H3 receptor-mediated inhibition of intestinal acetylcholine release: pharmacological characterization of signal transduction pathways.
The present study investigates the mechanisms through which prejunctional histamine H3 receptors modulate intestinal cholinergic neurotransmission. The experiments were performed on longitudinal muscle-myenteric plexus preparations of guinea pig ileum, preincubated with [3H]choline, superfused with physiological salt solution containing hemicholinium-3, and subjected to electrical field stimulation. The stimulation-induced outflow of radioactivity was taken as an index of endogenous acetylcholine release. The electrically induced [3H]acetylcholine release was inhibited by histamine (EC50)=33.5 nM) or the H3 receptor agonist R-alpha-methylhistamine (EC50=41.6 nM), whereas it was not affected by pyridylethylamine (H1 agonist), impromidine (H2 agonist), pyrilamine (H1 antagonist), cimetidine (H2 antagonist), thioperamide or clobenpropit (H3 antagonists). The inhibitory effects of histamine or R-alpha-methylhistamine were antagonized by thioperamide (pKd= 8.31 and 8.53, respectively) or clobenpropit (pKd=9.44 and 9.32, respectively), but not by pyrilamine or cimetidine. The modulatory action of histamine on the evoked tritium outflow was attenuated by pertussis toxin and abolished by N-ethylmaleimide, two selective blockers of Gi/Go proteins. Tetraethylammonium or 4-aminopyridine, acting as inhibitors of voltage-dependent K+ channels, enhanced the evoked tritium outflow when tested alone, and apparently counteracted the inhibitory effect of histamine. However, the blocking actions of tetraethylammonium and 4-aminopyridine were no longer evident when their enhancing actions were compensated by appropriate reductions of Ca2+ concentration in the superfusion medium. Histamine-induced inhibition of evoked tritium output was enhanced by omega-conotoxin, a selective blocker of N-type Ca2+ channels, or low Ca2+ concentration, whereas it was not modified by nifedipine, an antagonist of L-type Ca2+ channels. In addition, the inhibitory effect of histamine was not significantly affected by forskolin (activator of adenylyl cyclase), 8-bromo-cyclic AMP (a stable analog of cyclic AMP), rolipram (a selective blocker of type IV phosphodiesterase), phorbol myristate acetate (activator of protein kinase C), H-89 (N-(2-[p-bromocinnamylamino]ethyl)-5-isoquinolinesulfonamide, inhibitor of protein kinase A), Ro-31-8220 (2-(1-[3-(amidinothio)propyl]-1H-indol-3-yl)-3-(1-methylindol-3-yl)-maleimide, inhibitor of protein kinase C), KT5823 (N-methyl-(8R*,9S*,11S*)-(-)-9-methoxy-9-methoxycarbonyl-8-methyl-2,3,9,10-tetrahydro-8,11-epoxy-1H,8H,11H-2,7b,11a-triazadibenzo [a,g]cycloocta[c,d,e]-trinden-1-one, inhibitor of protein kinase G), or lavendustin A (inhibitor of tyrosine kinase). The present results indicate that histamine inhibits intestinal cholinergic neurotransmission through presynaptic H3 receptors coupled to Gi/Go proteins. It is suggested that adenylyl cyclase, serine-threonine protein kinase and tyrosine kinase pathways are not implicated in this regulatory action, and that Gi/Go proteins modulate the activity of N-type Ca2+ channels through a direct link, thus causing a reduced availability of extracellular Ca2+ at the level of ileal cholinergic nerve terminals.
Blandizzi
C
C
Department of Oncology, Transplants and Advanced Technologies in Medicine, University of Pisa, Italy.
Colucci
R
R
Tognetti
M
M
De Paolis
B
B
Del Tacca
M
M
D000109:Acetylcholine / Q000378:metabolism*; D000818:Animals; D002118:Calcium / Q000494:pharmacology; D015220:Calcium Channels / Q000187:drug effects / Q000502:physiology; D002799:Cholinergic Fibers / Q000187:drug effects* / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004558:Electric Stimulation; D019204:GTP-Binding Proteins / Q000187:drug effects / Q000378:metabolism; D006168:Guinea Pigs; D006632:Histamine / Q000494:pharmacology; D017442:Histamine Agonists / Q000494:pharmacology*; D006633:Histamine Antagonists / Q000494:pharmacology*; D007082:Ileum / Q000187:drug effects / Q000378:metabolism; D008297:Male; D008761:Methylhistamines / Q000494:pharmacology; D009197:Myenteric Plexus / Q000187:drug effects / Q000378:metabolism; D018100:Receptors, Histamine H3 / Q000187:drug effects* / Q000502:physiology; D015398:Signal Transduction / Q000187:drug effects* / Q000502:physiology
D015220:Calcium Channels; D017442:Histamine Agonists; D006633:Histamine Antagonists; D008761:Methylhistamines; D018100:Receptors, Histamine H3; C069357:alpha-methylhistamine; D006632:Histamine; D019204:GTP-Binding Proteins; D000109:Acetylcholine; D002118:Calcium