Genistein inhibits the regulation of active sodium-potassium transport by dopaminergic agonists in nonpigmented ciliary epithelium

PURPOSE. To determine whether dopamine receptor stimulation regulates Na,K-ATPase-mediated ion transport in cultured nonpigmented ciliary epithelium (NPE). METHODS. Using a rabbit NPE cell line, active Na-K transport activity was determined by measuring ouabain-sensitive potassium (⁸⁶Rb) uptake in cell monolayers. Western blot analysis of membrane material obtained from cell homogenates was conducted to examine tyrosine phosphorylation of membrane proteins. RESULTS. Ouabain-sensitive potassium (⁸⁶Rb) uptake was inhibited in the presence of either dopa- mine or the D₁-selective agonist SKF82958. The response was suppressed by SCH23390, a D₁ antagonist, but not by sulpiride, a D₂-selective antagonist. Quinpirole, a D₂-selective agonist, did not cause inhibition of ouabain-sensitive potassium (⁸⁶Rb) uptake. Cyclic adenosine monophosphate (cAMP) was detectably increased in SKF82958-treated cells, although the concentration of SKF required to elevate cell cAMP was higher than the concentration needed to inhibit ouabain-sensitive potassium (⁸⁶Rb) uptake. The protein kinase A inhibitor H89 prevented the ⁸⁶Rb uptake response to SKF82958. Genistein, an inhibitor of tyrosine kinases, also prevented the ⁸⁶Rb uptake response to SKF82958. Membrane material isolated from cells exposed to SKF82958 showed an increase in the density of several phosphotyrosine bands. These changes in phosphotyrosine immunoblot density were not observed in material isolated from cells that received either genistein or SCH23390 before SKF82958 treatment. CONCLUSIONS. The results of this study suggest D₁ agonists cause a reduction of Na,K-ATPase-mediated ion transport by a mechanism that could involve a tyrosine kinase step.