title: Evidence for facilitated diffusion of urea across the gill basolateral membrane of the rainbow trout (Oncorhynchus mykiss).
abstract: Recent in vivo evidence suggests that the mechanism of branchial urea excretion in the ammoniotelic rainbow trout (Oncorhynchus mykiss) is carrier-mediated. Further characterization of this proposed mechanism was achieved by using an in vitro isolated basolateral membrane vesicle (BLMV) preparation in which isolated gill membranes were used to determine a variety of physiological properties of the transporter. BLMV demonstrated two components of urea uptake, a linear component at concentrations up to 17.5 mmol x l(-1) and a saturable component (K(0.5)=0.35+/-0.01 mmol x l(-1); V(max)=0.14+/-0.02 micromol mg protein(-1) h(-1)) with a Hill constant of 1.35+/-0.18 at low, physiologically relevant urea concentrations (<2 mmol x l(-1)). Saturable uptake of urea at 1 mmol x l(-1) by BLMV was reduced by 88.5% when incubated with 0.25 mmol x l(-1) phloretin, a potent blocker of UT-type facilitated diffusion urea transport mechanisms. BLMV also demonstrated differential handling of urea versus urea analogues at 1 mmol x l(-1) concentrations and total analogue/total urea uptake ratios were 32% for acetamide and 84% for thiourea. Saturable urea uptake at 1 mmol x l(-1) was significantly reduced by almost 100% in the presence of 5 mmol x l(-1) thiourea but was not affected by 5 mmol x l(-1) acetamide or 5 mmol x l(-1) N-methylurea. Lastly, total urea uptake at 1 mmol x l(-1) by BLMV was sensitive to temperatures above and below the temperature of acclimation with a Q(10)>2 suggesting a protein carrier-mediated process. Combined, this evidence indicates that a facilitated diffusion urea transport mechanism is likely present in the basolateral membrane of the rainbow trout gill.
authors.affiliation: Department of Biology, McMaster University, Hamilton, Ontario, Canada, L8S 4K1. dmcdonald@rsmas.miami.edu
authors.forename: M Danielle
authors.initials: MD
authors.lastname: McDonald
authors.forename: Chris M
authors.initials: CM
authors.lastname: Wood
mesh_terms: D000064:Acclimatization / Q000502:physiology; D000081:Acetamides / Q000493:pharmacokinetics; D000818:Animals; D001693:Biological Transport, Active / Q000187:drug effects / Q000502:physiology; D002250:Carbon Radioisotopes; D002462:Cell Membrane / Q000378:metabolism*; D004058:Diffusion; D004847:Epithelial Cells / Q000378:metabolism; D005880:Gills / Q000378:metabolism*; D017686:Oncorhynchus mykiss / Q000378:metabolism* / Q000502:physiology; D010693:Phloretin / Q000494:pharmacology; D013696:Temperature; D013890:Thiourea / Q000493:pharmacokinetics; D014508:Urea / Q000031:analogs & derivatives / Q000493:pharmacokinetics*
chemical_list: D000081:Acetamides; D002250:Carbon Radioisotopes; D014508:Urea; D013890:Thiourea; D010693:Phloretin