We have demonstrated that the kidney plays an important role in iron balance and that metabolically significant reabsorption of this ion occurs in the loop of Henle and the collecting ducts [Wareing M, Ferguson CJ, Green R, Riccardi D, and Smith CP. J Physiol (Lond) 524: 581-586, 2000]. To test the possibility that the divalent metal transporter DMT1 (Gunshin H, Mackenzie B, Berger UV, Gunshin Y, Romero MF, Boron WF, Nussberger S, Gollan JL, and Hediger MA. Nature 388: 482-488, 1997) could represent the apical route for iron entry in the kidney, we raised and affinity-purified an anti-DMT-1 polyclonal antibody and determined DMT-1 distribution in rat kidney by Western analysis, immunofluorescence, and confocal microscopy. The strongest DMT1-specific (i.e., peptide-protectable) immunoreactivity was found in the collecting ducts, in both principal and intercalated cells. Thick ascending limbs of Henle's loop and, more intensely, distal convoluted tubules exhibited apical immunostaining. Considerable intracellular DMT-1 immunoreactivity was seen throughout the nephron, particularly in S3 segments. The described distribution of DMT-1 protein is in agreement with our previous identification of nephron sites of iron reabsorption, suggesting that DMT-1 provides the molecular mechanism for apical iron entry in the distal nephron but not in the proximal tubule. Basolateral iron exit may be facilitated by a different system.

SUMMARY1. In rats receiving 200 gl. min-sodium chloride, glomerular filtration rate, single nephron glomerular filtration rate, and fractional reabsorption were measured at various points along the nephron, and ionic concentrations measured in early distal tubular fluid in virgin and 6, 12 and 19 day pregnant rats.2. Glomerular filtration increased progressively until the twelfth day ofpregnancy. At 19 days of pregnancy the glomerular filtration rate, while still above virgin levels, was reduced below 12 day pregnant levels.3. Single nephron glomerular filtration rates measured in proximal and distal tubules were different at both 12 and 19 days of pregnancy, indicating an alteration of tubuloglomerular feed-back. 4. A change in the ratio ofglomerular filtration rate: distal single nephron filtration rate indicated a redistribution of glomerular filtrate to juxtamedullary nephrons by the sixth day of pregnancy.5. Fluid reabsorption is similar up to the early distal tubule but it is not possible to say whether reabsorption is the same in proximal tubules. More fluid is reabsorbed by late distal tubules and collecting ducts in 12 and 19 day pregnant animals than in the virgin and 6 day pregnant animals.6. Changes in ion reabsorption by the loop of Henle occurred during pregnancy; sodium reabsorption was increased by the sixth day of pregnancy and potassium reabsorption by the twelfth day.

The aims of this study were to investigate (a) if renal Na+ handling was normal in Cftrtm2camΔF508 cystic fibrosis mice, (b) whether adaptation to dietary salt depletion was preserved and (c) whether Cftrtm2camΔF508 mice exhibited enhanced amiloride‐sensitive Na+ absorption. In Na+‐replete animals (maintained on a 0.32 % NaCl diet) given a 150 mM NaCl i.v. maintenance infusion, there was no difference in fractional Na+ excretion (FENa) between wild‐type (0.42 ± 0.06 %, n= 12) and Cftrtm2camΔF508 mice (0.47 ± 0.13 %, n= 7). Amiloride infusion significantly increased FENa in both wild‐type (3.14 ± 0.83 %, n= 6) and Cftrtm2camΔF508 mice (3.47 ± 0.63 %, n= 9), though with no significant difference between genotypes. A 14 day dietary salt restriction (animals maintained on a 0.03 % NaCl diet) and maintenance infusion with a 15 mM NaCl vehicle caused a reduction in FENa to 0.14 ± 0.05 %, n= 8 in wild‐type mice and 0.14 ± 0.04 %, n= 8 in Cftrtm2camΔF508 mice. No significant difference in the ability to adapt to low salt conditions was apparent comparing the two genotypes. Treatment of salt‐restricted mice with amiloride resulted in a blunted natriuresis in both wild‐type mice (FENa= 1.10 ± 0.16 %, n= 7) and Cftrtm2camΔF508 mice (FENa= 1.97 ± 0.29 %, n= 9). The natriuresis induced by amiloride was significantly greater in Cftrtm2camΔF508 mice than in wild‐type controls. In conclusion, Cftrtm2camΔF508 mice exhibit normal renal salt excretion when either salt replete or salt restricted. Enhanced amiloride‐sensitive FENa is consistent with increased Na+ absorption via the amiloride‐sensitive sodium channel ENaC, in cystic fibrosis kidney, but this was only observed during salt restriction.

The role of diffusion in transepithelial potassium flux and the importance of potassium channels in the luminal cell membrane to this process were examined by applying a luminal microperfusion technique to surface tubules in kidneys of anesthetized rats. Potassium concentration gradients were applied by altering the concentration of KCl in perfusates. To some perfusates, 2 mmol/l BaCl2 was added to block potassium channels in the luminal cell membrane. The mean applied potassium concentration gradient was highly predictive of net potassium transport in the absence of any change in fluid reabsorption, with an apparent potassium permeability of 22 x 10(-5) cm/s. Thus potassium transport in the proximal tubule may have an important diffusive component. Luminal barium significantly reduced the concentration of potassium in collected fluid under conditions of net potassium secretion, although a substantial barium-insensitive potassium permeability was also observed. However, the site of action of luminally applied barium is uncertain in proximal tubule, since barium was reabsorbed by the tubule at a rate of 13.6 pmol.mm-1.min-1. We conclude that diffusion is a significant driving force for potassium reabsorption in proximal tubule and that most diffusive potassium transport occurs via a barium-insensitive route, possibly the paracellular pathway.

Changes in proximal tubule function have been reported in cystic fibrosis patients. The aim of this study was to investigate proximal tubule function in the Cftrtm2camΔF508 cystic fibrosis (CF) mouse model. A range of techniques were used including renal clearance studies, in situ microperfusion, RT‐PCR and whole‐cell patch clamping. Renal Na+ clearance was similar in wild‐type (1.4 ± 0.3 μl min−1, number of animals, N= 12) and CF mice (1.6 ± 0.4 μl min−1, N= 7) under control conditions. Acute extracellular volume expansion resulted in significant natriuresis in wild‐type (7.0 ± 0.8 μl min−1, N= 8) and CF mice (9.3 ± 1.4 μl min−1, N= 9); no difference between genotypes was observed. In situ microperfusion revealed that fluid absorptive rate (Jv) was similar under control conditions between wild‐type (2.2 ± 0.4 nl mm−1 min−1, n= 10) and CF mice (1.9 ± 0.3 nl mm−1 min−1, n= 11). Addition of a forskolin‐dibutyryl cAMP (db‐cAMP) cocktail to the perfusate caused no significant change in Jv in either wild‐type (2.6 ± 0.7 nl mm−1 min−1, n= 10) or Cftrtm2camΔF508 mice (2.0 ± 0.5 nl mm−1 min−1, n= 10). CFTR expression was confirmed in samples of outer cortex using RT‐PCR. However, no evidence for functional CFTR was obtained when outer cortical cells were stimulated with protein kinase A or forskolin‐db‐cAMP using whole‐cell patch clamping. In conclusion, no functional deficit in proximal tubule function was found in Cftrtm2camΔF508 mice. This may be a consequence of a lack of whole‐cell cAMP‐dependent Cl− conductance in mouse proximal tubule cells.