title: Attenuated aortic vasodilation and sympathetic prejunctional facilitation in epinephrine-deficient mice: selective impairment of β2-adrenoceptor responses.
abstract: It has been suggested that there is a link between epinephrine synthesis and the development of β2-adrenoceptor-mediated effects, but it remains to be determined whether this development is triggered by epinephrine. The aim of this study was to characterize β-adrenoceptor-mediated relaxation and facilitation of norepinephrine release in the aorta of phenylethanolamine-N-methyltransferase-knockout (Pnmt-KO) mice. Catecholamines were quantified by reverse-phase high-performance liquid chromatography-electrochemical detection. Aortic rings were mounted in a myograph to determine concentration-response curves to selective β1- or β2-adrenoceptor agonists in the absence or presence of selective β1- or β2-adrenoceptor antagonists. Aortic rings were also preincubated with [(3)H]norepinephrine to measure tritium overflow elicited by electrical stimulation in the presence of increasing concentrations of nonselective β- or selective β2-adrenoceptor agonists. β2-Adrenoceptor protein density was evaluated by Western blotting and β2-adrenoceptor localization by immunohistochemistry. Epinephrine is absent in Pnmt-KO mice. The potency and the maximal effect of the β2-adrenoceptor agonist terbutaline were lower in Pnmt-KO than in wild-type (WT) mice. The selective β2-adrenoceptor antagonist ICI 118,551 [(±)-erythro-(S*,S*)-1-[2,3-(dihydro-7-methyl-1H-inden-4-yl)oxy]-3-[(1-methylethyl)amino]-2-butanol hydrochloride] antagonized the relaxation caused by terbutaline in WT but not in Pnmt-KO mice. Isoproterenol and terbutaline induced concentration-dependent increases in tritium overflow in WT mice only. β2-Adrenoceptor protein density was decreased in membrane aorta homogenates of Pnmt-KO mice, and this finding was supported by immunofluorescence confocal microscopy. In conclusion, epinephrine is crucial for β2-adrenoceptor-mediated vasodilation and facilitation of norepinephrine release. In the absence of epinephrine, β2-adrenoceptor protein density was decreased in aorta cell membranes, thus potentially hindering its functional activity.
authors.affiliation: Laboratory of General Physiology (M.M.-R.) and Laboratory of Pharmacology and Neurobiology (F.F., P.C.), Unit for Multidisciplinary Investigation in Biomedicine, Institute of Biomedical Sciences Abel Salazar, University of Porto; Department of Pharmacology and Therapeutics, Faculty of Medicine, University of Porto (A.L.G., M.F., J.A., P.S., D.M.); Neuropharmacology, Institute of Molecular and Cellular Biology, University of Porto (M.M., D.M.); Center for Drug Discovery and Innovative Medicines, University of Porto (M.M.-R., A.L.G., M.F., J.A., P.S., M.M., F.F., P.C., D.M.); Laboratory of Pharmacology, Department of Drug Sciences, Faculty of Pharmacy, University of Porto and Rede de Química e Tecnologia (REQUIMTE), Porto, Portugal (M.M.); and Burnett School of Biomedical Sciences, College of Medicine, University of Central Florida, Orlando, Florida (S.N.E.). mirodrigues@icbas.up.pt.
authors.forename: Mónica
authors.identifier: http://orcid.org/0000-0003-1373-9202
authors.initials: M
authors.lastname: Moreira-Rodrigues
authors.affiliation: Laboratory of General Physiology (M.M.-R.) and Laboratory of Pharmacology and Neurobiology (F.F., P.C.), Unit for Multidisciplinary Investigation in Biomedicine, Institute of Biomedical Sciences Abel Salazar, University of Porto; Department of Pharmacology and Therapeutics, Faculty of Medicine, University of Porto (A.L.G., M.F., J.A., P.S., D.M.); Neuropharmacology, Institute of Molecular and Cellular Biology, University of Porto (M.M., D.M.); Center for Drug Discovery and Innovative Medicines, University of Porto (M.M.-R., A.L.G., M.F., J.A., P.S., M.M., F.F., P.C., D.M.); Laboratory of Pharmacology, Department of Drug Sciences, Faculty of Pharmacy, University of Porto and Rede de Química e Tecnologia (REQUIMTE), Porto, Portugal (M.M.); and Burnett School of Biomedical Sciences, College of Medicine, University of Central Florida, Orlando, Florida (S.N.E.).
authors.forename: Ana L
authors.initials: AL
authors.lastname: Graça
authors.affiliation: Laboratory of General Physiology (M.M.-R.) and Laboratory of Pharmacology and Neurobiology (F.F., P.C.), Unit for Multidisciplinary Investigation in Biomedicine, Institute of Biomedical Sciences Abel Salazar, University of Porto; Department of Pharmacology and Therapeutics, Faculty of Medicine, University of Porto (A.L.G., M.F., J.A., P.S., D.M.); Neuropharmacology, Institute of Molecular and Cellular Biology, University of Porto (M.M., D.M.); Center for Drug Discovery and Innovative Medicines, University of Porto (M.M.-R., A.L.G., M.F., J.A., P.S., M.M., F.F., P.C., D.M.); Laboratory of Pharmacology, Department of Drug Sciences, Faculty of Pharmacy, University of Porto and Rede de Química e Tecnologia (REQUIMTE), Porto, Portugal (M.M.); and Burnett School of Biomedical Sciences, College of Medicine, University of Central Florida, Orlando, Florida (S.N.E.). mirodrigues@icbas.up.pt.
