Pharmacological characterization of the human histamine H₂ receptor stably expressed in Chinese hamster ovary cells

Abstract

4 Displacement studies with histamine showed that a limited fraction (32 ± 6%) of the binding sites showed a high affinity for histamine (2 ± 1.2 gM); the shallow displacement curves were reflected by a Hill-coefficient significantly different from unity (nH = 0.58 ± 0.09). The addition of 100 ILM Gpp(NH)p resulted in a steepening of the displacement curve (nH = 0.79 ± 0.02) and a loss of high affinity sites for histamine. 5 Displacement studies with other agonists indicated that the recently developed specific H2 agonists, amthamine and amselamine, showed an approximately 4-5 fold higher affinity for the human H2 receptor than histamine. 6 Stimulation of CHOhumH2 cells with histamine resulted in a rapid rise of the intracellular cyclic AMP levels. After 10 min an approximately 10 fold increase in cyclic AMP could be measured. The EC50 value for this response was 7 ± 1 nM for histamine. This response was effectively blocked by tiotidine and cimetidine, resulting in Ki values of 8 ± 1 nM and 0.56 ± 0.24 LM respectively.

Evaluation of Histamine H1-, H2-, and H3-Receptor Ligands at the Human Histamine H4 Receptor: Identification of 4-Methylhistamine as the First Potent and Selective H4 Receptor Agonist

“…Within the series of known H 2 R agonists that we have tested, only some ligands retain H 4 R activity. Replacement of the imidazole ring of histamine in the selective H 2 R agonists amthamine and amselamine (Leurs et al, 1994) results in a total loss of hH 4 R activity at concentrations up to 10 M. Dimaprit, a H 2 R agonist/H 3 R antagonist lacking an imidazole group, binds the H 4 R with moderate affinity, showing a pK i value of 6.5 Ϯ 0.1, and exerts partial H 4 R agonistic activity (Table 1). Impromidine, which was reported to bind to both H 2 R and H 3 R, also binds potently to the hH 4 R with a pK i value of 7.6 Ϯ 0.1 and acts as a partial H 4 R agonist (␣ ϭ 0.5).…”

“…In our hands the isothiourea-based H 3 R antagonists clobenpropit and iodophenpropit both potently bind to the hH 4 R (Table 3). Yet, both ligands have very distinguishable intrinsic activities at the H 4 R. Clobenpropit, which acts as inverse I͔iodoaminopotentidine displacement assay (Leurs et al, 1994). b pK i values determined with ͓…”

8R-Lisuride Is a Potent Stereospecific Histamine H1-Receptor Partial Agonist

“…125 I]aminopotentidine ( 125 I-APT) displacement studies using cell homogenates of COS-7 cells transiently transfected with cDNA encoding the human H 2 receptor (pcDNA 3 -H 2 ) as described previously (Leurs et al, 1994). Cell homogenates were incubated for 90 min at 30°C in a total volume of 400 l 50 mM Na 2 /K ϩ -phosphate buffer containing gelatin (0.1%; pH 7.4 at 30°C) with approximately 0.5 nM 125 I-APT.…”

Cloning and Expression of a Complementary DNA Encoding a Molluscan Octopamine Receptor That Couples to Chloride Channels in HEK293 Cells

“…The medium was aspirated and the cells were lysed by sonication in 200 l of ice-cold 0.1 N HCl. Then, 75 l of neutralization buffer (230 mM NaOH, 560 mM Tris, 140 mM NaCl, 56 mM EDTA, 0.35 M HEPES) was added, and the concentration of cAMP was determined as described previously (25).…”