Thiazide diuretics enhance renal Na⁺ excretion by blocking the Na⁺-Cl^– cotransporter (NCC), and mutations in NCC result in Gitelman syndrome. The mechanisms underlying the accompanying hypocalciuria and hypomagnesemia remain debated. Here, we show that enhanced passive Ca²⁺ transport in the proximal tubule rather than active Ca²⁺ transport in distal convolution explains thiazide-induced hypocalciuria. First, micropuncture experiments in mice demonstrated increased reabsorption of Na⁺ and Ca²⁺ in the proximal tubule during chronic hydrochlorothiazide (HCTZ) treatment, whereas Ca²⁺ reabsorption in distal convolution appeared unaffected. Second, HCTZ administration still induced hypocalciuria in transient receptor potential channel subfamily V, member 5–knockout (Trpv5-knockout) mice, in which active distal Ca²⁺ reabsorption is abolished due to inactivation of the epithelial Ca²⁺ channel Trpv5. Third, HCTZ upregulated the Na⁺/H⁺ exchanger, responsible for the majority of Na⁺ and, consequently, Ca²⁺ reabsorption in the proximal tubule, while the expression of proteins involved in active Ca²⁺ transport was unaltered. Fourth, experiments addressing the time-dependent effect of a single dose of HCTZ showed that the development of hypocalciuria parallels a compensatory increase in Na⁺ reabsorption secondary to an initial natriuresis. Hypomagnesemia developed during chronic HCTZ administration and in NCC-knockout mice, an animal model of Gitelman syndrome, accompanied by downregulation of the epithelial Mg²⁺ channel transient receptor potential channel subfamily M, member 6 (Trpm6). Thus, Trpm6 downregulation may represent a general mechanism involved in the pathogenesis of hypomagnesemia accompanying NCC inhibition or inactivation.

: The family of epithelial Ca²⁺ channels consists of two highly homologues members, TRPV5 and TRPV6, which constitute the apical Ca²⁺ entry mechanism in active Ca²⁺ (re)absorption in kidney and small intestine. In kidney, TRPV5 expression has been extensively studied, whereas TRPV6 localization and regulation has been largely confined to the small intestine. The present study investigated the renal distribution of TRPV6 and regulation by 1,25-dihydroxyvitamin D₃ (1,25(OH)₂D₃). In mouse kidney, TRPV6 was detected by immunohistochemistry at the apical domain of the distal convoluted tubules (DCT2), connecting tubules (CNT), and cortical and medullary collecting ducts (CD). Furthermore, several putative vitamin D-responsive elements were detected upstream of the mouse TRPV6 start codon, and 1,25(OH)₂D₃ treatment significantly increased renal TRPV6 mRNA and protein expression. In DCT2 and CNT, TRPV6 co-localizes with the other known Ca²⁺ transport proteins, including TRPV5 and calbindin-D_28K. Together, these data suggest a role for TRPV6 in 1,25(OH)₂D₃-stimulated Ca²⁺ reabsorption in these segments. Interestingly, distribution of TRPV6 extended to the CD, where it localized to the apical domain of principal and intercalated cells, which are not generally implicated in active Ca²⁺ reabsorption. In addition, TRPV6 mRNA levels were quantified in a large set of tissues, and in the order of decreasing expression level were detected: prostate > stomach, brain > lung > duodenum, kidney, bone, cecum, heart > colon > skeletal muscle > pancreas. Therefore, additional physiologic functions for TRPV6 are feasible. In conclusion, TRPV6 is expressed along the apical domain of DCT2, CNT, and CD, where TRPV6 expression is positively regulated by 1,25(OH)₂D₃. E-mail: r.bindels@ncmls.kun.nl

