Ouabain-sensitive Na+-K+ ATPase pumps in cultured human retinal pigment epithelium

The expression of Na+-K+ ATPase pumps was studied in cultured human retinal pigment epithelium (RPE). Pump site number was measured by quantitation of the specific binding of [3H]ouabain to cultures of varying density. Specific binding of [3H]ouabain was time- and concentration-dependent, and was inhibited by potassium and by excess unlabeled ouabain. Estimates of pump site number based upon specific [3H]ouabain binding indicated that the number of pumps per RPE cell was maximal in sparse cultures and declined six-fold as cultures became confluent. Pump activity, determined by measurement of specific 86Rb (rubidium) uptake, was also greater in sparse than in dense cultures. Quantitation of [3H]thymidine incorporation as a measure of cell proliferation demonstrated that proliferation in RPE cultures decreased logarithmically as culture density increased. Increased pump site number in cultured RPE, therefore, correlated with increased cell proliferation and decreased culture density. We conclude that human RPE express ouabain-sensitive Na+-K+ ATPase in vitro and maximal expression is observed in sparse, proliferating cultures.