Isoform-specific function and distribution of Na/K pumps in the frog lens epithelium.
Epithelial cells from the anterior and equatorial surfaces of the frog lens were isolated and used the same day for studies of the Na/K ATPase. RNase protection assays showed that all cells express alpha(1)- and alpha(2)-isoforms of the Na/K pump but not the alpha(3)-isoform, however the alpha(2)-isoform dominates in anterior cells whereas the alpha(1)-isoform dominates in equatorial cells. The whole cell patch-clamp technique was used to record functional properties of the Na/K pump current (I(P)), defined as the current specifically inhibited by dihydro-ouabain (DHO). DHO-I(P) blockade data indicate the alpha(1)-isoform has a dissociation constant of 100 microm DHO whereas for the alpha(2)-isoform it is 0.75 microm DHO. Both alpha(1)- and alpha(2)-isoforms are half maximally activated at an intracellular Na(+)-concentration of 9 mm. The alpha(1)-isoform is half maximally activated at an extracellular K(+)-concentration of 3.9 mm whereas for the alpha(2)-isoform, half maximal activation occurs at 0.4 mm. Lastly, transport by the alpha(1)-isoform is inhibited by a drop in extracellular pH, which does not affect transport by the alpha(2)-isoform. Under normal physiological conditions, I(P) in equatorial cells is approximately 0.23 microA/microF, and in anterior cells it is about 0.14 microA/microF. These current densities refer to the area of cell membrane assuming a capacitance of around 1 microF/cm(2). Because cell size and geometry are different at the equatorial vs. anterior surface of the intact lens, we estimate Na/K pump current density per area of lens surface to be around 10 microA/cm(2) at the equator vs. 0.5 microA/cm(2) at the anterior pole.
Gao
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J
Department of Physiology & Biophysics, SUNY at Stony Brook, NY 11794-8661, USA.
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Wymore
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D000595:Amino Acid Sequence; D000818:Animals; D004553:Electric Conductivity; D004848:Epithelium / Q000187:drug effects / Q000201:enzymology; D006863:Hydrogen-Ion Concentration; D017136:Ion Transport; D007908:Lens, Crystalline / Q000166:cytology / Q000187:drug effects / Q000201:enzymology*; D008954:Models, Biological; D008969:Molecular Sequence Data; D023282:Nuclease Protection Assays; D010042:Ouabain / Q000031:analogs & derivatives* / Q000494:pharmacology; D018408:Patch-Clamp Techniques; D020033:Protein Isoforms / Q000037:antagonists & inhibitors / Q000235:genetics / Q000502:physiology; D012333:RNA, Messenger / Q000096:biosynthesis; D011894:Rana pipiens; D016415:Sequence Alignment; D000254:Sodium-Potassium-Exchanging ATPase / Q000037:antagonists & inhibitors / Q000235:genetics / Q000502:physiology*
D020033:Protein Isoforms; D012333:RNA, Messenger; C000977:dihydroouabain; D010042:Ouabain; D000254:Sodium-Potassium-Exchanging ATPase