Controversies and practical management of patients with gout and chronic kidney disease.

Uric acid is a toxin retained with advancing kidney disease. Clinical manifestations of hyperuricemia include gout and systemic inflammation that are associated with increased risk for cardiovascular mortality. As many as one third of all patients with chronic kidney disease (CKD) have a history of gout, yet <25% of these patients are effectively treated to target serum urate levels of ≤6 mg/dL. A major reason for ineffective management of gout and hyperuricemia is the complexity in managing these patients, with some medications contraindicated, others requiring special dosing, potential drug interactions, and other factors. Consequently, many nephrologists do not primarily manage gout despite it being a common complication of CKD, leaving management to the primary physician or rheumatologist. We believe that kidney specialists should consider gout as a major complication of CKD and actively manage it in their patients. Here, we present insights from nephrologists and rheumatologists on a team approach to gout management that includes the nephrologist.

Genetic and Physiological Effects of Insulin-Like Growth Factor-1 (IGF-1) on Human Urate Homeostasis.

SIGNIFICANCE STATEMENT: Hyperinsulinemia induces hyperuricemia by activating net renal urate reabsorption in the renal proximal tubule. The basolateral reabsorptive urate transporter GLUT9a appears to be the dominant target for insulin. By contrast, IGF-1 infusion reduces serum urate (SU), through mechanisms unknown. Genetic variants of IGF1R associated with reduced SU have increased IGF-1R expression and interact with genes encoding the GLUT9 and ABCG2 urate transporters, in a sex-specific fashion, which controls the SU level. Activation of IGF-1/IGF-1R signaling in Xenopus oocytes modestly activates GLUT9a and inhibits insulin's stimulatory effect on the transporter, which also activates multiple secretory urate transporters-ABCG2, ABCC4, OAT1, and OAT3. The results collectively suggest that IGF-1 reduces SU by activating secretory urate transporters and inhibiting insulin's action on GLUT9a. BACKGROUND: Metabolic syndrome and hyperinsulinemia are associated with hyperuricemia. Insulin infusion in healthy volunteers elevates serum urate (SU) by activating net urate reabsorption in the renal proximal tubule, whereas IGF-1 infusion reduces SU by mechanisms unknown. Variation within the IGF1R gene also affects SU levels. METHODS: Colocalization analyses of a SU genome-wide association studies signal at IGF1R and expression quantitative trait loci signals in cis using COLOC2, RT-PCR, Western blotting, and urate transport assays in transfected HEK 293T cells and in Xenopus laevis oocytes. RESULTS: Genetic association at IGF1R with SU is stronger in women and is mediated by control of IGF1R expression. Inheritance of the urate-lowering homozygous genotype at the SLC2A9 locus is associated with a differential effect of IGF1R genotype between men and women. IGF-1, through IGF-1R, stimulated urate uptake in human renal proximal tubule epithelial cells and transfected HEK 293T cells, through activation of IRS1, PI3/Akt, MEK/ERK, and p38 MAPK; urate uptake was inhibited in the presence of uricosuric drugs, specific inhibitors of protein tyrosine kinase, PI3 kinase (PI3K), ERK, and p38 MAPK. In X. laevis oocytes expressing ten individual urate transporters, IGF-1 through endogenous IGF-1R stimulated urate transport mediated by GLUT9, OAT1, OAT3, ABCG2, and ABCC4 and inhibited insulin's stimulatory action on GLUT9a and OAT3. IGF-1 significantly activated Akt and ERK. Specific inhibitors of PI3K, ERK, and PKC significantly affected IGF-1 stimulation of urate transport in oocytes. CONCLUSIONS: The combined results of infusion, genetics, and transport experiments suggest that IGF-1 reduces SU by activating urate secretory transporters and inhibiting insulin's action.

Genomic dissection of 43 serum urate-associated loci provides multiple insights into molecular mechanisms of urate control.

