Rapid modulation of Na+/K+-ATPase activity in osmoregulatory tissues of a salmonid fish.
The effects of cyclic AMP on Na+/K+-ATPase activity were studied in the gill and kidney of the euryhaline brown trout Salmo trutta using two different experimental approaches. In the first series of experiments, in situ Na+/K+-ATPase activity was analyzed by measuring the ouabain-sensitive uptake of non-radioactive rubidium (Rb+) into gill cells and blocks of gill and kidney tissue. Rubidium uptake was linear for at least 30 min and was significantly inhibited by 1 mmol x l(-1) ouabain. Several agents presumed to increase the intracellular cyclic AMP concentration inhibited ouabain-sensitive Rb+ uptake in both gill (0.5 and 2 mmol x l(-1) dibutyryl-cyclic AMP, 1 mmol x l(-1) theophylline, 10 micromol x l(-1) forskolin and 10 micromol x l(-1 )isoproterenol) and kidney (10 micromol x l(-1) forskolin) tissue from freshwater-acclimated fish. In a separate series of experiments, ATP hydrolase activity was assayed in a permeabilised gill membrane preparation after incubation of tissue blocks with 10 micromol x l(-1 )forskolin. Forskolin elevated gill cyclic AMP levels 40-fold, inhibited maximal enzymatic Na+/K+-ATPase activity (Vmax) in gill tissue from both freshwater- and seawater-acclimated fish and reduced the apparent K+ affinity in the gills of seawater-acclimated fish, demonstrating that the effects are mediated through modifications of the enzyme itself. The protein phosphatase inhibitors okadaic acid and cyclosporin A did not affect forskolin-induced inhibition of Na+/K+-ATPase activity, indicating that forskolin-mediated modulation was stable for the duration of assay. We suggest that cyclic-AMP-mediated phosphorylation through protein kinases may underlie the rapid modulation of Na+/K+-ATPase activity in the osmoregulatory tissues of euryhaline teleosts.
Tipsmark
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Institute of Biology, University of Southern Denmark, Odense University, Campusvej 55, DK-5230 Odense M, Denmark.
Madsen
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D000818:Animals; D003994:Bucladesine / Q000494:pharmacology; D005576:Colforsin / Q000494:pharmacology*; D000242:Cyclic AMP / Q000378:metabolism*; D016572:Cyclosporine / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D005618:Fresh Water / Q000737:chemistry; D005880:Gills / Q000166:cytology / Q000187:drug effects / Q000201:enzymology*; D066298:In Vitro Techniques; D007668:Kidney / Q000166:cytology / Q000187:drug effects / Q000201:enzymology*; D019319:Okadaic Acid / Q000494:pharmacology; D010042:Ouabain / Q000494:pharmacology; D010726:Phosphodiesterase Inhibitors / Q000494:pharmacology; D012414:Rubidium Radioisotopes / Q000378:metabolism; D012623:Seawater / Q000737:chemistry; D000254:Sodium-Potassium-Exchanging ATPase / Q000378:metabolism*; D013806:Theophylline / Q000494:pharmacology; D014337:Trout / Q000378:metabolism*
D004791:Enzyme Inhibitors; D010726:Phosphodiesterase Inhibitors; D012414:Rubidium Radioisotopes; D005576:Colforsin; D019319:Okadaic Acid; D010042:Ouabain; D003994:Bucladesine; D016572:Cyclosporine; D013806:Theophylline; D000242:Cyclic AMP; D000254:Sodium-Potassium-Exchanging ATPase