title: Activation of Na+/H+ and K+/H+ exchange by calyculin A in Amphiuma tridactylum red blood cells: implications for the control of volume-induced ion flux activity.
abstract: Alteration in cell volume of vertebrates results in activation of volume-sensitive ion flux pathways. Fine control of the activity of these pathways enables cells to regulate volume following osmotic perturbation. Protein phosphorylation and dephosphorylation have been reported to play a crucial role in the control of volume-sensitive ion flux pathways. Exposing Amphiuma tridactylu red blood cells (RBCs) to phorbol esters in isotonic medium results in a simultaneous, dose-dependent activation of both Na(+)/H(+) and K(+)/H(+) exchangers. We tested the hypothesis that in Amphiuma RBCs, both shrinkage-induced Na(+)/H(+) exchange and swelling-induced K(+)/H(+) exchange are activated by phosphorylation-dependent reactions. To this end, we assessed the effect of calyculin A, a phosphatase inhibitor, on the activity of the aforementioned exchangers. We found that exposure of Amphiuma RBCs to calyculin-A in isotonic media results in simultaneous, 1-2 orders of magnitude increase in the activity of both K(+)/H(+) and Na(+)/H(+) exchangers. We also demonstrate that, in isotonic media, calyculin A-dependent increases in net Na(+) uptake and K(+) loss are a direct result of phosphatase inhibition and are not dependent on changes in cell volume. Whereas calyculin A exposure in the absence of volume changes results in stimulation of both the Na(+)/H(+) and K(+)/H(+) exchangers, superimposing cell swelling or shrinkage and calyculin A treatment results in selective activation of K(+)/H(+) or Na(+)/H(+) exchange, respectively. We conclude that kinase-dependent reactions are responsible for Na(+)/H(+) and K(+)/H(+) exchange activity, whereas undefined volume-dependent reactions confer specificity and coordinated control.
authors.affiliation: Dept. of Natural Sciences, Univ. of Puerto Rico, Utuado, Puerto Rico 00641. ortizacevedo@yahoo.com
authors.forename: Alejandro
authors.initials: A
authors.lastname: Ortiz-Acevedo
authors.forename: Robert R
authors.initials: RR
authors.lastname: Rigor
authors.forename: Hector M
authors.initials: HM
authors.lastname: Maldonado
authors.forename: Peter M
authors.initials: PM
authors.lastname: Cala
mesh_terms: D000584:Amiloride / Q000031:analogs & derivatives / Q000494:pharmacology; D000818:Animals; D048429:Cell Size / Q000187:drug effects*; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D004912:Erythrocytes / Q000187:drug effects* / Q000201:enzymology; D008387:Marine Toxins; D009997:Osmotic Pressure; D010080:Oxazoles / Q000494:pharmacology*; D010749:Phosphoprotein Phosphatases / Q000037:antagonists & inhibitors* / Q000378:metabolism; D010766:Phosphorylation; D011188:Potassium / Q000378:metabolism; D027961:Potassium-Hydrogen Antiporters / Q000819:agonists* / Q000378:metabolism; D011494:Protein Kinases / Q000378:metabolism; D012428:Ruthenium / Q000378:metabolism; D012964:Sodium / Q000378:metabolism; D017923:Sodium-Hydrogen Exchangers / Q000819:agonists* / Q000037:antagonists & inhibitors / Q000378:metabolism; D013997:Time Factors; D014562:Caudata / Q000097:blood*
chemical_list: D004791:Enzyme Inhibitors; D008387:Marine Toxins; D010080:Oxazoles; D027961:Potassium-Hydrogen Antiporters; D017923:Sodium-Hydrogen Exchangers; C059041:calyculin A; D000584:Amiloride; D012428:Ruthenium; D012964:Sodium; D011494:Protein Kinases; D010749:Phosphoprotein Phosphatases; D011188:Potassium; C039614:ethylisopropylamiloride
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