High serum urate is a prerequisite for gout and associated with metabolic disease. Genome-wide association studies (GWAS) have reported dozens of loci associated with serum urate control; however, there has been little progress in understanding the molecular basis of the associated loci. Here, we employed trans-ancestral meta-analysis using data from European and East Asian populations to identify 10 new loci for serum urate levels. Genome-wide colocalization with cis-expression quantitative trait loci (eQTL) identified a further five new candidate loci. By cis- and trans-eQTL colocalization analysis, we identified 34 and 20 genes, respectively, where the causal eQTL variant has a high likelihood that it is shared with the serum urate-associated locus. One new locus identified was SLC22A9 that encodes organic anion transporter 7 (OAT7). We demonstrate that OAT7 is a very weak urate-butyrate exchanger. Newly implicated genes identified in the eQTL analysis include those encoding proteins that make up the dystrophin complex, a scaffold for signaling proteins and transporters at the cell membrane; MLXIP that, with the previously identified MLXIPL, is a transcription factor that may regulate serum urate via the pentose-phosphate pathway and MRPS7 and IDH2 that encode proteins necessary for mitochondrial function. Functional fine mapping identified six loci (RREB1, INHBC, HLF, UBE2Q2, SFMBT1 and HNF4G) with colocalized eQTL containing putative causal SNPs. This systematic analysis of serum urate GWAS loci identified candidate causal genes at 24 loci and a network of previously unidentified genes likely involved in control of serum urate levels, further illuminating the molecular mechanisms of urate control.

Zoledronic Acid-Associated Fanconi Syndrome in Patients With Cancer.

Zoledronic acid (ZA) is an antiresorptive agent typically used for fracture prevention in postmenopausal osteoporosis, malignancy-associated metastatic bone lesions, and as a treatment for hypercalcemia. ZA is excreted almost entirely by the kidney; as a result, a reduction in renal clearance can lead to its accumulation and potential renal toxicity. Although uncommon, acute kidney injury (AKI) from intravenous bisphosphonates has been described, with different patterns including tubulointerstitial nephritis, acute tubular necrosis, as well as focal segmental glomerulosclerosis. Here we present 4 patients with an underlying malignancy who each developed evidence of generalized proximal tubular dysfunction, also known as Fanconi syndrome, approximately 1 week after receiving treatment with ZA. On presentation, all patients had AKI, low serum bicarbonate levels, abnormal urinary acidification, hypophosphatemia, hypokalemia, and increased urine amino acid excretion or renal glycosuria. Based on the temporal association between ZA infusion and the development of these electrolyte abnormalities, each case is highly suggestive of ZA-associated Fanconi syndrome. Due to the severity of presentation, all required discontinuation of ZA and ongoing electrolyte repletion. Nephrologists and oncologists should be aware of this complication and consider ZA as a possible trigger of new-onset Fanconi syndrome.

Mutations in HOGA1 do Not Confer a Dominant Phenotype Manifesting as Kidney Stone Disease.

PURPOSE: The etiology of calcium-oxalate kidney stone formation remains elusive. Biallelic mutations in HOGA1 are responsible for primary hyperoxaluria type 3 and result in oxalate overproduction and kidney stone disease. Our previous study showed that carriers of HOGA1 mutations have elevated urinary levels of oxalate precursors. In this study we explored the possibility that mutations in HOGA1 confer a dominant phenotype in the form of kidney stone disease or hyperoxaluria. MATERIALS AND METHODS: An observational analytic case control study was designed to determine the prevalence of pathogenic HOGA1 mutations among adults with calcium-oxalate kidney stone disease. Given the high prevalence of HOGA1 mutations among Ashkenazi Jews, this group was evaluated separately. Carrier frequency of any of the 52 reported pathogenic mutations was compared to data derived from gnomAD for the corresponding ethnic group. Sanger sequencing of HOGA1 gene was performed on DNA samples from the following groups: 60 Ashkenazi Jews and 86 nonAshkenazi calcium-oxalate stone formers, 150 subjects with low and 150 with high urinary oxalate levels. RESULTS: The carrier prevalence of pathogenic mutations among the Ashkenazi Jews was 1.7% compared to 2.8% in the corresponding control group (p=0.9 OR=0.6 95% CI 0.01-3.51). We did not detect any mutation among the nonAshkenazi study group. No correlation was detected between hyperoxaluria and HOGA1 variants. CONCLUSIONS: This study shows that mutations in HOGA1 do not confer a dominant phenotype in the form of calcium-oxalate kidney stone disease or hyperoxaluria.

