Aquaporin-1 is recruited to the plasma membrane by hyperosmotic stimuli via a protein kinase A-dependent pathway in rat peritoneal mesothelial cells.
Aquaporin-1 (AQP1) has been reported to play an important role in water permeability in peritoneal dialysis. To determine the mechanism involved in this process, we used cultured rat peritoneal mesothelial cells (RPMCs) to examine the glucose-induced translocation of AQP1 to the plasma membrane. Cultured RPMCs obtained from male Sprague-Dawley rats were incubated in a combination of Dulbecco modified Eagle medium (DMEM) and F12 medium at 37 degrees C for 15 minutes. The plasma membrane of the RPMCs was separated by Percoll gradient, and the quantity of AQP1 in the membrane fraction was determined by Western blot analysis. The amount of AQP1 was significantly increased by the addition of 5% glucose (139.5% +/- 38.7% of control, p < 0.05) or of dibutyryl cyclic adenosine monophosphate (db-cAMP), a cAMP analog to the medium (139.5% +/- 21.9% of control, p < 0.05). However glucose-induced enhancement of AQP1 disappeared with the addition of H-89, a protein kinase A (PKA)--specific inhibitor (103% +/- 17.5% of control, p < 0.05 as compared with 5% glucose). We also examined the effect of 5% glucose on PKA activity separately in the cytosol fraction, the crude membrane fraction, and the pure plasma membrane fraction. In the cytosol fraction of 5% glucose-stimulated RPMCs, PKA activity was decreased (70.5% +/- 11.5% of control, p < 0.01), but in the crude membrane fraction, it was significantly increased (143.9% +/- 52.9% of control, p < 0.01). In the pure plasma membrane fraction, PKA activity did not change. From those findings, we hypothesize that 5% glucose augments the PKA-dependent translocation of AQP1 to the plasma membrane, mediated by PKA translocation to the intracellular AQP1 store.
Sakurada
Tsutomu
T
St. Marianna University School of Medicine, Division of Nephrology and Hypertension, Kawasaki, Japan.
Kuboshima
Shingo
S
Ogimoto
Goichi
G
Fujino
Tomoya
T
Sato
Takeo
T
Maeba
Teruhiko
T
Kimura
Kenjiro
K
D000818:Animals; D051398:Aquaporin 1; D020346:Aquaporins / Q000378:metabolism*; D015153:Blotting, Western; D002462:Cell Membrane / Q000378:metabolism*; D002478:Cells, Cultured; D017868:Cyclic AMP-Dependent Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D003995:Dibutyryl Cyclic GMP / Q000494:pharmacology; D004789:Enzyme Activation; D004848:Epithelium / Q000648:ultrastructure; D005947:Glucose / Q000494:pharmacology; D007546:Isoquinolines / Q000494:pharmacology; D008297:Male; D009994:Osmolar Concentration; D010537:Peritoneum / Q000378:metabolism* / Q000648:ultrastructure; D051381:Rats; D017207:Rats, Sprague-Dawley; D013449:Sulfonamides
C496022:Aqp1 protein, rat; D020346:Aquaporins; D007546:Isoquinolines; D013449:Sulfonamides; D051398:Aquaporin 1; D003995:Dibutyryl Cyclic GMP; D017868:Cyclic AMP-Dependent Protein Kinases; D005947:Glucose; C063509:N-(2-(4-bromocinnamylamino)ethyl)-5-isoquinolinesulfonamide