Alteration of active Na-K transport on protein kinase C activation in cultured ciliary epithelium

Purpose. Experiments were conducted to test whether protein kinase C activation causes changes in active sodium-potassium transport in an established SV-10 transformed line (ODM2) of cultured human nonpigmented ciliary epithelial cells. Methods. Rubidium-86 (⁸⁶Rb) uptake was measured and the data used to determine the rate of potassium entry into the cells. Results. Protein kinase C activator, phorbol dibutyrate (PDBu), caused a stimulation of ouabain-sensitive ⁸⁶Rb uptake. Inhibition of protein kinase C by 1-(5-isoquinolinylsulfonyl) 2-methylpiperazine (H-7), or down-regulation of protein kinase C activation by prolonged exposure of PDBu, decreased the PDBu response. These results suggest that protein kinase C plays a role in Na-K pump activation. The Na⁺/H⁺ exchanger inhibitor, amiloride, also reduced the stimulation of the ouabain-sensitive ⁸⁶Rb uptake by PDBu. ⁸⁶Rb efflux was not altered by protein kinase C activation. At the same time that PDBu increased the ouabain-sensitive ⁸⁶Rb uptake, it also decreased the ouabain-insensitive ⁸⁶Rb uptake. The ouabain-insensitive ⁸⁶Rb uptake component could be inhibited by bumetanide, suggesting that protein kinase C activation decreases the activity of a Na/K/2Cl cotransporter. Conclusions. These findings suggest that activation of protein kinase C may stimulate Na,K-ATPase activity mainly by a mechanism involving increased Na⁺ influx mediated by the Na⁺/H⁺ exchanger.