Epithelial sodium conductance in rabbit preimplantation trophectodermal cells.
We examined the development of epithelial Na+ conductance in 6- and 7-day post coitus (p.c.) preimplantation rabbit embryos using the whole-cell patch-clamp technique on dissociated rabbit trophectodermal cells and by immunocytochemical localization using a polyclonal antibody directed against subunits of an apical epithelial Na+ channel on the intact blastocyst. In Day 6 and 7 p.c. trophectodermal cells, we observed an outwardly rectified whole-cell Na+ current. The current-voltage characteristics did not differ between the 6- and the 7-day p.c. cells. Replacement of Na+ with the impermeant cation N-methyl-D-glucamine in the pipette or bath reduced outward currents and inward currents, respectively, indicating that the current was Na(+)-dependent. Treatment of 7-day p.c. cells with 100 microM amiloride, benzamil, or ethylisopropyl amiloride (EIPA) blocked the whole-cell currents within 5 min. However, the current of the Day 6 p.c. embryo was not blocked by amiloride. The amiloride block at Day 7 p.c. was only partially reversible after 15 min of continuous perfusion of the bath with an amiloride-free solution. The apparent dissociation constant (Ki) for amiloride, benzamil, and EIPA was 12, 50, and 16 microM, respectively, when measured 5 min after drug addition. Immunolocalization studies of blastocysts with a polyclonal antibody raised against a high amiloride affinity Na+ channel isolated from bovine kidney revealed no specific binding to the trophectodermal cells at Day 6 p.c.(ABSTRACT TRUNCATED AT 250 WORDS)
Robinson
D H
DH
Department of Physiology and Biophysics, University of Alabama, Birmingham 35294.
Bubien
J K
JK
Smith
P R
PR
Benos
D J
DJ
D000584:Amiloride / Q000031:analogs & derivatives / Q000494:pharmacology; D000818:Animals; D001693:Biological Transport, Active / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D004553:Electric Conductivity / Q000187:drug effects; D007150:Immunohistochemistry; D066298:In Vitro Techniques; D008536:Meglumine / Q000494:pharmacology; D011817:Rabbits / Q000196:embryology*; D012964:Sodium / Q000493:pharmacokinetics*; D015222:Sodium Channels / Q000502:physiology; D014327:Trophoblasts / Q000502:physiology*
D015222:Sodium Channels; C013407:benzamil; D008536:Meglumine; D000584:Amiloride; D012964:Sodium; C039614:ethylisopropylamiloride