authors.forename: Marlene
authors.initials: M
authors.lastname: Ferreira
authors.affiliation: Laboratory of General Physiology (M.M.-R.) and Laboratory of Pharmacology and Neurobiology (F.F., P.C.), Unit for Multidisciplinary Investigation in Biomedicine, Institute of Biomedical Sciences Abel Salazar, University of Porto; Department of Pharmacology and Therapeutics, Faculty of Medicine, University of Porto (A.L.G., M.F., J.A., P.S., D.M.); Neuropharmacology, Institute of Molecular and Cellular Biology, University of Porto (M.M., D.M.); Center for Drug Discovery and Innovative Medicines, University of Porto (M.M.-R., A.L.G., M.F., J.A., P.S., M.M., F.F., P.C., D.M.); Laboratory of Pharmacology, Department of Drug Sciences, Faculty of Pharmacy, University of Porto and Rede de Química e Tecnologia (REQUIMTE), Porto, Portugal (M.M.); and Burnett School of Biomedical Sciences, College of Medicine, University of Central Florida, Orlando, Florida (S.N.E.).
authors.forename: Joana
authors.identifier: http://orcid.org/0000-0001-9941-0613
authors.initials: J
authors.lastname: Afonso
authors.affiliation: Laboratory of General Physiology (M.M.-R.) and Laboratory of Pharmacology and Neurobiology (F.F., P.C.), Unit for Multidisciplinary Investigation in Biomedicine, Institute of Biomedical Sciences Abel Salazar, University of Porto; Department of Pharmacology and Therapeutics, Faculty of Medicine, University of Porto (A.L.G., M.F., J.A., P.S., D.M.); Neuropharmacology, Institute of Molecular and Cellular Biology, University of Porto (M.M., D.M.); Center for Drug Discovery and Innovative Medicines, University of Porto (M.M.-R., A.L.G., M.F., J.A., P.S., M.M., F.F., P.C., D.M.); Laboratory of Pharmacology, Department of Drug Sciences, Faculty of Pharmacy, University of Porto and Rede de Química e Tecnologia (REQUIMTE), Porto, Portugal (M.M.); and Burnett School of Biomedical Sciences, College of Medicine, University of Central Florida, Orlando, Florida (S.N.E.).
authors.forename: Paula
authors.identifier: http://orcid.org/0000-0002-3663-7500
authors.initials: P
authors.lastname: Serrão
authors.affiliation: Laboratory of General Physiology (M.M.-R.) and Laboratory of Pharmacology and Neurobiology (F.F., P.C.), Unit for Multidisciplinary Investigation in Biomedicine, Institute of Biomedical Sciences Abel Salazar, University of Porto; Department of Pharmacology and Therapeutics, Faculty of Medicine, University of Porto (A.L.G., M.F., J.A., P.S., D.M.); Neuropharmacology, Institute of Molecular and Cellular Biology, University of Porto (M.M., D.M.); Center for Drug Discovery and Innovative Medicines, University of Porto (M.M.-R., A.L.G., M.F., J.A., P.S., M.M., F.F., P.C., D.M.); Laboratory of Pharmacology, Department of Drug Sciences, Faculty of Pharmacy, University of Porto and Rede de Química e Tecnologia (REQUIMTE), Porto, Portugal (M.M.); and Burnett School of Biomedical Sciences, College of Medicine, University of Central Florida, Orlando, Florida (S.N.E.).
authors.forename: Manuela
authors.identifier: http://orcid.org/0000-0002-9509-0613
authors.initials: M
authors.lastname: Morato
authors.affiliation: Laboratory of General Physiology (M.M.-R.) and Laboratory of Pharmacology and Neurobiology (F.F., P.C.), Unit for Multidisciplinary Investigation in Biomedicine, Institute of Biomedical Sciences Abel Salazar, University of Porto; Department of Pharmacology and Therapeutics, Faculty of Medicine, University of Porto (A.L.G., M.F., J.A., P.S., D.M.); Neuropharmacology, Institute of Molecular and Cellular Biology, University of Porto (M.M., D.M.); Center for Drug Discovery and Innovative Medicines, University of Porto (M.M.-R., A.L.G., M.F., J.A., P.S., M.M., F.F., P.C., D.M.); Laboratory of Pharmacology, Department of Drug Sciences, Faculty of Pharmacy, University of Porto and Rede de Química e Tecnologia (REQUIMTE), Porto, Portugal (M.M.); and Burnett School of Biomedical Sciences, College of Medicine, University of Central Florida, Orlando, Florida (S.N.E.).
