title: Constitutive activity of the A2A adenosine receptor and compartmentalised cyclic AMP signalling fine-tune noradrenaline release.
abstract: Neuroblastoma SH-SY5Y (SH) cells endogenously express A(2A) adenosine receptors and can be differentiated into a sympathetic neuronal phenotype, capable of depolarisation-dependent noradrenaline release. Using differentiated SH culture, we here explored the link between A(2A)-receptor signalling and neurotransmitter release. In response to the receptor agonist CGS21680, the cells produced cyclic AMP (cAMP), and when depolarised, they released increased amounts of noradrenaline. An A(2A)-receptor antagonist, XAC, as well as an inhibitor of cAMP-dependent protein kinase A (PKA), H89, depressed agonist-dependent release. In the presence of XAC or H89, noradrenaline release was found to be below basal values. This suggested that release facilitation also owes to constitutive receptor activity. We demonstrate that even in the absence of an agonist, the native A(2A)-receptor stimulated cAMP production, leading to the activation of PKA and enhanced noradrenaline release. Ancillary, non-cAMP-dependent effects of the receptor (i.e. phosphorylation of CREB, of Rabphilin3A) were refractory to constitutive activation. PKA-dependent facilitation of noradrenaline release was recapitulated with membrane-permeable 8-Br-cAMP; in addition to facilitation, 8-Br-cAMP caused marked inhibition of release, an effect not observed upon receptor activation. Inhibition by receptor-independent cAMP was likely due to suppression of voltage-dependent calcium current (VDCC) and increased activity of Src-family kinases. Receptor-mediated release facilitation was reproduced in the presence of tetrodotoxin (blocking action potentials); hence, the signalling occurred at the active zone comprising release sites. Our findings thus support (1) presynaptic localisation of the A(2A)-receptor and (2) suggest that compartmentalised pathways transmit cAMP signalling in order to facilitate depolarisation-dependent neurotransmitter release.
authors.affiliation: Centre for Physiology and Pharmacology, Institute of Pharmacology, Medizinische Universität Wien, Währinger Strasse 13A, 1090, Wien, Austria.
authors.forename: Edin
authors.initials: E
authors.lastname: Ibrisimovic
authors.forename: Helmut
authors.initials: H
authors.lastname: Drobny
authors.forename: Qiong
authors.initials: Q
authors.lastname: Yang
authors.forename: Thomas
authors.initials: T
authors.lastname: Höfer
authors.forename: Stefan
authors.initials: S
authors.lastname: Boehm
authors.forename: Christian
authors.initials: C
authors.lastname: Nanoff
authors.forename: Klaus
authors.initials: K
authors.lastname: Schicker
mesh_terms: D000241:Adenosine / Q000031:analogs & derivatives / Q000494:pharmacology; D058908:Adenosine A2 Receptor Agonists / Q000494:pharmacology*; D002454:Cell Differentiation / Q000502:physiology; D002460:Cell Line; D000242:Cyclic AMP / Q000378:metabolism*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism*; D006801:Humans; D009474:Neurons / Q000187:drug effects / Q000378:metabolism; D009638:Norepinephrine / Q000378:metabolism*; D010627:Phenethylamines / Q000494:pharmacology; D043705:Receptor, Adenosine A2A / Q000378:metabolism*; D015398:Signal Transduction; D009435:Synaptic Transmission / Q000187:drug effects
chemical_list: D058908:Adenosine A2 Receptor Agonists; D010627:Phenethylamines; D043705:Receptor, Adenosine A2A; C061282:2-(4-(2-carboxyethyl)phenethylamino)-5'-N-ethylcarboxamidoadenosine; D000242:Cyclic AMP; D017868:Cyclic AMP-Dependent Protein Kinases; D000241:Adenosine; D009638:Norepinephrine