title: Mechanistic characterization of gastric copper transport in rainbow trout.
abstract: An in vitro gut-sac technique and (64)Cu as a radiotracer were used to characterize gastric copper (Cu) transport. Cu transport was stimulated by low luminal pH (4.0 vs. 7.4), to a greater extent than explained by the increased availability of the free Cu(2+) ion. At pH = 4.0, uptake kinetics were indicative of a low affinity (K (m) = 525 μmol L(-1)), saturable carrier-mediated component superimposed on a large linear (diffusive and/or convective) component, with about 50% occurring by each pathway at Cu = 50 μmol L(-1). Osmotic gradient experiments showed that solvent drag via fluid transport may play a role in Cu uptake via the stomach, in contrast to the intestine. Also unlike the intestine, neither the Na(+) gradient, high Ag, nor phenamil had any influence on gastric Cu transport, and a tenfold excess of Fe and Zn failed to inhibit Cu uptake. These findings indicate that neither Na(+)-dependent pathways nor DMT1 are likely candidates for carrier-mediated Cu transport in the stomach. We have cloned a partial cDNA sequence for the copper transporter Ctr1, and show its mRNA expression in all segments of the trout gastrointestinal tract, including the stomach. Based on the fact that this transporter is functional at low pH conventionally found in the stomach lumen, we suggest Ctr1 is a pathway for gastric Cu transport in trout. Extreme hypoxia inhibited Cu uptake. High P(CO₂) levels (7.5 torr) increased Cu uptake and acetazolamide (100 μmol L(-1)) significantly inhibited Cu uptake, indicating carbonic anhydrase activity was involved in gastric Cu transport. Transport of Cu was insensitive to bafilomycin (10 μmol L(-1)) suggesting a V-ATPase did not play a direct role in the process. Expression (mRNA) of H (+) , K (+)-ATPase, carbonic anhydrase 2, and the α-3 isoform of Na (+)-K (+)-ATPase were observed in the stomach. We suggest these enzymes facilitate Cu transport in the stomach indirectly as part of a physiological mechanism exporting H(+) to the cell exterior. However, pre-treatment with the H (+) , K (+)-ATPase proton pump blocker omeprazole did not affect gastric Cu transport, suggesting that other mechanisms must also be involved.
authors.affiliation: Department of Biology, McMaster University, Hamilton, ON L8S 4K1, Canada. nadellsr@mcmaster.ca
authors.forename: Sunita R
authors.initials: SR
authors.lastname: Nadella
authors.forename: Carrie C Y
authors.initials: CC
authors.lastname: Hung
authors.forename: Chris M
authors.initials: CM
authors.lastname: Wood
mesh_terms: D000818:Animals; D001692:Biological Transport / Q000187:drug effects; D002256:Carbonic Anhydrases / Q000378:metabolism; D003300:Copper / Q000097:blood / Q000493:pharmacokinetics / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D041981:Gastrointestinal Tract / Q000187:drug effects* / Q000201:enzymology / Q000378:metabolism; D017506:H(+)-K(+)-Exchanging ATPase / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D017686:Oncorhynchus mykiss / Q000097:blood / Q000378:metabolism*
chemical_list: D003300:Copper; D017506:H(+)-K(+)-Exchanging ATPase; D002256:Carbonic Anhydrases
references.citation: Am J Physiol. 1978 Mar;234(3):E228-35
references.pmid: 24346
references.citation: J Morphol. 1978 Nov;158(2):155-67
references.pmid: 731703
references.citation: Comp Biochem Physiol C Toxicol Pharmacol. 2009 Sep;150(3):349-60
references.pmid: 19527800
references.citation: J Exp Biol. 1999 Aug;202(Pt 15):2103-10
references.pmid: 10393825
references.citation: Am J Physiol Regul Integr Comp Physiol. 2006 Dec;291(6):R1764-72
references.pmid: 16902189
