Characterization of adenosine A1 receptor in a cell line (28A) derived from rabbit collecting tubule

W. S. Spielman, K. N. Klotz, L. J. Arend, B. A. Olson, D. G. LeVier, U. Schwabe

Research output: Contribution to journalArticlepeer-review

25 Scopus citations


We have previously reported that in several renal cell types, adenosine receptor agonists inhibit adenylyl cyclase and activate phospholipase C via a pertussis toxin-sensitive G protein. In the present study, in 28A cells, both of these adenosine receptor-mediated responses were inhibited by 8- cyclopentyl-1,3-dipropylxanthine (DPCPX), a highly selective A1 adenosine receptor antagonist. The binding characteristics of the adenosine A1 receptor in the 28A renal cell line were studied using the radiolabeled antagonist [3H]DPCPX to determine whether two separate binding sites could account for these responses. Saturation binding of [3H]DPCPX to 28A cell membranes revealed a single class of A1 binding sites with an apparent K(d) value of 1.4 nM and maximal binding capacity of 64 fmol/mg protein. Competition experiments with a variety of adenosine agonists gave biphasic displacement curves with a pharmacological profile characteristic of A1 receptors. Comparison of [3H]DPCPX competition binding data from 28A cell membranes with rabbit brain membranes, a tissue with well-characterized A1 receptors, reveals that the A1 receptor population in 28A cells has similar agonist binding affinities to the receptor population in brain but has a considerably lower density. Addition of guanosine 5'-triphosphate (100 μM) to 28A cell membranes caused the competition curves to shift from biphasic to monophasic, indicating that the A1 receptors exist in two interconvertible affinity states because of their couplinng to G proteins. In the absence of evidence for subpopulations of the A1 receptor, it appears that in 28A cells, a single A1 receptor population, as defined by ligand binding characteristics, couples via one or more pertussis toxin-sensitive guanine nucleotide binding proteins to two different biological signaling mechanisms.

Original languageEnglish (US)
Pages (from-to)C502-C508
JournalAmerican Journal of Physiology - Cell Physiology
Issue number2 32-2
StatePublished - 1992


  • G proteins
  • adenosine 3',5'-cyclic monophosphate
  • calcium
  • phosphoinositides
  • receptor binding
  • signal transduction

ASJC Scopus subject areas

  • Physiology
  • Cell Biology


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