Development of an electronic health record-based chronic kidney disease registry to promote population health management.

BACKGROUND: Electronic health record (EHR) based chronic kidney disease (CKD) registries are central to population health strategies to improve CKD care. In 2015, Partners Healthcare System (PHS), encompassing multiple academic and community hospitals and outpatient care facilities in Massachusetts, developed an EHR-based CKD registry to identify opportunities for quality improvement, defined as improvement on both process measures and outcomes measures associated with clinical care. METHODS: Patients are included in the registry based on the following criteria: 1) two estimated glomerular filtration rate (eGFR) results < 60 ml/min/1.73m2 separated by 90 days, including the most recent eGFR being < 60 ml/min/1.73m2; or 2) the most recent two urine protein values > 300 mg protein/g creatinine on either urine total protein/creatinine ratio or urine albumin/creatinine ratio; or 3) an EHR problem list diagnosis of end stage renal disease (ESRD). The registry categorizes patients by CKD stage and includes rates of annual testing for eGFR and proteinuria, blood pressure control, use of angiotensin converting enzyme inhibitors (ACE-Is) or angiotensin receptor blockers (ARBs), nephrotoxic medication use, hepatitis B virus (HBV) immunization, vascular access placement, transplant status, CKD progression risk; number of outpatient nephrology visits, and hospitalizations. RESULTS: The CKD registry includes 60,503 patients and has revealed several opportunities for care improvement including 1) annual proteinuria testing performed for 17% (stage 3) and 31% (stage 4) of patients; 2) ACE-I/ARB used in 41% (stage 3) and 46% (stage 4) of patients; 3) nephrotoxic medications used among 23% of stage 4 patients; and 4) 89% of stage 4 patients lack HBV immunity. For advanced CKD patients there are opportunities to improve vascular access placement, transplant referrals and outpatient nephrology contact. CONCLUSIONS: A CKD registry can identify modifiable care gaps across the spectrum of CKD care and enable population health strategy implementation. No linkage to Social Security Death Master File or US Renal Data System (USRDS) databases limits our ability to track mortality and progression to ESRD.

The molecular physiology of uric acid homeostasis.

Uric acid, generated from the metabolism of purines, has proven and emerging roles in human disease. Serum uric acid is determined by production and the net balance of reabsorption or secretion by the kidney and intestine. A detailed understanding of epithelial absorption and secretion of uric acid has recently emerged, aided in particular by the results of genome-wide association studies of hyperuricemia. Novel genetic and regulatory networks with effects on uric acid homeostasis have also emerged. These developments promise to lead to a new understanding of the various diseases associated with hyperuricemia and to novel, targeted therapies for hyperuricemia.

Hyperuricemia, Acute and Chronic Kidney Disease, Hypertension, and Cardiovascular Disease: Report of a Scientific Workshop Organized by the National Kidney Foundation.

