Cloning and characterization of the monkey histamine H3 receptor isoforms

We have recently identified three splice isoforms of the histamine H3 receptor in multiple brain regions of cynomolgus monkey (Macaca fascicularis). Two of the novel isoforms displayed a deletion in the third intracellular loop (H3(413) and H3(410)), the third isoform H3(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 H3(445) mRNA as well as H3(413), H3(410), and H3(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 H3(445) was predominant in all of the regions tested, followed by H3(335), with the H3(413) and H3(410) being of low abundance. When expressed in C6 cells, H3(445), H3(413), and H3(410) exhibit high affinity binding to the agonist ligand [3H]-(N)-α-methylhistamine with respective pKD values of 9.7, 9.7, and 9.6. As expected, the H3(335) isoform did not display any saturable binding with [3H]-(N)-α-methylhistamine. The histamine H3 receptor agonists histamine, (R)-α-methylhistamine, imetit and proxyfan were able to activate calcium mobilization responses through H3(445), H3(413) and H3(410) receptors when they were co-expressed with the chimeric Gαqi5-protein in HEK293 cells, while no response was elicited in cells expressing the H3(335) isoform. The existence of multiple H3 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 H3 receptor antagonists.