title: Cloning and characterization of Na(+)/H(+) Exchanger isoforms NHE2 and NHE3 from the gill of Pacific dogfish Squalus suckleyi.
abstract: Na(+)/H(+) Exchanger (NHE) proteins mediate cellular and systemic homeostasis of sodium and acid and may be the major sodium uptake method for fishes. We cloned and sequenced NHE2 and NHE3 from the gill of the North Pacific Spiny Dogfish shark Squalus suckleyi and expressed them in functional form in NHE-deficient (AP-1) cell lines. Estimated IC50 for inhibition of NHE activity by amiloride and EIPA were 55 μmol l(-1) and 4.8 μmol l(-1), respectively, for NHE2 and 9 μmol l(-1) and 24 μmol l(-1), respectively, for NHE3. Phenamil at 100 μmol l(-1) caused less than 16% inhibition of activity for each isoform. Although the IC50 are similar for the two isoforms, dfNHE2 is less sensitive than human NHE2 to inhibition by amiloride and EIPA, while dfNHE3 is more sensitive than human NHE3. These IC50 estimates should be considered when selecting inhibitor doses for fishes and for reinterpretation of previous studies that use these pharmacological agents.
authors.affiliation: Department of Biological Sciences, Z512 Biological Sciences Bldg, University of Alberta, Edmonton, AB T6G 2E9, Canada; Bamfield Marine Sciences Centre, 100 Pachena Road, Bamfield, BC V0R 1B0, Canada. Electronic address: guffey0@purdue.edu.
authors.forename: Samuel C
authors.initials: SC
authors.lastname: Guffey
authors.affiliation: Department of Biochemistry, 347 Medical Sciences Bldg, University of Alberta, Edmonton, AB T6G 2H7, Canada. Electronic address: lfliegel@ualberta.ca.
authors.forename: Larry
authors.initials: L
authors.lastname: Fliegel
authors.affiliation: Department of Biological Sciences, Z512 Biological Sciences Bldg, University of Alberta, Edmonton, AB T6G 2E9, Canada; Bamfield Marine Sciences Centre, 100 Pachena Road, Bamfield, BC V0R 1B0, Canada. Electronic address: ggoss@ualberta.ca.
authors.forename: Greg G
authors.initials: GG
authors.lastname: Goss
mesh_terms: D000584:Amiloride / Q000031:analogs & derivatives / Q000494:pharmacology; D000818:Animals; D001693:Biological Transport, Active / Q000187:drug effects*; D002460:Cell Line; D003001:Cloning, Molecular / Q000379:methods; D006224:Cricetinae; D004284:Dogfish / Q000235:genetics*; D062686:Epithelial Sodium Channel Blockers / Q000494:pharmacology*; D029941:Fish Proteins / Q000235:genetics / Q000378:metabolism*; D005880:Gills / Q000378:metabolism; D020128:Inhibitory Concentration 50; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D017923:Sodium-Hydrogen Exchangers / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*
chemical_list: D062686:Epithelial Sodium Channel Blockers; D029941:Fish Proteins; D020033:Protein Isoforms; D017923:Sodium-Hydrogen Exchangers; C047088:phenylamil; D000584:Amiloride; C039614:ethylisopropylamiloride
keywords: Cell; Elasmobranch; Homeostasis; Proton; Transport