Urate is a cause of gout, kidney stones, and acute kidney injury from tumor lysis syndrome, but its relationship to kidney disease, cardiovascular disease, and diabetes remains controversial. A scientific workshop organized by the National Kidney Foundation was held in September 2016 to review current evidence. Cell culture studies and animal models suggest that elevated serum urate concentrations can contribute to kidney disease, hypertension, and metabolic syndrome. Epidemiologic evidence also supports elevated serum urate concentrations as a risk factor for the development of kidney disease, hypertension, and diabetes, but differences in methodologies and inpacts on serum urate concentrations by even subtle changes in kidney function render conclusions uncertain. Mendelian randomization studies generally do not support a causal role of serum urate in kidney disease, hypertension, or diabetes, although interpretation is complicated by nonhomogeneous populations, a failure to consider environmental interactions, and a lack of understanding of how the genetic polymorphisms affect biological mechanisms related to urate. Although several small clinical trials suggest benefits of urate-lowering therapies on kidney function, blood pressure, and insulin resistance, others have been negative, with many trials having design limitations and insufficient power. Thus, whether uric acid has a causal role in kidney and cardiovascular diseases requires further study.

Insulin and SGK1 reduce the function of Na+/monocarboxylate transporter 1 (SMCT1/SLC5A8).

SMCTs move several important fuel molecules that are involved in lipid, carbohydrate, and amino acid metabolism, but their regulation has been poorly studied. Insulin controls the translocation of several solutes that are involved in energetic cellular metabolism, including glucose. We studied the effect of insulin on the function of human SMCT1 expressed in Xenopus oocytes. The addition of insulin reduced α-keto-isocaproate (KIC)-dependent 22Na+ uptake by 29%. Consistent with this result, the coinjection of SMCT1 with SGK1 cRNA decreased the KIC-dependent 22Na+ uptake by 34%. The reduction of SMCT1 activity by SGK1 depends on its kinase activity, and it was observed that the coinjection of SMCT1 with S442D-SGK1 (a constitutively active mutant) decreased the KIC-dependent 22Na+ uptake by 50%. In contrast, an SMCT1 coinjection with K127M-SGK1 (an inactive mutant) had no effect on the KIC-dependent Na+ uptake. The decreasing SMCT1 function by insulin or SGK1 was corroborated by measuring [1-14C]acetate uptake and the electric currents of SMCT1-injected oocytes. Previously, we found that SMCT2/Slc5a12-mRNA, but not SMCT1/Slc5a8-mRNA, is present in zebrafish pancreas (by in situ hybridization); however, SLC5a8 gene silencing was associated with the development of human pancreatic cancer. We confirmed that the mRNA and protein of both transporters were present in rat pancreas using RT-PCR with specific primers, Western blot analysis, and immunohistochemistry. Additionally, significant propionate-dependent 22Na+ uptake occurred in pancreatic islets and was reduced by insulin treatment. Our data indicate that human SMCT1 is regulated by insulin and SGK1 and that both SMCTs are present in the mammalian pancreas.

An electronic alert to decrease Kayexalate ordering.

Important safety concerns have recently emerged regarding the use of sodium polystyrene sulfonate (Kayexalate), a cation-exchange resin commonly used for the treatment of hyperkalemia. We implemented an electronic alert system at a tertiary care academic medical center to warn providers of the safety concerns of Kayexalate. We assessed the number of Kayexalate prescriptions per month, as well as the number of grams of Kayexalate ordered per month, one year before versus one year after implementing the alert. The mean (±SD) number of Kayexalate orders decreased from 123 (±12) to 76 (±14) orders/month (38% absolute reduction, p < 0.001) after implementing the alert. Additionally, the mean (±SD) amount of Kayexalate prescribed decreased from 3332 (±329) to 1885 (±358) g/month (43% absolute reduction, p < 0.001). We conclude that an electronic alert is an effective tool to decrease Kayexalate ordering.

Conformational change in transfer RNA is an early indicator of acute cellular damage.

