Cloning and characterization of the monkey histamine H3 receptor isoforms

We have recently identified three splice isoforms of the histamine H(3) receptor in multiple brain regions of cynomolgus monkey (Macaca fascicularis). Two of the novel isoforms displayed a deletion in the third intracellular loop (H(3)(413) and H(3)(410)), the third isoform H(3)(335) displayed a deletion in the i3 intracellular loop and a complete deletion of the putative fifth transmembrane domain TM5. We have confirmed by RT-PCR the expression of full-length H(3)(445) mRNA as well as H(3)(413), H(3)(410), and H(3)(335) splice isoform mRNA in multiple monkey brain regions including the frontal, parietal and occipital cortex, parahippocampal gyrus, hippocampus, amygdala, caudate nucleus, putamen, thalamus, hypothalamus, and cerebellum. The full-length isoform H(3)(445) was predominant in all of the regions tested, followed by H(3)(335), with the H(3)(413) and H(3)(410) being of low abundance. When expressed in C6 cells, H(3)(445), H(3)(413), and H(3)(410) exhibit high affinity binding to the agonist ligand [(3)H]-(N)-alpha-methylhistamine with respective pK(D) values of 9.7, 9.7, and 9.6. As expected, the H(3)(335) isoform did not display any saturable binding with [(3)H]-(N)-alpha-methylhistamine. The histamine H(3) receptor agonists histamine, (R)-alpha-methylhistamine, imetit and proxyfan were able to activate calcium mobilization responses through H(3)(445), H(3)(413) and H(3)(410) receptors when they were co-expressed with the chimeric G alpha(qi5)-protein in HEK293 cells, while no response was elicited in cells expressing the H(3)(335) isoform. The existence of multiple H(3) receptor splice isoforms across species raises the possibility that isoform specific properties including ligand affinity, signal transduction coupling, and brain localization may differentially contribute to observed in vivo effects of histamine H(3) receptor antagonists.