FK506 (tacrolimus) and dexamethasone are potent immunosuppressants known to induce significant side effects on mineral homeostasis, including hypercalciuria and hypomagnesemia. However, the underlying molecular mechanisms remain unknown. The present study investigated the effects of FK506 and dexamethasone on the expression of proteins involved in active Ca²⁺ reabsorption: the epithelial Ca²⁺ channel TRPV5 and the cytosolic Ca²⁺-binding protein calbindin-D_28K. In addition, the renal expression of the putative Mg²⁺ channel TRPM6, suggested to be involved in transcellular Mg²⁺ reabsorption, was determined. Administration of FK506 to rats by daily oral gavage during 7 d significantly enhanced the urinary excretion of Ca²⁺ and Mg²⁺ and induced a significant hypomagnesemia. FK506 significantly decreased the renal mRNA expression of TRPV5 (62 ± 7% relative to controls), calbindin-D_28K (9 ± 1%), and TRPM6 (52 ± 8%), as determined by real-time quantitative PCR analysis. Furthermore, semiquantitative immunohistochemistry showed reduced renal protein abundance of TRPV5 (24 ± 5%) and calbindin-D_28K (29 ± 4%), altogether suggesting that downregulation of these transport proteins is responsible for the FK506-induced Ca²⁺ and Mg²⁺ wasting. In contrast, dexamethasone significantly enhanced renal TRPV5 (150 ± 15%), calbindin-D_28K (177 ± 23%), and TRPM6 (156 ± 20%) mRNA levels along with TRPV5 (211 ± 8%) and calbindin-D_28K (176 ± 5%) protein abundance in the presence of significantly increased Ca²⁺ and Mg²⁺ excretion. This indicated that these proteins are directly or indirectly regulated by dexamethasone. In conclusion, FK506 and dexamethasone induce renal Ca²⁺ and Mg²⁺ wasting, albeit by different mechanisms. Downregulation of specific Ca²⁺ and Mg²⁺ transport proteins provides a molecular mechanism for FK506-induced hypercalciuria and hypomagnesemia, whereas dexamethasone positively regulates these proteins.

Hepatocyte nuclear factor 1β (HNF1β)–associated disease is a recently recognized clinical entity with a variable multisystem phenotype. Early reports described an association between HNF1B mutations and maturity-onset diabetes of the young. These patients often presented with renal cysts and renal function decline that preceded the diabetes, hence it was initially referred to as renal cysts and diabetes syndrome. However, it is now evident that many more symptoms occur, and diabetes and renal cysts are not always present. The multisystem phenotype is probably attributable to functional promiscuity of the HNF1β transcription factor, involved in the development of the kidney, urogenital tract, pancreas, liver, brain, and parathyroid gland. Nephrologists might diagnose HNF1β-associated kidney disease in patients referred with a suspected diagnosis of autosomal dominant polycystic kidney disease, medullary cystic kidney disease, diabetic nephropathy, or CKD of unknown cause. Associated renal or extrarenal symptoms should alert the nephrologist to HNF1β-associated kidney disease. A considerable proportion of these patients display hypomagnesemia, which sometimes mimics Gitelman syndrome. Other signs include early onset diabetes, gout and hyperparathyroidism, elevated liver enzymes, and congenital anomalies of the urogenital tract. Because many cases of this disease are probably undiagnosed, this review emphasizes the clinical manifestations of HNF1β-associated disease for the nephrologist.

Chronic metabolic acidosis results in renal Ca²⁺ and Mg²⁺ wasting, whereas chronic metabolic alkalosis is known to exert the reverse effects. It was hypothesized that these adaptations are mediated at least in part by the renal Ca²⁺ and Mg²⁺ transport proteins. The aim of this study, therefore, was to determine the effect of systemic acid-base status on renal expression of the epithelial Ca²⁺ channel TRPV5, the Ca²⁺-binding protein calbindin-D_28K, and the epithelial Mg²⁺ channel TRPM6 in relation to Ca²⁺ and Mg²⁺ excretion. Chronic metabolic acidosis that was induced by NH₄Cl loading or administration of the carbonic anhydrase inhibitor acetazolamide for 6 d enhanced calciuresis accompanied by decreased renal TRPV5 and calbindin-D_28K mRNA and protein abundance in wild-type mice. In contrast, metabolic acidosis did not affect Ca²⁺ excretion in TRPV5 knockout (TRPV5^−/−) mice, in which active Ca²⁺ reabsorption is effectively abolished. This demonstrates that downregulation of renal Ca²⁺ transport proteins is responsible for the hypercalciuria. Conversely, chronic metabolic alkalosis that was induced by NaHCO₃ administration for 6 d increased the expression of Ca²⁺ transport proteins accompanied by diminished urine Ca²⁺ excretion in wild-type mice. However, this Ca²⁺-sparing action persisted in TRPV5^−/− mice, suggesting that additional mechanisms apart from upregulation of active Ca²⁺ transport contribute to the hypocalciuria. Furthermore, chronic metabolic acidosis decreased renal TRPM6 expression, increased Mg²⁺ excretion, and decreased serum Mg²⁺ concentration, whereas chronic metabolic alkalosis resulted in the exact opposite effects. In conclusion, these data suggest that regulation of Ca²⁺ and Mg²⁺ transport proteins contributes importantly to the effects of acid-base status on renal divalent handling.