Distinguishing among HCO 3- , CO 3= , and H + as Substrates of Proteins That Appear To Be "Bicarbonate" Transporters.

BACKGROUND: Differentiating among HCO 3- , CO 3= , and H + movements across membranes has long seemed impossible. We now seek to discriminate unambiguously among three alternate mechanisms: the inward flux of 2 HCO 3- (mechanism 1), the inward flux of 1 CO 3= (mechanism 2), and the CO 2 /HCO 3- -stimulated outward flux of 2 H + (mechanism 3). METHODS: As a test case, we use electrophysiology and heterologous expression in Xenopus oocytes to examine SLC4 family members that appear to transport "bicarbonate" ("HCO 3- "). RESULTS: First, we note that cell-surface carbonic anhydrase should catalyze the forward reaction CO 2 +OH - →HCO 3- if HCO 3- is the substrate; if it is not, the reverse reaction should occur. Monitoring changes in cell-surface pH ( Δ pH S ) with or without cell-surface carbonic anhydrase, we find that the presumed Cl-"HCO 3 " exchanger AE1 (SLC4A1) does indeed transport HCO 3- (mechanism 1) as long supposed, whereas the electrogenic Na/"HCO 3 " cotransporter NBCe1 (SLC4A4) and the electroneutral Na + -driven Cl-"HCO 3 " exchanger NDCBE (SLC4A8) do not. Second, we use mathematical simulations to show that each of the three mechanisms generates unique quantities of H + at the cell surface (measured as Δ pH S ) per charge transported (measured as change in membrane current, ΔIm ). Calibrating ΔpH S /Δ Im in oocytes expressing the H + channel H V 1, we find that our NBCe1 data align closely with predictions of CO 3= transport (mechanism 2), while ruling out HCO 3- (mechanism 1) and CO 2 /HCO 3- -stimulated H + transport (mechanism 3). CONCLUSIONS: Our surface chemistry approach makes it possible for the first time to distinguish among HCO 3- , CO 3= , and H + fluxes, thereby providing insight into molecular actions of clinically relevant acid-base transporters and carbonic-anhydrase inhibitors.

Left Ventricular Systolic Dysfunction in NBCe1-B/C-Knockout Mice.

Congenital proximal renal tubular acidosis (pRTA) is a rare systemic disease caused by mutations in the SLC4A4 gene that encodes the electrogenic sodium bicarbonate cotransporter, NBCe1. The major NBCe1 protein variants are designated NBCe1-A, NBCe1-B, and NBCe1-C. NBCe1-A expression is kidney-specific, NBCe1-B is broadly expressed and is the only NBCe1 variant expressed in the heart, and NBCe1-C is a splice variant of NBCe1-B that is expressed in the brain. No cardiac manifestations have been reported from patients with pRTA, but studies in adult rats with virally induced reduction in cardiac NBCe1-B expression indicate that NBCe1-B loss leads to cardiac hypertrophy and prolonged QT intervals in rodents. NBCe1-null mice die shortly after weaning, so the consequence of congenital, global NBCe1 loss on the heart is unknown. To circumvent this issue, we characterized the cardiac function of NBCe1-B/C-null (KOb/c) mice that survive up to 2 months of age and which, due to the uninterrupted expression of NBCe1-A, do not exhibit the confounding acidemia of the globally null mice. In contrast to the viral knockdown model, cardiac hypertrophy was not present in KOb/c mice as assessed by heart-weight-to-body-weight ratios and cardiomyocyte cross-sectional area. However, echocardiographic analysis revealed reduced left ventricular ejection fraction, and intraventricular pressure-volume measurements demonstrated reduced load-independent contractility. We also observed increased QT length variation in KOb/c mice. Finally, using the calcium indicator Fura-2 AM, we observed a significant reduction in the amplitude of Ca2+ transients in paced KOb/c cardiomyocytes. These data indicate that congenital, global absence of NBCe1-B/C leads to impaired cardiac contractility and increased QT length variation in juvenile mice. It remains to be determined whether the cardiac phenotype in KOb/c mice is influenced by the absence of NBCe1-B/C from neuronal and endocrine tissues.

Corneal dystrophy mutations R125H and R804H disable SLC4A11 by altering the extracellular pH dependence of the intracellular pK that governs H+(OH-) transport.

Mutations in the H+(OH-) conductor SLC4A11 result in corneal endothelial dystrophy. In previous studies using mouse Slc4a11, we showed that the pK value that governs the intracellular pH dependence of SLC4A11 (pKi) is influenced by extracellular pH (pHe). We also showed that some mutations result in acidic or alkaline shifts in pKi, indicating that the pH dependence of SLC4A11 is important for physiological function. An R125H mutant, located in the cytosolic amino terminus of SLC4A11, apparently causes a complete loss of function, yet the anion transport inhibitor 4,4'-diisothiocyano-2,2'-stilbenedisulfonic acid (DIDS) can partially rescue SLC4A11/R125H activity. In the present study we set out to determine whether the effect of R125H is explained by an extreme shift in pKi. In Xenopus oocytes, we measured SLC4A11-mediated H+(OH-) conductance while monitoring pHi. We find that 1) the human corneal variant SLC4A11-B has a more acidic pKi than mouse Slc4a11, likely due to the presence of an NH2-terminal appendage; 2) pKi for human SLC4A11 is acid-shifted by raising pHe to 10.00; and 3) R125H and R804H mutants mediate substantial H+(OH-) conductances at pHe = 10.00, with pKi shifted into the wild-type range. These data suggest that the defect in each is a shift in pKi at physiological pHe, brought about by a disconnection in the mechanisms by which pHe influences pKi. Using de novo modeling, we show that R125 is located at the cytosolic dimer interface and suggest that this interface is critical for relaying the influence of pHe on the external face of the transmembrane domain to the intracellular, pKi-determining regions.

