Guanosine 5'-(gamma-[35S]thio)triphosphate autoradiography allows selective detection of histamine H3 receptor-dependent G protein activation in rat brain tissue sections.
Histamine elicits its biological effects via three distinct G protein-coupled receptors, termed H1, H2, and H3. We have used guanosine 5'-(gamma-[35S]thio)triphosphate (GTPgamma[35S]) autoradiography to localize histamine receptor-dependent G protein activation in rat brain tissue sections. Initial studies revealed that in basal conditions, adenosine was present in tissue sections in sufficient concentrations to generate an adenosine A1 receptor-dependent GTPgamma[35S] signal in several brain regions. All further incubations therefore contained 8-cyclopentyl-1,3-dipropylxanthine (10 microM), a selective A1 receptor antagonist. Histamine elicited dose-dependent increments in GTPgamma[35S] binding to discrete anatomical structures, most notably the caudate putamen, cerebral cortex, and substantia nigra. The overall anatomical pattern of the histamine-evoked binding response closely reflects the known distribution of H3 binding sites and was faithfully mimicked by N(alpha)-methylhistamine, (R)-alpha-methylhistamine, and immepip, three H3-selective agonists. In all regions examined, the GTPgamma[35S] signal was reversed with thioperamide and clobenpropit, two potent H3-selective antagonists, whereas mepyramine, a specific H1 antagonist, and cimetidine, a prototypic H2 antagonist, proved ineffective. These data indicate that in rat brain tissue sections, GTPgamma[35S] autoradiography selectively detects H3 receptor-dependent signaling in response to histamine stimulation. As the existing evidence suggests that GTPgamma[35S] autoradiography preferentially reveals responses to G(i/o)-coupled receptors, our data indicate that most, if not all, central H3 binding sites represent functional receptors coupling to G(i/o), the inhibitory class of G proteins. Besides allowing more detailed studies on H3 receptor signaling within anatomically restricted regions of the CNS, GTPgamma[35S] autoradiography offers a novel approach for functional in vitro screening of H3 ligands.
Department of Physiology, University of Kuopio, Finland.
J T
JT
Laitinen
M
M
Jokinen
D000241:Adenosine / Q000494:pharmacology; D000818:Animals; D001345:Autoradiography; D001479:Basal Ganglia / Q000737:chemistry / Q000378:metabolism; D001923:Brain Chemistry / Q000502:physiology*; D002540:Cerebral Cortex / Q000737:chemistry / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000378:metabolism*; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000494:pharmacology*; D006153:Guanosine Diphosphate / Q000494:pharmacology; D006632:Histamine / Q000494:pharmacology; D006633:Histamine Antagonists / Q000494:pharmacology; D006634:Histamine H1 Antagonists / Q000494:pharmacology; D008297:Male; D010880:Piperidines / Q000494:pharmacology; D011485:Protein Binding / Q000187:drug effects; D011738:Pyrilamine / Q000494:pharmacology; D051381:Rats; D017208:Rats, Wistar; D018100:Receptors, Histamine H3 / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology; D013378:Substantia Nigra / Q000737:chemistry / Q000378:metabolism; D013462:Sulfur Radioisotopes; D014970:Xanthines / Q000494:pharmacology
D006633:Histamine Antagonists; D006634:Histamine H1 Antagonists; D010880:Piperidines; D018100:Receptors, Histamine H3; D013462:Sulfur Radioisotopes; D014970:Xanthines; D006153:Guanosine Diphosphate; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D006632:Histamine; C051360:1,3-dipropyl-8-cyclopentylxanthine; D019204:GTP-Binding Proteins; D011738:Pyrilamine; C052075:thioperamide; D000241:Adenosine