively high levels of H2-HR (Bmax=26 pmol/mg protein), histamine and dimaprit are full agonists in eliciting cAMP responses with pEC50 values of 9.30 and 7.72 that are 1000-fold more potent than their respective pEC50 values of 6.13 and 4.91 for increasing [Ca2+]i. The agonist potencies decrease for both responses at lower H2-HR density (5 pmol/mg protein) and dimaprit exhibits partial agonist behavior for the [Ca2+]i response. The inverse agonists ranitidine and cimetidine more potently inhibit cAMP production in the higher expressing H2-HR line. Histamine also activated both signaling pathways via human H1-HRs highly expressed (Bmax=17 pmol/mg protein) in HEK cells, with a 1000-fold greater potency for [Ca2+]i vs. cAMP responses (pEC50=7.86 and 4.82, respectively). These studies demonstrate a markedly different potency for activation of multiple signaling pathways by H1- and H2-HRs that may contribute to the selectivity of histamine responses in vivo.