Department of Biology, McMaster University, Hamilton, ON L8S 4K1, Canada.
J Comp Physiol B. 2011 Jan;181(1):27-41. doi: 10.1007/s00360-010-0510-x. Epub 2010 Sep 3.
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.
采用离体肠囊技术和 64Cu 作为示踪剂，研究了胃铜（Cu）转运。低腔内 pH（4.0 对 7.4）刺激 Cu 转运，其程度大于游离 Cu2+离子增加的程度。在 pH = 4.0 时，摄取动力学表明低亲和力（Km = 525 μmol L-1），饱和载体介导的成分叠加在大的线性（扩散和/或对流）成分上，在 Cu = 50 μmol L-1 时，每种途径分别发生约 50%。渗透梯度实验表明，通过流体运输的溶剂拖曳可能在胃中通过胃吸收铜起作用，与肠相反。与肠不同，Na+梯度、高 Ag 或苯甲酰胺对胃 Cu 转运均无影响，Fe 和 Zn 的十倍过量也不能抑制 Cu 摄取。这些发现表明，Na+依赖性途径或 DMT1 均不太可能是胃中载体介导的 Cu 转运的候选者。我们已经克隆了铜转运蛋白 Ctr1 的部分 cDNA 序列，并显示其 mRNA 在鳟鱼胃肠道的所有节段表达，包括胃。基于该转运蛋白在胃腔中通常发现的低 pH 下具有功能的事实，我们认为 Ctr1 是鳟鱼胃铜转运的途径。极度缺氧抑制 Cu 摄取。高 P(CO₂)水平（7.5 torr）增加 Cu 摄取，乙酰唑胺（100 μmol L-1）显著抑制 Cu 摄取，表明碳酸酐酶活性参与胃 Cu 转运。Cu 转运对巴弗洛霉素（10 μmol L-1）不敏感，表明 V-ATPase 未直接参与该过程。胃中观察到 H(+)、K(+)-ATPase、碳酸酐酶 2 和 Na(+)-K(+)-ATPase 的 α-3 同工型的表达（mRNA）。我们认为这些酶通过将 H(+)输出到细胞外，作为一种生理机制间接促进胃中 Cu 转运。然而，预先用 H(+)、K(+)-ATPase 质子泵抑制剂奥美拉唑处理并不影响胃 Cu 转运，这表明还必须有其他机制参与。
