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The thick ascending limb (TAL) of Henle’s loop drives paracellular Na+, Ca2+, and Mg2+ reabsorption via the tight junction (TJ). The TJ is composed of claudins that consist of four transmembrane segments, two extracellular segments (ECS1 and -2), and one intracellular loop. Claudins interact within the same (cis) and opposing (trans) plasma membranes. The claudins Cldn10b, -16, and -19 facilitate cation reabsorption in the TAL, and their absence leads to a severe disturbance of renal ion homeostasis. We combined electrophysiological measurements on microperfused mouse TAL segments with subsequent analysis of claudin expression by immunostaining and confocal microscopy. Claudin interaction properties were examined using heterologous expression in the TJ-free cell line HEK 293, live-cell imaging, and Förster/FRET. To reveal determinants of interaction properties, a set of TAL claudin protein chimeras was created and analyzed. Our main findings are that (i) TAL TJs show a mosaic expression pattern of either cldn10b or cldn3/cldn16/cldn19 in a complex; (ii) TJs dominated by cldn10b prefer Na+ over Mg2+, whereas TJs dominated by cldn16 favor Mg2+ over Na+; (iii) cldn10b does not interact with other TAL claudins, whereas cldn3 and cldn16 can interact with cldn19 to form joint strands; and (iv) further claudin segments in addition to ECS2 are crucial for trans interaction. We suggest the existence of at least two spatially distinct types of paracellular channels in TAL: a cldn10b-based channel for monovalent cations such as Na+ and a spatially distinct site for reabsorption of divalent cations such as Ca2+ and Mg2+. [ABSTRACT FROM AUTHOR]

Large deletions in the first intron of the With No lysine (K) 1 (WNK1) gene are responsible for Familial Hyperkalemic Hypertension (FHHt), a rare form of human hypertension associated with hyperkalemia and hyperchloremic metabolic acidosis. We generated a mouse model of WNK1-associated FHHt to explore the consequences of this intronic deletion. WNK1+/FHHt mice display all clinical and biological signs of FHHt. This phenotype results from increased expression of long WNK1 (L-WNK1), the ubiquitous kinase isoform of WNK1, in the distal convoluted tubule, which in turn, stimulates the activity of the Na-Cl cotransporter. We also show that the activity of the epithelial sodium channel is not altered in FHHt mice, suggesting that other mechanisms are responsible for the hyperkalemia and acidosis in this model. Finally, we observe a decreased expression of the renal outer medullary potassium channel in the late distal convoluted tubule of WNK1+/FHHt mice, which could contribute to the hyperkalemia. In summary, our study provides insights into the in vivo mechanisms underlying the pathogenesis of WNK1-mediated FHHt and further corroborates the importance of WNK1 in ion homeostasis and blood pressure. [ABSTRACT FROM AUTHOR]

In the kidney, tight junction proteins contribute to segment specific selectivity and permeability of paracellular ¡on transport. In the thick ascending limb (TAL) of Henle's loop, chloride is reabsorbed transcellularly, whereas sodium reabsorption takes transcellular and paracellular routes. TAL salt transport maintains the concentrating ability of the kidney and generates a transepithelial voltage that drives the reabsorption of calcium and magnesium. Thus, functionality of TAL ion transport depends strongly on the properties of the paracellular pathway. To elucidate the role of the tight junction protein claudin-10 in TAL function, we generated mice with a deletion of Cldn10 in this segment. We show that claudin-10 determines paracellular sodium permeability, and that its 10ss leads to hypermagnesemia and nephrocalcinosis. In isolated perfused TAL tubules of claudin-10-deficient mice, paracellular permeability of sodium is decreased, and the relative permeability of calcium and magnesium is increased. Moreover, furosemide-inhibitable transepithelial voltage is increased, leading to a shift from paracellular sodium transport to paracellular hyperabsorption of calcium and magnesium. These data identify claudin-10 as a key factor in control of cation selectivity and transport in the TAL, and deficiency in this pathway as a cause of nephrocalcinosis. [ABSTRACT FROM AUTHOR]

The thick ascending limb (TAL) of Henle's loop drives paracellular Na + , Ca 2+ , and Mg 2+ reabsorption via the tight junction (TJ). The TJ is composed of claudins that consist of four transmembrane segments, two extracellular segments (ECS1 and -2), and one intracellular loop. Claudins interact within the same (cis) and opposing (trans) plasma membranes. The claudins Cldn10b, -16, and -19 facilitate cation reabsorption in the TAL, and their absence leads to a severe disturbance of renal ion homeostasis. We combined electrophysiological measurements on microperfused mouse TAL segments with subsequent analysis of claudin expression by immunostaining and confocal microscopy. Claudin interaction properties were examined using heterologous expression in the TJ-free cell line HEK 293, live-cell imaging, and Förster/FRET. To reveal determinants of interaction properties, a set of TAL claudin protein chimeras was created and analyzed. Our main findings are that (i) TAL TJs show a mosaic expression pattern of either cldn10b or cldn3/cldn16/cldn19 in a complex; (ii) TJs dominated by cldn10b prefer Na + over Mg 2+ , whereas TJs dominated by cldn16 favor Mg 2+ over Na + ; (iii) cldn10b does not interact with other TAL claudins, whereas cldn3 and cldn16 can interact with cldn19 to form joint strands; and (iv) further claudin segments in addition to ECS2 are crucial for trans interaction. We suggest the existence of at least two spatially distinct types of paracellular channels in TAL: a cldn10b-based channel for monovalent cations such as Na + and a spatially distinct site for reabsorption of divalent cations such as Ca 2+ and Mg 2 ., Competing Interests: The authors declare no conflict of interest.

In the kidney, tight junction proteins contribute to segment specific selectivity and permeability of paracellular ion transport. In the thick ascending limb (TAL) of Henle's loop, chloride is reabsorbed transcellularly, whereas sodium reabsorption takes transcellular and paracellular routes. TAL salt transport maintains the concentrating ability of the kidney and generates a transepithelial voltage that drives the reabsorption of calcium and magnesium. Thus, functionality of TAL ion transport depends strongly on the properties of the paracellular pathway. To elucidate the role of the tight junction protein claudin-10 in TAL function, we generated mice with a deletion of Cldn10 in this segment. We show that claudin-10 determines paracellular sodium permeability, and that its loss leads to hypermagnesemia and nephrocalcinosis. In isolated perfused TAL tubules of claudin-10-deficient mice, paracellular permeability of sodium is decreased, and the relative permeability of calcium and magnesium is increased. Moreover, furosemide-inhibitable transepithelial voltage is increased, leading to a shift from paracellular sodium transport to paracellular hyperabsorption of calcium and magnesium. These data identify claudin-10 as a key factor in control of cation selectivity and transport in the TAL, and deficiency in this pathway as a cause of nephrocalcinosis.