Basic characterization of an ouabain-resistant, bumetanide-sensitive K+ carrier-mediated transport system in J774.2 mouse macrophage-like cell line and in variants deficient in adenylate cyclase and cAMP-dependent protein kinase activities

Aline Bourrit, Henri Atlan, Ilana Fromer, Raphael N. Melmed, David Lichtstein*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

13 Scopus citations

Abstract

86Rb(K+) transport across the plasma membrane of macrophage-like cells was studied. The cells used were the wild-type J774.2 and its two variants, CT2 cells, deficient in adenylate cyclase, and J7H1 cells, deficient in cAMP-dependent protein kinase. In the three cell lines about 15% of the total 86Rb(K+) influx is transported by the K+ carrier-mediated transport system. The 86Rb(K+) efflux carried by the same transporter is negligible when measured in the absence of ouabain in the medium. Therefore this carrier conducts a net inward flux of K+ under the experimental conditions used. The transporter is sensitive to extracellular Na+ and inhibited by 'loop' diuretics; bumetanide inhibits ouabain-resistant 86Rb(K+) influx with IC50 of 0.1, 5.0, and 0.05 μM for J774.2, CT2 and J7H1 macrophages, respectively. The membrane potential of the three cells was measured, using the distribution of [3H]tetraphenylphosphonium ([3H]TPP+) across the plasma membrane, and found to be -80.1, -108.5 and -105.1 mV for J774.2, CT2 and J7H1 cells, respectively. The addition of bumetanide to the cell medium does not alter [3H]TPP+ uptake indicating that the transporter is electrically silent. It is concluded that despite the differences in cAMP metabolism by the three macrophages, the basic characteristics of K+ carrier-mediated transport system of the three cells are very similar.

Original languageEnglish
Pages (from-to)85-94
Number of pages10
JournalBiochimica et Biophysica Acta - Biomembranes
Volume817
Issue number1
DOIs
StatePublished - 11 Jul 1985

Keywords

  • (Mouse macrophage)
  • Bumetanide
  • K transport
  • Membrane potential
  • Ouabain resistance
  • Potassium carrier-mediated transport

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