Raymond G. Booth

Previously, (−)-trans-1-phenyl-3-N,N-dimethylamino-1,2,3,4-tetrahydronaphthalene ([−]-trans-H₂-PAT) was shown to activate stereospecifically histamine H₁ receptors coupled to modulation of tyrosine hydroxylase activity in guinea pig and rat forebrain in vitro and in vivo. Furthermore, the novel radioligand [³H](−)-trans-H₂-PAT was shown to label selectively H₁ receptors in guinea pig and rat brain with high affinity (K_D, ∼0.1 and 0.5 nM, respectively) and a B_max about 50 and 15%, respectively, of that observed for the H₁antagonist radioligand [³H]mepyramine. In the current study, [³H](−)-trans-H₂-PAT-labeled cloned guinea pig and human H₁ receptors in Chinese hamster ovary (CHO) cell membranes with high affinity (K_D, ∼0.08 and 0.23 nM, respectively) and a B_max about 15% of that observed for [³H]mepyramine. The binding of H₂-PAT to H₁ receptors in both CHO-H₁ cell lines was stereoselective with the (−)-trans-isomer having affinity (K_i, ∼1.5 nM) about 4-, 20-, and 50-times higher than the (−)-cis-, (+)-trans-, and (+)-cis-isomers, respectively; the affinity of (−)-trans-H₂-PAT was unaffected by excess GTP. In functional assays, (−)-trans-H₂-PAT was a full antagonist of histamine H₁-mediated stimulation of phospholipase C (PLC) and [³H]inositol phosphates (IP) formation in CHO-H₁ cells, a full inverse agonist of constitutively active H₁ receptors in COS-7-H₁cells, and a full competitive antagonist (pA₂ = 9.2) of histamine H₁-mediated contraction of guinea pig ileum. It is concluded that (−)-trans-H₂-PAT is an antagonist at H₁ receptors coupled to PLC/IP formation and smooth muscle contraction. Meanwhile, the observation that [³H](−)-trans-H₂-PAT labels only a subpopulation of H₁ receptors and that (−)-trans-H₂-PAT activates H₁receptors coupled to modulation of tyrosine hydroxylase suggests that there may be post-translational H₁ receptor heterogeneity.