Tissue damage by oxidative stress is a key pathogenic mechanism in various diseases, including AKI and CKD. Thus, early detection of oxidative tissue damage is important. Using a tRNA-specific modified nucleoside 1-methyladenosine (m1A) antibody, we show that oxidative stress induces a direct conformational change in tRNA structure that promotes subsequent tRNA fragmentation and occurs much earlier than DNA damage. In various models of tissue damage (ischemic reperfusion, toxic injury, and irradiation), the levels of circulating tRNA derivatives increased rapidly. In humans, the levels of circulating tRNA derivatives also increased under conditions of acute renal ischemia, even before levels of other known tissue damage markers increased. Notably, the level of circulating free m1A correlated with mortality in the general population (n=1033) over a mean follow-up of 6.7 years. Compared with healthy controls, patients with CKD had higher levels of circulating free m1A, which were reduced by treatment with pitavastatin (2 mg/d; n=29). Therefore, tRNA damage reflects early oxidative stress damage, and detection of tRNA damage may be a useful tool for identifying organ damage and forming a clinical prognosis.

Glycosylation regulates the function and membrane localization of KCC4.

Glycosylation plays a role in regulating many biological activities, including protein folding and cell surface expression of biomolecules. However, the importance of glycosylation for KCC4 function has not previously been demonstrated. Site-directed mutagenesis was performed on the four putative extracellular N-linked glycosylation sites of KCC4 to determine the role of these sites in KCC4 half-life, cell surface expression, and transporter activity, as well as in KCC4-dependent tumor formation. We showed that triple (N312/331/344/Q) and quadruple (N312/331/344/360/Q) mutations of N-linked glycosylation sites disrupt the N-linked glycosylation of KCC4, resulting in the accumulation of KCC4, predominantly in the endoplasmic reticulum (ER) and not at the cell surface. Further investigation indicated that mutations of the central two (N331/344/Q) N-linked glycosylation sites inhibit the membrane trafficking of KCC4. Our data suggest that the glycan moieties at the N331 and N344 sites were Endo H-resistant, complex-form structures, and that the N312 and N360 sites were Endo H-sensitive, high mannose-containing structures. Under hypotonic stress conditions, the ability to adapt to changes in intracellular chloride ion concentrations and RVD (regulatory volume decrease) activities were less efficient in cells containing the deglycosylated form of KCC4 that were not expressed at the cell surface. Deglycosylated forms of KCC4 also demonstrated decreased tumor formation and lung colonization in mouse xenografts. The difference in glycan complexity may account for the differential impact of each branch on the biological effects of KCC4. We propose that glycosylation is essential for the surface expression, stabilization, and bioactivity of KCC4.

Caffeine inhibits both basal and insulin-activated urate transport.

OBJECTIVE: Caffeine, an adenosine receptor antagonist, is a potent central nervous system stimulant that also impairs insulin signaling. Recent studies have suggested that coffee consumption lowers serum urate (SU) and protects against gout, by unknown mechanisms. We hypothesized that caffeine lowers serum urate by affecting activity of urate transporters. METHODS: We examined the effect of caffeine and adenosine on basal and insulin-stimulation of net 14C-urate uptake in the human renal proximal tubule cell line PTC-05, and on individual urate transporters expressed in Xenopus laevis oocytes. RESULTS: We found that caffeine and adenosine efficiently inhibited both basal and insulin-stimulation of net 14C-urate uptake mediated by endogenous urate transporters in PTC-05 cells. In oocytes expressing individual urate transporters, caffeine (>0.2 mM) more efficiently inhibited the basal urate transport activity of GLUT9 isoforms, OAT4, OAT1, OAT3, NPT1, ABCG2 and ABCC4 than did adenosine, without significantly affecting URAT1 and OAT10. However, unlike adenosine, caffeine at lower concentrations (<0.2 mM), very effectively inhibited insulin-activation of urate transport activity of GLUT9, OAT10, OAT1, OAT3, NPT1, ABCG2 and ABCC4 by blocking activation of Akt and ERK. CONCLUSIONS: We postulate that inhibition of urate transport activity of the reabsorptive transporters GLUT9, OAT10, and OAT4 by caffeine is a key mechanism in its urate-lowering effects. Additionally, the ability of caffeine to block insulin-activated urate transport by GLUT9a and OAT10 suggests greater relative inhibition of these transporters in hyperinsulinemia.

