title: Calcium-dependent inhibition of synaptosomal serotonin transport by the alpha 2-adrenoceptor agonist 5-bromo-N-[4,5-dihydro-1H-imidazol-2-yl]-6-quinoxalinamine (UK14304).
abstract: Termination of serotonergic transmission is the function of the plasma membrane 5-hydroxytryptamine (serotonin, 5-HT) transporter (SERT), which is also a high-affinity target in vivo for antidepressants, amphetamines, and cocaine. Studies show that SERT is regulated by protein kinase- and phosphataselinked pathways. In contrast, receptor-linked modulation of SERT is only minimally defined. Because noradrenergic stimulation is reported to influence 5-HT release, we explored possible presynaptic adrenoceptor-mediated regulation of SERT. In mouse forebrain synaptosomes, alpha2-adrenoceptor agonists, particularly 5-bromo-N-[4,5-dihydro-1H-imidazol-2-yl]-6-quinoxalinamine (UK14304), triggered a concentration- and time-dependent decrease in 5-HT transport. In contrast, 5-HT uptake was unaffected by pharmacological alpha1-adrenoceptor activation. Kinetically, UK14304 significantly decreased the apparent substrate affinity, Km without altering transport capacity, Vmax. At concentrations of UK14304 supporting maximal inhibition of SERT in synaptosomes, no effect on SERT in transfected cells was observed, suggesting that UK14304 acts indirectly to reduce SERT activity. The effect of UK14304 on 5-HT uptake was not shared by other Na+ and Cl--dependent transporters. UK14304-mediated inhibition of SERT function was yohimbine-sensitive, as was inhibition triggered by norepinephrine, and was abolished in the absence of added Ca2+. Moreover, UK14304 effects were attenuated by voltage-sensitive Ca2+ channel antagonists, consistent with a role for Ca2+ in UK14304 effects. In agreement with altered 5-HT transport activity in vitro, in vivo chronoamperometry studies revealed that UK14304 significantly prolonged 5-HT clearance. Our findings suggest that UK14304 modulates SERT function in vitro and in vivo via signaling pathways, possibly supported by an influx of Ca2+ through voltage-sensitive Ca2+ channels.
authors.affiliation: Department of Pharmacology, Meharry Medical College, 1005 D.B. Todd Blvd., Nashville, TN 37208, USA. tansah@mmc.edu
authors.forename: Twum-Ampofo
authors.initials: TA
authors.lastname: Ansah
authors.forename: Sammanda
authors.initials: S
authors.lastname: Ramamoorthy
authors.forename: Sylvia
authors.initials: S
authors.lastname: Montañez
authors.forename: Lynette C
authors.initials: LC
authors.lastname: Daws
authors.forename: Randy D
authors.initials: RD
authors.lastname: Blakely
mesh_terms: D058647:Adrenergic alpha-2 Receptor Agonists; D000316:Adrenergic alpha-Agonists / Q000494:pharmacology*; D000818:Animals; D001692:Biological Transport / Q000187:drug effects; D000068438:Brimonidine Tartrate; D019556:COS Cells; D002118:Calcium / Q000378:metabolism; D002352:Carrier Proteins / Q000187:drug effects / Q000378:metabolism; D008297:Male; D008562:Membrane Glycoproteins / Q000187:drug effects / Q000378:metabolism; D026901:Membrane Transport Proteins; D051379:Mice; D008810:Mice, Inbred C57BL; D009419:Nerve Tissue Proteins; D011810:Quinoxalines / Q000494:pharmacology*; D012701:Serotonin / Q000378:metabolism*; D050486:Serotonin Plasma Membrane Transport Proteins; D013574:Synaptosomes / Q000187:drug effects* / Q000378:metabolism
chemical_list: D058647:Adrenergic alpha-2 Receptor Agonists; D000316:Adrenergic alpha-Agonists; D002352:Carrier Proteins; D008562:Membrane Glycoproteins; D026901:Membrane Transport Proteins; D009419:Nerve Tissue Proteins; D011810:Quinoxalines; D050486:Serotonin Plasma Membrane Transport Proteins; C493231:Slc6a4 protein, mouse; D012701:Serotonin; D000068438:Brimonidine Tartrate; D002118:Calcium