title: Neuropeptide Y and the retinal pigment epithelium: receptor subtypes, signaling, and bioelectrical responses.
abstract: OBJECTIVE
  To characterize the potential for neuropeptide Y (NPY) signaling in the retinal pigment epithelium (RPE) by identifying the NPY receptor subtypes present, determining the effect of NPY on second-messenger production and membrane conductance, and establishing the neural retina as a site of NPY gene expression.
  
  
  METHODS
  Neuropeptide Y receptors present in bovine and human RPE were identified using ribonuclease protection assays and reverse transcriptase-coupled polymerase chain reaction. Assays of cyclic adenosine monophosphate (cAMP) and inositol phosphate production were performed using anion exchange chromatography and RPE cultures labeled with tritiated adenine or myo-inositol, respectively. Open-circuit recordings of transepithelial potential and resistance were performed using intact bovine RPE-choroid preparations. Neuropeptide Y-expressing cells in the retina were identified by staining for beta-galactosidase activity in eyes from mice in which lacZ replaces a portion of the NPY gene.
  
  
  RESULTS
  Human RPE contained transcripts encoding Y1, Y2, and Y5 receptors, the predominant subtypes present in the central nervous system. Bovine RPE contained transcripts encoding Y2 receptors but not Y1 receptors. However, cultured cells contained transcripts encoding Y1 and Y2 receptors. Neuropeptide Y signaling in cultured bovine RPE occurred predominately through the Y2 receptor subtype, because nanomolar amounts of NPY and NPY13-36, but not [Leu31,Pro34]NPY, significantly inhibited isoproterenol-induced cAMP accumulation. Apical application of NPY increased the transepithelial potential in RPE-choroid preparations. This response was greatly diminished after basolateral membrane Cl- channels were blocked or changes in intracellular Ca2+ concentration were prevented with a Ca2+ chelator. The NPY gene was expressed in amacrine cells of the inner nuclear and ganglion cell layers of the mouse retina.
  
  
  CONCLUSIONS
  The discovery of functionally coupled NPY receptors in the RPE represents the identification of a novel site of expression of this receptor family. The effects of NPY on the electrophysiologic properties of the bovine RPE are consistent with a potential paracrine role in regulating basolateral membrane Ca2+-sensitive Cl- conductance linked to Cl- and fluid transport.
authors.affiliation: Department of Biological Chemistry, The University of Michigan Medical School, Ann Arbor 48105-0714, USA.
authors.forename: D A
authors.initials: DA
authors.lastname: Ammar
authors.forename: B A
authors.initials: BA
authors.lastname: Hughes
authors.forename: D A
authors.initials: DA
authors.lastname: Thompson
mesh_terms: D000818:Animals; D001485:Basement Membrane; D015220:Calcium Channels / Q000378:metabolism; D002417:Cattle; D002478:Cells, Cultured; D018118:Chloride Channels / Q000378:metabolism; D000242:Cyclic AMP / Q000378:metabolism; D004553:Electric Conductivity; D015870:Gene Expression; D006801:Humans; D007295:Inositol Phosphates / Q000378:metabolism; D008564:Membrane Potentials / Q000187:drug effects; D009478:Neuropeptide Y / Q000235:genetics / Q000494:pharmacology*; D010857:Pigment Epithelium of Eye / Q000187:drug effects / Q000378:metabolism*; D016133:Polymerase Chain Reaction; D012194:RNA-Directed DNA Polymerase; D017476:Receptors, Neuropeptide Y / Q000145:classification / Q000378:metabolism*; D012160:Retina / Q000187:drug effects / Q000378:metabolism; D015398:Signal Transduction; D001616:beta-Galactosidase / Q000378:metabolism
chemical_list: D015220:Calcium Channels; D018118:Chloride Channels; D007295:Inositol Phosphates; D009478:Neuropeptide Y; D017476:Receptors, Neuropeptide Y; D000242:Cyclic AMP; D012194:RNA-Directed DNA Polymerase; D001616:beta-Galactosidase