Hyperpolarizing GABAergic transmission requires the KCC2 C-terminal ISO domain.

KCC2 is the neuron-specific member of the of K(+)-Cl(-) cotransporter gene family. It is also the only member of its family that is active under physiologically normal conditions, in the absence of osmotic stress. By extruding Cl(-) from the neuron under isotonic conditions, this transporter maintains a low concentration of neuronal Cl(-), which is essential for fast inhibitory synaptic transmission by GABA and glycine in the mature nervous system. The other members of this K(+)-Cl(-) cotransporter gene family are exclusively swelling-activated. Here we demonstrate that a 15 aa region near the end of the C terminus, unique to KCC2 (termed the ISO domain), is required for KCC2 to cotransport K(+) and Cl(-) out of the neuron under isotonic conditions. We made this discovery by overexpressing chimeric KCC2-KCC4 cDNA constructs in cultured hippocampal neurons prepared from Sprague Dawley rat embryos and assaying neuronal Cl(-) through gramicidin perforated patch-clamp recordings. We found that when neurons had been transfected with a chimeric KCC2 that lacked the unique ISO domain, hyperpolarizing responses to GABA were abolished. This finding indicates that the ISO domain is required for neuronal Cl(-) regulation. Furthermore, we discovered that when KCC2 lacks the ISO domain, it still retains swelling-activated transport, which demonstrates that there are exclusive molecular determinants of isotonic and swelling-induced K(+)-Cl(-) cotransport in neurons.

Molecular evidence for a role for K(+)-Cl(-) cotransporters in the kidney.

K(+)-Cl(-) cotransporter (KCC) isoforms 3 (KCC3) and 4 (KCC4) are expressed at the basolateral membrane of proximal convoluted tubule cells, and KCC4 is present in the basolateral membrane of the thick ascending loop of Henle's limb and α-intercalated cells of the collecting duct. Little is known, however, about the physiological roles of these transporters in the kidney. We evaluated KCC3 and KCC4 mRNA and protein expression levels and intrarenal distribution in male Wistar rats or C57 mice under five experimental conditions: hyperglycemia after a single dose of streptozotocin, a low-salt diet, metabolic acidosis induced by ammonium chloride in drinking water, and low- or high-K(+) diets. Both KCC3 mRNA and protein expression were increased during hyperglycemia in the renal cortex and at the basolateral membrane of proximal tubule cells but not with a low-salt diet or acidosis. In contrast, KCC4 protein expression was increased by a low-sodium diet in the whole kidney and by metabolic acidosis in the renal outer medulla, specifically at the basolateral membrane of α-intercalated cells. The increased protein expression of KCC4 by a low-salt diet was also observed in WNK4 knockout mice, suggesting that upregulation of KCC4 in these circumstances is not WNK4 dependent. No change in KCC3 or KCC4 protein expression was observed under low- or high-K(+) diets. Our data are consistent with a role for KCC3 in the proximal tubule glucose reabsorption mechanism and for KCC4 in salt reabsorption of the thick ascending loop of Henle's loop and acid secretion of the collecting duct.

The kidney in hyperuricemia and gout.

PURPOSE OF REVIEW: Gout is a painful inflammatory arthritis associated with hyperuricemia, with a prevalence of almost 10 million in the USA. Reduced renal excretion of urate is the underlying hyperuricemic mechanism in the vast majority of gout patients; most of the genes that affect serum urate level (SUA) encode urate transporters or associated regulatory proteins. Acquired influences can also modulate SUA and renal urate excretion, sometimes precipitating acute gout. Coincidentally, the prevalence of renal comorbidities in gout - hypertension, chronic kidney disease (CKD), and nephrolithiasis - is very high. RECENT FINDINGS: Recent advances in genetics and molecular physiology have greatly enhanced the understanding of renal reabsorption and secretion of filtered urate. Moreover, baseline SUA appears to be set by the net balance of absorption and secretion across epithelial cells in the kidney and intestine. There have also been substantial advances in the management of gout in patients with CKD. SUMMARY: The stage is set for an increasingly molecular understanding of baseline and regulated urate transport by the kidney and intestine. The increasing prevalence of gout with CKD will be balanced by an expanding spectrum of therapeutic options for this important disease.

