Na(+)-K(+)-2Cl- cotransport, Na+/H+ exchange, and cell volume in ferret erythrocytes.
Ferrets have high-Na+ and low-K+ erythrocytes (113 and 5.4 mmol/l cell water) due to the lack of Na(+)-K+ pumps. Because ferret erythrocytes have a high capacity for Na(+)-K(+)-2Cl- cotransport, the present study was undertaken to evaluate cell volume-related changes in cotransport activity and its role in volume regulation. With cell shrinkage, Na(+)-K(+)-2Cl- cotransport is activated about twofold. A large bumetanide-insensitive Na+ uptake component that has not yet been described is found in shrunken erythrocytes. Its inhibition by amiloride (concn inhibiting 50% of maximal response = 12 microM) and the Na+ dependence of amiloride-sensitive extracellular pH changes measured in cells suspended in hypertonic unbuffered medium indicate that this flux represents Na+/H+ exchange. Shrinkage activation of both transporters follows a time lag of approximately 3 min and also requires normal levels of ATP. ATP depletion inhibits Na(+)-K(+)-2Cl- cotransport even at normal cell volume. Both transporters are partially inhibited by the protein kinase inhibitors staurosporine and K252a, and activators of protein kinases A and C do not affect transport. Okadaic acid inhibition of protein phosphatases activates Na(+)-K(+)-2Cl- cotransport to its maximal activity (same after shrinkage), but shrinkage and okadaic acid activation are not additive. In contrast, okadaic acid activates Na+/H+ exchange even in shrunken cells. These results indicate that cell shrinkage activates Na(+)-K(+)-2Cl- cotransport and Na+/H+ exchange probably by phosphorylation processes.
Mairbäurl
H
H
Department of Sports Medicine, University of Heidelberg, Germany.
Herth
C
C
D000255:Adenosine Triphosphate / Q000097:blood; D000818:Animals; D002034:Bumetanide / Q000494:pharmacology; D002227:Carbazoles / Q000494:pharmacology; D002352:Carrier Proteins / Q000097:blood*; D002712:Chlorides / Q000097:blood; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004910:Erythrocyte Membrane / Q000378:metabolism*; D004912:Erythrocytes / Q000166:cytology / Q000187:drug effects / Q000502:physiology*; D005289:Ferrets; D066298:In Vitro Techniques; D026121:Indole Alkaloids; D007700:Kinetics; D008297:Male; D008565:Membrane Proteins / Q000097:blood; D011188:Potassium / Q000097:blood; D047428:Protein Kinase Inhibitors; D012964:Sodium / Q000097:blood*; D017923:Sodium-Hydrogen Exchangers / Q000097:blood*; D028021:Sodium-Potassium-Chloride Symporters; D019311:Staurosporine / Q000494:pharmacology
D002227:Carbazoles; D002352:Carrier Proteins; D002712:Chlorides; D004791:Enzyme Inhibitors; D026121:Indole Alkaloids; D008565:Membrane Proteins; D047428:Protein Kinase Inhibitors; D017923:Sodium-Hydrogen Exchangers; D028021:Sodium-Potassium-Chloride Symporters; D002034:Bumetanide; D000255:Adenosine Triphosphate; C049985:staurosporine aglycone; D012964:Sodium; D019311:Staurosporine; D011188:Potassium