GUANOSINE 5'-(GAMMA-[S-35]THIO)TRIPHOSPHATE AUTORADIOGRAPHY ALLOWS SELECTIVE DETECTION OF HISTAMINE H-3 RECEPTOR-DEPENDENT G-PROTEIN ACTIVATION IN RAT-BRAIN TISSUE-SECTIONS

Histamine elicits its biological effects via three distinct G protein- coupled receptors, termed H-1, H-2, and H-3. We have used guanosine 5' -(gamma-[S-35]thio)triphosphate (GTP gamma[S-35]) autoradiography to l ocalize histamine receptor-dependent G protein activation in rat brain tissue sections. Initial studies revealed that in basal conditions, a denosine was present in tissue sections in sufficient concentrations t o generate an adenosine A(1) receptor-dependent GTP gamma[S-35] signal in several brain regions. All further incubations therefore contained 8-cyclopentyl- 1-,3-dipropylxanthine (10 mu M), a selective A(1) rece ptor antagonist. Histamine elicited dose-dependent increments in GTP g amma[S-35] binding to discrete anatomical structures, most notably the caudate putamen, cerebral cortex, and substantia nigra. The overall a natomical pattern of the histamine-evoked binding response closely ref lects the known distribution of H, binding sites and was faith fully m imicked by N-alpha-methylhistamine, (R)-alpha-methylhistamine, and imm epip, three H,-selective agonists. In all regions examined, the GTP ga mma[S-35] signal was reversed with thioperamide and clobenpropit, two potent H-2-selective antagonists, whereas mepyramine, a specific H-1 a ntagonist, and cimetidine, a prototypic H-2 antagonist, proved ineffec tive. These data indicate that in rat brain tissue sections, GTP gamma [S-35] autoradiography selectively detects H-3 receptor-dependent sign aling in response to histamine stimulation. As the existing evidence s uggests that GTP gamma[S-35] autoradiography preferentially reveals re sponses to G(i/o) -coupled receptors, our data indicate that most, if not all, central H-3 binding sites represent lunctional receptors coup ling to G(i/o), the inhibitory class of G proteins. Besides allowing m ore detailed studies on H-3 receptor signaling within anatomically res tricted regions of the CNS, GTP gamma[S-35] autoradiography offers a n ovel approach for functional in vitro screening of H-3 ligands.