Pregnancy-associated polyuria in familial renal glycosuria.

A pregnant woman presented at gestational week 28 with loss of consciousness and profound polyuria. Further characterization revealed osmotic diuresis due to massive glycosuria without hyperglycemia. Glycosuria reduced substantially postpartum, from approximately 100 to approximately 30 g/1.73 m2 per day. DNA sequencing analysis of the SLC5A2 gene encoding the renal glucose transporter SGLT2 showed a homozygous frame-shift mutation (occurring after the glutamine at amino acid 168 and leading to premature termination of the protein at amino acid 186) diagnostic of familial renal glycosuria. Pregnant women with familial renal glycosuria can be at risk of profound polyuria during pregnancy due to the associated increase in glycosuria. These findings also have implications for the use of SGLT2 inhibitors in clinical practice.

Deficiency of the calcium-sensing receptor in the kidney causes parathyroid hormone-independent hypocalciuria.

Rare loss-of-function mutations in the calcium-sensing receptor (Casr) gene lead to decreased urinary calcium excretion in the context of parathyroid hormone (PTH)-dependent hypercalcemia, but the role of Casr in the kidney is unknown. Using animals expressing Cre recombinase driven by the Six2 promoter, we generated mice that appeared grossly normal but had undetectable levels of Casr mRNA and protein in the kidney. Baseline serum calcium, phosphorus, magnesium, and PTH levels were similar to control mice. When challenged with dietary calcium supplementation, however, these mice had significantly lower urinary calcium excretion than controls (urinary calcium to creatinine, 0.31±0.03 versus 0.63±0.14; P=0.001). Western blot analysis on whole-kidney lysates suggested an approximately four-fold increase in activated Na(+)-K(+)-2Cl(-) cotransporter (NKCC2). In addition, experimental animals exhibited significant downregulation of Claudin14, a negative regulator of paracellular cation permeability in the thick ascending limb, and small but significant upregulation of Claudin16, a positive regulator of paracellular cation permeability. Taken together, these data suggest that renal Casr regulates calcium reabsorption in the thick ascending limb, independent of any change in PTH, by increasing the lumen-positive driving force for paracellular Ca(2+) transport.

The K-Cl cotransporter KCC3 is mutant in a severe peripheral neuropathy associated with agenesis of the corpus callosum.

Peripheral neuropathy associated with agenesis of the corpus callosum (ACCPN) is a severe sensorimotor neuropathy associated with mental retardation, dysmorphic features and complete or partial agenesis of the corpus callosum. ACCPN is transmitted in an autosomal recessive fashion and is found at a high frequency in the province of Quebec, Canada. ACCPN has been previously mapped to chromosome 15q. The gene SLC12A6 (solute carrier family 12, member 6), which encodes the K+-Cl- transporter KCC3 and maps within the ACCPN candidate region, was screened for mutations in individuals with ACCPN. Four distinct protein-truncating mutations were found: two in the French Canadian population and two in non-French Canadian families. The functional consequence of the predominant French Canadian mutation (2436delG, Thr813fsX813) was examined by heterologous expression of wildtype and mutant KCC3 in Xenopus laevis oocytes; the truncated mutant is appropriately glycosylated and expressed at the cellular membrane, where it is non-functional. Mice generated with a targeted deletion of Slc12a6 have a locomotor deficit, peripheral neuropathy and a sensorimotor gating deficit, similar to the human disease. Our findings identify mutations in SLC12A6 as the genetic lesion underlying ACCPN and suggest a critical role for SLC12A6 in the development and maintenance of the nervous system.