title: Radiotracers for cardiac sympathetic innervation: transport kinetics and binding affinities for the human norepinephrine transporter.
abstract: BACKGROUND
  Most radiotracers for imaging of cardiac sympathetic innervation are substrates of the norepinephrine transporter (NET). The goal of this study was to characterize the NET transport kinetics and binding affinities of several sympathetic nerve radiotracers, including [(11)C]-(-)-meta-hydroxyephedrine, [(11)C]-(-)-epinephrine, and a series of [(11)C]-labeled phenethylguanidines under development in our laboratory. For comparison, the NET transport kinetics and binding affinities of some [(3)H]-labeled biogenic amines were also determined.
  
  
  METHODS
  Transport kinetics studies were performed using rat C6 glioma cells stably transfected with the human norepinephrine transporter (C6-hNET cells). For each radiolabeled NET substrate, saturation transport assays with C6-hNET cells measured the Michaelis-Menten transport constants Km and Vmax for NET transport. Competitive inhibition binding assays with homogenized C6-hNET cells and [(3)H]mazindol provided estimates of binding affinities (KI) for NET.
  
  
  RESULTS
  Km, Vmax and KI values were determined for each NET substrate with a high degree of reproducibility. Interestingly, C6-hNET transport rates for 'tracer concentrations' of substrate, given by the ratio Vmax/Km, were found to be highly correlated with neuronal transport rates measured previously in isolated rat hearts (r(2)=0.96). This suggests that the transport constants Km and Vmax measured using the C6-hNET cells accurately reflect in vivo transport kinetics.
  
  
  CONCLUSIONS
  The results of these studies show how structural changes in NET substrates influence NET binding and transport constants, providing valuable insights that can be used in the design of new tracers with more optimal kinetics for quantifying regional sympathetic nerve density.
authors.affiliation: Division of Nuclear Medicine, Department of Radiology, 2276 Medical Sciences I Building, University of Michigan Medical School, 1301 Catherine Street, SPC 5610, Ann Arbor, MI 48109, USA. raffel@umich.edu
authors.forename: David M
authors.initials: DM
authors.lastname: Raffel
authors.forename: Wei
authors.initials: W
authors.lastname: Chen
authors.forename: Yong-Woon
authors.initials: YW
authors.lastname: Jung
authors.forename: Keun Sam
authors.initials: KS
authors.lastname: Jang
authors.forename: Guie
authors.initials: G
authors.lastname: Gu
authors.forename: Nicholas V
authors.initials: NV
authors.lastname: Cozzi
mesh_terms: D000818:Animals; D001692:Biological Transport; D045744:Cell Line, Tumor; D004809:Ephedrine / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism; D004837:Epinephrine / Q000737:chemistry / Q000378:metabolism*; D006321:Heart / Q000294:innervation*; D006801:Humans; D007700:Kinetics; D050484:Norepinephrine Plasma Membrane Transport Proteins / Q000378:metabolism*; D049268:Positron-Emission Tomography; D011485:Protein Binding; D011849:Radioactive Tracers; D051381:Rats; D013329:Structure-Activity Relationship; D013564:Sympathetic Nervous System / Q000000981:diagnostic imaging*
chemical_list: D050484:Norepinephrine Plasma Membrane Transport Proteins; D011849:Radioactive Tracers; C065363:3-hydroxyephedrine; D004809:Ephedrine; D004837:Epinephrine