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J.M. Sanchez, P. Iserovich, K. Kuang, J. Fischbarg; Fluid Transport by Opossum Kidney Cell Layers, and the Effects of Translayer Electrical Currents . Invest. Ophthalmol. Vis. Sci. 2003;44(13):3465.
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Purpose. We have presented evidence suggesting that endothelial fluid transport results from electro-osmosis across intercellular junctions. As the endothelium is a secretory epithelium, for the present study, we tested our theory using a putative absorptive epithelium, the one formed by Opossum kidney epithelium cells, which are gaining acceptance as a model for kidney proximal tubule. Methods. Opossum Kidney Cells (OKC) (ATCC Catalog # CRL-1840) were grown to confluence on 24 mm permeable membrane inserts (Transwell Costar #3450). We utilized MEM with Earle’s salts, 25mM HEPES and NaHCO3, 10ml of L-Glutamine [4 mM] (SIGMA Product # M7278), 6% FBS, penicillin (100 U/ml) and streptomycin (100 ng/ml). The inserts with the confluent cell layers were supported by a stainless steel mesh and the rate of fluid movement was determined by volume clamp using a nanoinjector. Electrical current was sent across the preparations following the following convention: positive current (+) went from apical to basal, and vice-versa for negative current (-). Results. We found that the layers transported fluid in the apical to basal direction, as would be expected from proximal tubule layers. The transport rates varied from 3 to 20 µL/(h cm2). Up to 70 % of the fluid movements through the preparations could be inhibited by ouabain [0.5 mM, 3 inserts]. Importantly, fluid movement was decreased by passing "positive" current, and was increased by passing negative current. Current values used went from ±50 to ±250 µAmp, and the same pattern of effects was observed at all current values (6 inserts). Conclusions. Although the OCK cells behave in many respects as proximal kidney, fluid transport across them had apparently not been demonstrated until this study. Our results thus reaffirm that these cells constitute a valid model for kidney proximal tubule. As for the effects of currents, to be noted, the negative current goes in a direction opposite to that of fluid transport and stimulates it, and vice-versa. Although local osmosis or an osmotic artifact cannot be excluded, the simplest explanation for the effects found is that the imposed currents induce electro-osmotic fluid movements across putative anion selective junctions.
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