title: Presynaptic P2X1-3 and α3-containing nicotinic receptors assemble into functionally interacting ion channels in the rat hippocampus.
abstract: Previous studies documented a cross-talk between purinergic P2X (P2XR) and nicotinic acetylcholine receptors (nAChR) in heterologous expression systems and peripheral preparations. We now investigated if this occurred in native brain preparations and probed its physiological function. We found that P2XR and nAChR were enriched in hippocampal terminals, where both P2X1-3R and α3, but not α4, nAChR subunits were located in the active zone and in dopamine-β-hydroxylase-positive hippocampal terminals. Notably, P2XR ligands displaced nAChR binding and nAChR ligands displaced P2XR binding to hippocampal synaptosomes. In addition, a negative P2XR/nAChR cross-talk was observed in the control of the evoked release of noradrenaline from rat hippocampal synaptosomes, characterized by a less-than-additive facilitatory effect upon co-activation of both receptors. This activity-dependent cross-inhibition was confirmed in Xenopus oocytes transfected with P2X1-3Rs and α3β2 (but not α4β2) nAChR. Besides, P2X2 co-immunoprecipitated α3β2 (but not α4β2) nAChR, both in HEK cells and rat hippocampal membranes indicating that this functional interaction is supported by a physical association between P2XR and nAChR. Moreover, eliminating extracellular ATP with apyrase in hippocampal slices promoted the inhibitory effect of the nAChR antagonist tubocurarine on noradrenaline release induced by high- but not low-frequency stimulation. Overall, these results provide integrated biochemical, pharmacological and functional evidence showing that P2X1-3R and α3β2 nAChR are physically and functionally interconnected at the presynaptic level to control excessive noradrenergic terminal activation upon intense synaptic firing in the hippocampus.
authors.affiliation: CNC-Center for Neuroscience and Cell Biology, University of Coimbra, 3004-504, Portugal. Electronic address: ricardojrodrigues@gmail.com.
authors.forename: Ricardo J
authors.initials: RJ
authors.lastname: Rodrigues
authors.affiliation: CNC-Center for Neuroscience and Cell Biology, University of Coimbra, 3004-504, Portugal; Department of Biochemistry and Molecular Biology, Faculty of Biology, University of Barcelona, 08028, Spain; Department of Pathology and Experimental Therapeutics, Faculty of Medicine, IDIBELL, University of Barcelona, 08907, Spain.
authors.forename: Teresa
authors.initials: T
authors.lastname: Almeida
authors.affiliation: Department of Biochemistry and Molecular Biology, Faculty of Veterinary, Complutense University of Madrid, 28040, Spain.
authors.forename: Miguel
authors.initials: M
authors.lastname: Díaz-Hernández
authors.affiliation: CNC-Center for Neuroscience and Cell Biology, University of Coimbra, 3004-504, Portugal.
authors.forename: Joana M
authors.initials: JM
authors.lastname: Marques
authors.affiliation: Department of Biochemistry and Molecular Biology, Faculty of Biology, University of Barcelona, 08028, Spain; CIBERNED, Centro de Investigación en Red, Enfermedades Neurodegenerativas, Instituto de Salud Carlos III, Madrid, Spain.
authors.forename: Rafael
authors.initials: R
authors.lastname: Franco
authors.affiliation: Department of Pathology and Experimental Therapeutics, Faculty of Medicine, IDIBELL, University of Barcelona, 08907, Spain.
authors.forename: Carles
authors.initials: C
authors.lastname: Solsona
authors.affiliation: Department of Biochemistry and Molecular Biology, Faculty of Veterinary, Complutense University of Madrid, 28040, Spain.
authors.forename: María Teresa
authors.initials: MT
authors.lastname: Miras-Portugal
authors.affiliation: Department of Pathology and Experimental Therapeutics, Faculty of Medicine, IDIBELL, University of Barcelona, 08907, Spain; Department of Physiology, Faculty of Sciences, University of Ghent, B-9000, Belgium.
authors.forename: Francisco
authors.initials: F
authors.lastname: Ciruela
authors.affiliation: CNC-Center for Neuroscience and Cell Biology, University of Coimbra, 3004-504, Portugal; Faculty of Medicine, University of Coimbra, 3004-504, Portugal.
authors.forename: Rodrigo A
authors.initials: RA
authors.lastname: Cunha
mesh_terms: D000818:Animals; D004299:Dopamine beta-Hydroxylase / Q000378:metabolism; D057809:HEK293 Cells; D006624:Hippocampus / Q000254:growth & development* / Q000502:physiology*; D006801:Humans; D007473:Ion Channels / Q000502:physiology*; D008297:Male; D009411:Nerve Endings / Q000378:metabolism; D009638:Norepinephrine / Q000378:metabolism; D009865:Oocytes; D051381:Rats; D017208:Rats, Wistar; D020239:Receptor Cross-Talk / Q000502:physiology; D011978:Receptors, Nicotinic / Q000502:physiology*; D017661:Receptors, Presynaptic / Q000502:physiology*; D058469:Receptors, Purinergic P2X / Q000502:physiology*; D058474:Receptors, Purinergic P2X1 / Q000502:physiology; D058476:Receptors, Purinergic P2X2 / Q000502:physiology; D058477:Receptors, Purinergic P2X3 / Q000502:physiology; D013574:Synaptosomes / Q000378:metabolism; D014981:Xenopus; D064569:alpha7 Nicotinic Acetylcholine Receptor / Q000378:metabolism
chemical_list: D007473:Ion Channels; D011978:Receptors, Nicotinic; D017661:Receptors, Presynaptic; D058469:Receptors, Purinergic P2X; D058474:Receptors, Purinergic P2X1; D058476:Receptors, Purinergic P2X2; D058477:Receptors, Purinergic P2X3; D064569:alpha7 Nicotinic Acetylcholine Receptor; D004299:Dopamine beta-Hydroxylase; D009638:Norepinephrine
keywords: ATP (PubChem CID:5957); Cytisine (PubChem CID:10235); DHβE (PubChem CID:31762); Epibatidine (PubChem CID:3073763); Hippocampus; Methyllycaconitine (PubChem CID:5288811); NF023 (PubChem CID:4465); Neuromodulation; Nicotine (PubChem CID:942); Nicotinic receptors; Noradrenaline; P2X receptors; PPADS (PubChem CID:6093163); Presynaptic; Tubocurarine (PubChem CID:6000); α,β-methylene ATP (PubChem CID:91557)