Role of protein kinase C in regulation of Na+- and K +-dependent ATPase activity and pump function in corneal endothelial cells.
OBJECTIVE
Na+- and K+-dependent ATPase (Na,K-ATPase) plays an important role in the pump function of the corneal endothelium. We investigated the possible role of protein kinase C (PKC) in regulation of Na,K-ATPase activity and pump function in corneal endothelial cells.


METHODS
Confluent monolayers of mouse corneal endothelial cells were exposed to phorbol 12,13-dibutyrate (PDBu) to induce activation of PKC. ATPase activity of the cells was evaluated by using ammonium molybdate in spectrophotometric measurement of phosphate released from ATP, with Na,K-ATPase activity being defined as the portion of total ATPase activity sensitive to ouabain. Pump function of the cells was measured with a Ussing chamber, with the pump function attributable to Na,K-ATPase activity being defined as the portion of the total short-circuit current sensitive to ouabain.


RESULTS
PDBu (10(-7) M) increased the Na,K-ATPase activity and pump function of the cultured cells. These effects of PDBu were potentiated by the cyclooxygenase inhibitor indomethacin and the cytochrome P(450) inhibitor resorufin and were blocked by okadaic acid, an inhibitor of protein phosphatases 1 and 2A.


CONCLUSIONS
Our results suggest that PKC bidirectionally regulates Na,K-ATPase activity in mouse corneal endothelial cells: it inhibits Na,K-ATPase activity in a cyclooxygenase- and cytochrome P(450)-dependent manner, whereas it stimulates such activity by activating protein phosphatases 1 or 2A.
Hatou
Shin
S
Division for Vision Research, National Institute of Sensory Organs, National Tokyo Medical Center, Tokyo, Japan. tr97469@zc4.so-net.ne.jp
Yamada
Masakazu
M
Mochizuki
Hiroshi
H
Nishida
Teruo
T
D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D027682:Cation Transport Proteins / Q000502:physiology*; D002478:Cells, Cultured; D004553:Electric Conductivity; D004728:Endothelium, Corneal / Q000166:cytology / Q000187:drug effects* / Q000201:enzymology; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D007213:Indomethacin / Q000494:pharmacology; D051379:Mice; D008809:Mice, Inbred C3H; D019319:Okadaic Acid / Q000494:pharmacology; D010042:Ouabain / Q000494:pharmacology; D010078:Oxazines / Q000494:pharmacology; D015240:Phorbol 12,13-Dibutyrate / Q000494:pharmacology*; D011188:Potassium / Q000378:metabolism; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000502:physiology*; D012964:Sodium / Q000378:metabolism; D000254:Sodium-Potassium-Exchanging ATPase / Q000037:antagonists & inhibitors / Q000378:metabolism*
D027682:Cation Transport Proteins; D004791:Enzyme Inhibitors; D010078:Oxazines; D019319:Okadaic Acid; D015240:Phorbol 12,13-Dibutyrate; D010042:Ouabain; C014180:resorufin; D000255:Adenosine Triphosphate; D012964:Sodium; D011493:Protein Kinase C; D000254:Sodium-Potassium-Exchanging ATPase; D011188:Potassium; D007213:Indomethacin