authors.forename: Fátima
authors.identifier: http://orcid.org/http:orcid.org/0000-0002-0527-1323
authors.initials: F
authors.lastname: Ferreirinha
authors.affiliation: Laboratory of General Physiology (M.M.-R.) and Laboratory of Pharmacology and Neurobiology (F.F., P.C.), Unit for Multidisciplinary Investigation in Biomedicine, Institute of Biomedical Sciences Abel Salazar, University of Porto; Department of Pharmacology and Therapeutics, Faculty of Medicine, University of Porto (A.L.G., M.F., J.A., P.S., D.M.); Neuropharmacology, Institute of Molecular and Cellular Biology, University of Porto (M.M., D.M.); Center for Drug Discovery and Innovative Medicines, University of Porto (M.M.-R., A.L.G., M.F., J.A., P.S., M.M., F.F., P.C., D.M.); Laboratory of Pharmacology, Department of Drug Sciences, Faculty of Pharmacy, University of Porto and Rede de Química e Tecnologia (REQUIMTE), Porto, Portugal (M.M.); and Burnett School of Biomedical Sciences, College of Medicine, University of Central Florida, Orlando, Florida (S.N.E.).
authors.forename: Paulo
authors.initials: P
authors.lastname: Correia-de-Sá
authors.affiliation: Laboratory of General Physiology (M.M.-R.) and Laboratory of Pharmacology and Neurobiology (F.F., P.C.), Unit for Multidisciplinary Investigation in Biomedicine, Institute of Biomedical Sciences Abel Salazar, University of Porto; Department of Pharmacology and Therapeutics, Faculty of Medicine, University of Porto (A.L.G., M.F., J.A., P.S., D.M.); Neuropharmacology, Institute of Molecular and Cellular Biology, University of Porto (M.M., D.M.); Center for Drug Discovery and Innovative Medicines, University of Porto (M.M.-R., A.L.G., M.F., J.A., P.S., M.M., F.F., P.C., D.M.); Laboratory of Pharmacology, Department of Drug Sciences, Faculty of Pharmacy, University of Porto and Rede de Química e Tecnologia (REQUIMTE), Porto, Portugal (M.M.); and Burnett School of Biomedical Sciences, College of Medicine, University of Central Florida, Orlando, Florida (S.N.E.).
authors.forename: Steven N
authors.identifier: http://orcid.org/0000-0002-6114-9189
authors.initials: SN
authors.lastname: Ebert
authors.affiliation: Laboratory of General Physiology (M.M.-R.) and Laboratory of Pharmacology and Neurobiology (F.F., P.C.), Unit for Multidisciplinary Investigation in Biomedicine, Institute of Biomedical Sciences Abel Salazar, University of Porto; Department of Pharmacology and Therapeutics, Faculty of Medicine, University of Porto (A.L.G., M.F., J.A., P.S., D.M.); Neuropharmacology, Institute of Molecular and Cellular Biology, University of Porto (M.M., D.M.); Center for Drug Discovery and Innovative Medicines, University of Porto (M.M.-R., A.L.G., M.F., J.A., P.S., M.M., F.F., P.C., D.M.); Laboratory of Pharmacology, Department of Drug Sciences, Faculty of Pharmacy, University of Porto and Rede de Química e Tecnologia (REQUIMTE), Porto, Portugal (M.M.); and Burnett School of Biomedical Sciences, College of Medicine, University of Central Florida, Orlando, Florida (S.N.E.).
authors.forename: Daniel
authors.identifier: http://orcid.org/0000-0003-3848-3362
authors.initials: D
authors.lastname: Moura
mesh_terms: D058666:Adrenergic beta-2 Receptor Agonists / Q000494:pharmacology*; D058672:Adrenergic beta-2 Receptor Antagonists / Q000494:pharmacology*; D000818:Animals; D001011:Aorta / Q000187:drug effects* / Q000378:metabolism; D002395:Catecholamines / Q000378:metabolism; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D004558:Electric Stimulation / Q000379:methods; D004837:Epinephrine / Q000378:metabolism*; D007545:Isoproterenol / Q000494:pharmacology; D051379:Mice; D018345:Mice, Knockout; D009638:Norepinephrine / Q000378:metabolism; D010625:Phenylethanolamine N-Methyltransferase / Q000378:metabolism; D018343:Receptors, Adrenergic, beta-2 / Q000378:metabolism*; D013726:Terbutaline / Q000494:pharmacology; D014664:Vasodilation / Q000187:drug effects*
chemical_list: D058666:Adrenergic beta-2 Receptor Agonists; D058672:Adrenergic beta-2 Receptor Antagonists; D002395:Catecholamines; D018343:Receptors, Adrenergic, beta-2; D010625:Phenylethanolamine N-Methyltransferase; D007545:Isoproterenol; D013726:Terbutaline; D009638:Norepinephrine; D004837:Epinephrine