references.citation: Comp Biochem Physiol C Toxicol Pharmacol. 2009 Nov;150(4):442-9
references.pmid: 19584005
references.citation: Br J Nutr. 1987 May;57(3):479-88
references.pmid: 3593674
references.citation: Aquat Toxicol. 2008 Aug 11;89(1):55-64
references.pmid: 18619683
references.citation: Gene. 2004 Mar 17;328:113-20
references.pmid: 15019990
references.citation: Am J Physiol Cell Physiol. 2003 Jun;284(6):C1525-30
references.pmid: 12734107
references.citation: Biosci Rep. 2000 Apr;20(2):51-91
references.pmid: 10965965
references.citation: Yale J Biol Med. 1992 Nov-Dec;65(6):577-96
references.pmid: 1341065
references.citation: J Membr Biol. 1988 Oct;105(1):1-21
references.pmid: 2852254
references.citation: J Comp Physiol B. 2007 Apr;177(3):349-60
references.pmid: 17211667
references.citation: Int J Biochem Cell Biol. 2003 Mar;35(3):288-91
references.pmid: 12531240
references.citation: J Comp Physiol B. 2006 Nov;176(8):793-806
references.pmid: 16835745
references.citation: Comp Biochem Physiol C Toxicol Pharmacol. 2008 May;147(4):450-9
references.pmid: 18304880
references.citation: Aquat Toxicol. 2005 Mar 25;72(1-2):99-117
references.pmid: 15748750
references.citation: Proc Soc Exp Biol Med. 1991 Apr;196(4):410-4
references.pmid: 1848938
references.citation: J Pharmacol Exp Ther. 1957 Aug;120(4):528-39
references.pmid: 13476377
references.citation: Anal Chem. 1999 Aug 1;71(15):3140-4
references.pmid: 21662904
references.citation: J Biol Chem. 2002 Feb 8;277(6):4380-7
references.pmid: 11734551
references.citation: J Exp Biol. 2003 Jan;206(Pt 1):11-23
references.pmid: 12456693
references.citation: Biochim Biophys Acta. 1973 Apr 25;307(1):169-80
references.pmid: 4351147
references.citation: FEBS Lett. 2007 Jun 12;581(14):2599-604
references.pmid: 17509573
references.citation: J Membr Biol. 1985;87(1):45-54
references.pmid: 2414446
references.citation: J Comp Physiol B. 2007 May;177(4):433-46
references.pmid: 17279389
references.citation: J Exp Biol. 2001 Nov;204(Pt 21):3779-87
references.pmid: 11719541
references.citation: Cell Metab. 2006 Sep;4(3):235-44
references.pmid: 16950140
references.citation: J Exp Biol. 2009 Mar;212(Pt 5):684-92
references.pmid: 19218520
references.citation: J Nutr. 2006 Jan;136(1):21-6
references.pmid: 16365053
references.citation: Aquat Toxicol. 2007 Jun 5;83(1):10-23
references.pmid: 17448547
references.citation: J Comp Physiol B. 2008 Jan;178(1):101-9
references.pmid: 17724600
references.citation: Environ Toxicol Chem. 2005 Nov;24(11):2954-64
references.pmid: 16398134
references.citation: Nature. 1997 Jul 31;388(6641):482-8
references.pmid: 9242408
references.citation: Physiol Rev. 1948 Oct;28(4):433-50
references.pmid: 18894957
references.citation: J Exp Biol. 2003 Dec;206(Pt 24):4475-86
references.pmid: 14610032
references.citation: J Membr Biol. 2008 Jan;221(2):87-95
references.pmid: 18217179
references.citation: Physiol Rev. 1995 Jan;75(1):155-89
references.pmid: 7831396
references.citation: Hum Mol Genet. 1995 Nov;4(11):2117-23
references.pmid: 8589689
references.citation: Am J Physiol. 1998 Nov;275(5):F633-50
references.pmid: 9815123
references.citation: J Biol Chem. 1976 Dec 10;251(23):7690-8
references.pmid: 12175
references.citation: Aquat Toxicol. 2009 Jul 26;93(4):205-16
references.pmid: 19515435
references.citation: J Anim Sci. 1980 Jul;51(1):186-92
references.pmid: 7410271
references.citation: J Exp Biol. 2000 Aug;203(Pt 16):2455-66
references.pmid: 10903160
references.citation: Clin Sci Mol Med. 1975 Sep;49(3):237-45
references.pmid: 240530
references.citation: J Exp Biol. 2006 Oct;209(Pt 19):3719-28
references.pmid: 16985189
references.citation: Comp Biochem Physiol C Toxicol Pharmacol. 2006 Aug;143(4):394-401
references.pmid: 16765095