Cadmium is a dangerous metal distributed widely in the environment. Members of our laboratory recently identified the ZIP8 transporter protein, encoded by the mouse Slc39a8 gene, to be responsible for genetic differences in response to cadmium damage of the testis. Stable retroviral infection of the ZIP8 cDNA in mouse fetal fibroblast cultures (rvZIP8 cells) leads to as much as a 10-fold increase in the rate of intracellular cadmium influx and accumulation. In the present study, we showed that cadmium uptake operated maximally at pH 7.5 and a temperature of 37°C and was inhibited by cyanide. Of more than a dozen cations tested, manganese(II) was the best competitive cation for cadmium uptake. The K_m for Cd²⁺ uptake was 0.62 μM, and the K_m for Mn²⁺ uptake was 2.2 μM; thus, manganese is probably the physiological substrate for ZIP8. Cadmium uptake was independent of sodium, potassium or chloride ions, but strongly dependent on the presence of bicarbonate. By Western blot analysis of rvZIP8 cells, we showed that ZIP8 protein was glycosylated. Using Z-stack confocal microscopy in Madin-Darby canine kidney polarized epithelial cells, we found that ZIP8 was localized on the apical side—implying an important role for manganese or cadmium uptake and disposition. It is likely that ZIP8 is a symporter, that a gradient across the plasma membrane acts as the driving force for manganese uptake, and that cadmium is a rogue hitchhiker displacing manganese to cause cadmium-associated disease.

The mouse and human genomes contain 14 highly conserved SLC39 genes. Viewed from an evolutionary perspective, SLC39A14 and SLC39A8 are the most closely related, each having three noncoding exons 1. However, SLC39A14 has two exons 4, giving rise to Zrt- and Irt-related protein (ZIP)ZIP14A and ZIP14B alternatively spliced products. C57BL/6J mouse ZIP14A expression is highest in liver, duodenum, kidney, and testis; ZIP14B expression is highest in liver, duodenum, brain, and testis; and ZIP8 is highest in lung, testis, and kidney. We studied ZIP14 stably retroviral-infected mouse fetal fibroblast cultures and transiently transfected Madin-Darby canine kidney (MDCK) polarized epithelial cells. Our findings include: 1) ZIP14-mediated cadmium uptake is proportional to cell toxicity, but manganese is not; 2) ZIP14B has a higher affinity than ZIP14A toward Cd²⁺ (K_m = 0.14 versus 1.1 μM) and Mn²⁺ uptake (K_m = 4.4 versus 18.2 μM); 3) ZIP14A- and ZIP14B-mediated Cd²⁺ uptake is most inhibited by Zn²⁺, and next by Mn²⁺ and Cu²⁺; 4) like ZIP8, ZIP14A- and ZIP14B-mediated Cd²⁺ uptake is dependent on extracellular \(\mathrm{HCO}_{3}^{-}\) ; 5) like ZIP8, ZIP14 transporters are localized on the apical surface of MDCK-ZIP cells; and 6) like ZIP8, ZIP14 proteins are glycosylated. Tissues such as intestine and liver, located between the environment and the animal, show high levels of ZIP14; given the high affinity for ZIP14, Cd²⁺ is likely to act as a rogue hitchhiker—displacing Zn²⁺ or Mn²⁺ and entering the body to cause unwanted cell damage and disease.

The identities of the apical Cl⁻/base exchangers in kidney proximal tubule and cortical collecting duct (CCD) cells remain unknown. Pendrin (PDS), which is expressed at high levels in the thyroid and its mutation causes Pendred's syndrome, is shown to be an anion exchanger. We investigated the renal distribution of PDS and its function. Our results demonstrate that pendrin mRNA expression in the rat kidney is abundant and limited to the cortex. Proximal tubule suspensions isolated from kidney cortex were highly enriched in pendrin mRNA. Immunoblot analysis studies localized pendrin to cortical brush-border membranes. Nephron segment RT-PCR localized pendrin mRNA to proximal tubule and CCD. Expression studies in HEK-293 cells demonstrated that pendrin functions in the Cl⁻/OH⁻, Cl⁻/HCO₃⁻, and Cl⁻/formate exchange modes. The conclusion is that pendrin is an apical Cl⁻/base exchanger in the kidney proximal tubule and CCD and mediates Cl⁻/OH⁻, Cl⁻/HCO₃⁻, and Cl⁻/formate exchange.

The identity of the transporter responsible for fructose absorption in the intestine in vivo and its potential role in fructose-induced hypertension remain speculative. Here we demonstrate that Glut5 (Slc2a5) deletion reduced fructose absorption by ∼75% in the jejunum and decreased the concentration of serum fructose by ∼90% relative to wild-type mice on increased dietary fructose. When fed a control (60% starch) diet, Glut5^-/- mice had normal blood pressure and displayed normal weight gain. However, whereas Glut5^+/+ mice showed enhanced salt absorption in their jejuna in response to luminal fructose and developed systemic hypertension when fed a high fructose (60% fructose) diet for 14 weeks, Glut5^-/- mice did not display fructose-stimulated salt absorption in their jejuna, and they experienced a significant impairment of nutrient absorption in their intestine with accompanying hypotension as early as 3–5 days after the start of a high fructose diet. Examination of the intestinal tract of Glut5^-/- mice fed a high fructose diet revealed massive dilatation of the caecum and colon, consistent with severe malabsorption, along with a unique adaptive up-regulation of ion transporters. In contrast to the malabsorption of fructose, Glut5^-/- mice did not exhibit an absorption defect when fed a high glucose (60% glucose) diet. We conclude that Glut5 is essential for the absorption of fructose in the intestine and plays a fundamental role in the generation of fructose-induced hypertension. Deletion of Glut5 results in a serious nutrient-absorptive defect and volume depletion only when the animals are fed a high fructose diet and is associated with compensatory adaptive up-regulation of ion-absorbing transporters in the colon.

SLC26A6 (PAT1, CFEX) is an anion exchanger that is expressed on the apical membrane of the kidney proximal tubule and the small intestine. Modes of transport mediated by SLC26A6 include Cl⁻/formate exchange, Cl⁻/HCO₃⁻ exchange, and Cl⁻/oxalate exchange. To study its role in kidney and intestinal physiology, gene targeting was used to prepare mice lacking Slc26a6. Homozygous mutant Slc26a6^−/− mice appeared healthy and exhibited a normal blood pressure, kidney function, and plasma electrolyte profile. In proximal tubules microperfused with a low-HCO₃⁻/high-Cl⁻ solution, the baseline rate of fluid absorption ( J_v), an index of NaCl transport under these conditions, was the same in wild-type and null mice. However, the stimulation of J_v by oxalate observed in wild-type mice was completely abolished in Slc26a6-null mice ( P < 0.05). Formate stimulation of J_v was partially reduced in null mice, but the difference from the response in wild-type mice did not reach statistical significance. Apical membrane Cl⁻/base exchange activity, assayed with the pH-sensitive dye BCPCF in microperfused proximal tubules, was decreased by 58% in Slc26a6^−/− animals ( P < 0.001 vs. wild types). In the duodenum, the baseline rate of HCO₃⁻ secretion measured in mucosal tissue mounted in Ussing chambers was decreased by ∼30% ( P < 0.03), whereas the forskolin-stimulated component of HCO₃⁻ secretion was the same in wild-type and Slc26a6^−/− mice. We conclude that Slc26a6 mediates oxalate-stimulated NaCl absorption, contributes to apical membrane Cl⁻/base exchange in the kidney proximal tubule, and also plays an important role in HCO₃⁻ secretion in the duodenum.