Epithelial sodium channel in a human trophoblast cell line (BeWo).
The present study was performed to assay sodium currents in BeWo cells. These cells comprise a human trophoblast cell line which displays many of the biochemical and morphological properties similar to those reported for the in uterus proliferative cytotrophoblast. For whole-cell patch-clamp experiments, BeWo cells treated for 12 h with 100 nM aldosterone were exposed to 8Br-cAMP, a membrane-permeable cAMP analogue, to induce channel activity. Cells showed an amiloride-sensitive ion current (IC50 of 5.77 microM). Ion substitution experiments showed that the amiloride-sensitive current carried cations with a permeability rank order of Li+ > Na+ > K+ > NMDG (PLi/PNa = 1.3, PK/PNa = 0.6, PNMDG/PNa = 0.2). In cells pretreated with aldosterone, we observed that nearly half of successful patches had sodium channels with a linear conductance of 6.4 +/- 1.8 pS, a low voltage-independent Po and a PK/PNa of 0.19. Using RT-PCR, we determined that control cells express the alpha-, but not beta- and gamma-, epithelial sodium channel (ENaC) mRNA. When cells were treated with aldosterone (100 nM, 12 h), all alpha-, beta- and gamma-ENaC mRNAs were detected. The presence of ENaC subunit proteins in these cells was confirmed by Western blot analysis and immunolocalization with specific ENaC primary antibodies. In summary, our results suggest that BeWo cells express ENaC subunits and that aldosterone was able to modulate a selective response by generating amiloride-sensitive sodium currents similar to those observed in other human tissues.
del Mónaco
Silvana
S
Laboratorio de Neurofisiología, Instituto de Investigaciones Médicas A. Lanari, University of Buenos Aires, C. de Malvinas 3150, Buenos Aires 1427, Argentina.
Assef
Yanina
Y
Kotsias
Basilio A
BA
D015124:8-Bromo Cyclic Adenosine Monophosphate / Q000494:pharmacology; D000450:Aldosterone / Q000502:physiology; D000584:Amiloride / Q000494:pharmacology; D045744:Cell Line, Tumor; D062686:Epithelial Sodium Channel Blockers; D053503:Epithelial Sodium Channels / Q000235:genetics / Q000378:metabolism*; D005260:Female; D006801:Humans; D008564:Membrane Potentials / Q000187:drug effects / Q000235:genetics; D018408:Patch-Clamp Techniques; D012333:RNA, Messenger / Q000032:analysis / Q000037:antagonists & inhibitors; D026941:Sodium Channel Blockers / Q000494:pharmacology; D014327:Trophoblasts / Q000187:drug effects / Q000378:metabolism*
D062686:Epithelial Sodium Channel Blockers; D053503:Epithelial Sodium Channels; D012333:RNA, Messenger; C507045:SCNN1A protein, human; C507048:SCNN1B protein, human; C507052:SCNN1G protein, human; D026941:Sodium Channel Blockers; D015124:8-Bromo Cyclic Adenosine Monophosphate; D000450:Aldosterone; D000584:Amiloride