Potassium transport in nonpigmented epithelial cells of ocular ciliary body: Inhibition of a Na+, K+, Cl− cotransporter by protein kinase C

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Abstract
The mechanism by which 86Rb+ (used as a tracer for K+) enters human nonpigmented ciliary epithelial cells were investigated. Ouabain‐inhibitable bumetanide‐insensitive 86Rb+ transport accounted for ∼70–80% of total, whereas bumetanide‐inhibitable ouabain‐insensitive uptake accounted for 15–25% of total. K+ channel blockers such as BaCl2 reduced uptake by ∼5%. Bumetanide inhibited 86Rb+ uptake with an IC50 of 0.5 μM, while furosemide inhibited with an IC50 of about 20 μM. Bumetanide‐inhibitable 86Rb+ uptake was reduced in Na+‐free or Cl−‐free media, suggesting that Na+ and Cl− were required for optimal uptake via this mechanism. These characteristics are consistent with a Na+, K+, Cl− contransporter in NPE cells. Treatment of NPE cells for 15 min with phorbol 12‐myristate, 13‐acetate (PMA), an activator of protein kinase C, caused a 50–70% decrease in 86Rb+ uptake via the Na+, K+, Cl− cotransporter. Other 86Rb+ uptake mechanisms were not affected. 86Rb+ uptake via the Na+, K+, Cl− contransporter could be inhibited by other phorbol esters and by dioctanoylglycerol, an analog of diacylglycerol, but not by 4α phorbol didecanoate, an ineffective activator of protein kinase C. Staurosporine, a protein kinase C inhibitor, blocked phorbol ester inhibition of 86Rb+ uptake. These data suggest that a Na+, K+, Cl− cotransporter in NPE cells is inhibited by activation of protein kinase C.

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