TY - JOUR
T1 - 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
AU - Bourrit, Aline
AU - Atlan, Henri
AU - Fromer, Ilana
AU - Melmed, Raphael N.
AU - Lichtstein, David
PY - 1985/7/11
Y1 - 1985/7/11
N2 - 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.
AB - 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.
KW - (Mouse macrophage)
KW - Bumetanide
KW - K transport
KW - Membrane potential
KW - Ouabain resistance
KW - Potassium carrier-mediated transport
UR - http://www.scopus.com/inward/record.url?scp=84886639026&partnerID=8YFLogxK
U2 - 10.1016/0005-2736(85)90071-9
DO - 10.1016/0005-2736(85)90071-9
M3 - ???researchoutput.researchoutputtypes.contributiontojournal.article???
AN - SCOPUS:84886639026
SN - 0005-2736
VL - 817
SP - 85
EP - 94
JO - Biochimica et Biophysica Acta - Biomembranes
JF - Biochimica et Biophysica Acta - Biomembranes
IS - 1
ER -