Revisiting the Role of Ser982 Phosphorylation in Stoichiometry Shift of the Electrogenic Na+/qHCO3- Cotransporter NBCe1.

In most cell types and heterologous expression systems, the electrogenic sodium-bicarbonate cotransporter NBCe1 operates with a 1Na+-2HCO3- stoichiometry that, given typical transmembrane electrochemical gradients, promotes Na+ and HCO3- influx. However, NBCe1 in the kidney mediates HCO3- efflux (HCO3- reabsorption), a direction that has been predicted to be favored only if NBCe1 operates with a 1:3 stoichiometry. The phosphorylation state of Ser982 in the cytosolic carboxy-terminal domain of NBCe1 has been reported to be a key determinant of the transporter stoichiometry, with non-phosphorylated Ser982 favoring a 1:3 stoichiometry. Conversely, phosphoproteomic data from renal cortical preparations have revealed the presence of NBCe1 peptides including phosphoserine982 (pSer982) and/or pSer985 although it was not known what proportion of NBCe1 molecules were phosphorylated. In the present study, we report the generation, characterization, and application of a novel phosphospecific antibody raised against NBCe1/pSer982 and show that, contrary to expectations, Ser982 is more prevalently phosphorylated in murine kidneys (in which NBCe1 mediates HCO3- efflux) than in murine colons (in which NBCe1 mediates HCO3- influx). Using phosphomimetic mutants of murine NBCe1 expressed in Xenopus oocytes, we found no evidence that the phosphorylation state of Ser982 or Ser985 alone influences the transport stoichiometry or conductance. Furthermore, we found that the phosphorylation of NBCe1/Ser982 is enhanced in murine kidneys following a 24 h induction of metabolic acidosis. We conclude that the phosphorylation status of Ser982 is not a key determinant of NBCe1 stoichiometry but correlates with presumed NBCe1 activity.

pH dependence of the Slc4a11-mediated H+ conductance is influenced by intracellular lysine residues and modified by disease-linked mutations.

SLC4A11 is the only member of the SLC4 family that transports protons rather than bicarbonate. SLC4A11 is expressed in corneal endothelial cells, and its mutation causes corneal endothelial dystrophy, although the mechanism of pathogenesis is unknown. We previously demonstrated that the magnitude of the H+ conductance (Gm) mediated by SLC4A11 is increased by rises in intracellular as well as extracellular pH (pHi and pHe). To better understand this feature and whether it is altered in disease, we studied the pH dependence of wild-type and mutant mouse Slc4a11 expressed in Xenopus oocytes. Using voltage-clamp circuitry in conjunction with a H+-selective microelectrode and a microinjector loaded with NaHCO3, we caused incremental rises in oocyte pHi and measured the effect on Gm. We find that the rise of Gm has a steeper pHi dependence at pHe =8.50 than at pHe =7.50. Data gathered at pHe =8.50 can be fit to the Hill equation enabling the calculation of a pK value that reports pHi dependence. We find that mutation of lysine residues that are close to the first transmembrane span (TM1) causes an alkaline shift in pK. Furthermore, two corneal-dystrophy-causing mutations close to the extracellular end of TM1, E399K and T401K (E368K and T370K in mouse), cause an acidic shift in pK, while a third mutation in the fourth intracellular loop, R804H (R774H in mouse), causes an alkaline shift in pK. This is the first description of determinants of SLC4A11 pH dependence and the first indication that a shift in pH dependence could modify disease expressivity in some cases of corneal dystrophy.

Multiple acid-base and electrolyte disturbances upregulate NBCn1, NBCn2, IRBIT and L-IRBIT in the mTAL.

KEY POINTS: The roles of the Na+ /HCO3- cotransporters NBCn1 and NBCn2 as well as their activators IRBIT and L-IRBIT in the regulation of the mTAL transport of NH4+ , HCO3- , and NaCl are investigated. Dietary challenges of NH4 Cl, NaHCO3 or NaCl all increase the abundance of NBCn1 and NBCn2 in the outer medulla. The three challenges generally produce parallel increases in the abundance of IRBIT and L-IRBIT in the outer medulla. Both IRBIT and L-IRBIT powerfully stimulate the activities of the mTAL isoforms of NBCn1 and NBCn2 as expressed in Xenopus oocytes. Our findings support the hypothesis that NBCn1, NBCn2, IRBIT and L-IRBIT appropriately promote NH4+ shunting but oppose HCO3- and NaCl reabsorption in the mTAL, and thus are at the nexus of the regulation pathways for multiple renal transport processes. ABSTRACT: The medullary thick ascending limb (mTAL) plays a key role in urinary acid and NaCl excretion. NBCn1 and NBCn2 are present in the basolateral mTAL, where NBCn1 promotes NH4+ shunting. IRBIT and L-IRBIT (the IRBITs) are two powerful activators of certain acid-base transporters. Here we use western blotting and immunofluorescence to examine the effects of multiple acid-base and electrolyte disturbances on expression of NBCn1, NBCn2 and the IRBITs in rat kidney. We also use electrophysiology to examine the functional effects of IRBITs on NBCn1 and NBCn2 in Xenopus oocytes. NH4 Cl-induced metabolic acidosis (MAc) substantially increases protein expression of NBCn1 and NBCn2 in the outer medulla (OM) of rat kidney. Surprisingly, NaHCO3 -induced metabolic alkalosis (MAlk) and high-salt diet (HSD) also increase expression of NBCn1 and NBCn2 (effect of NaHCO3  > HSD). Moreover, all three challenges generally increase OM expression of the IRBITs. In Xenopus oocytes, the IRBITs substantially increase the activities of NBCn1 and NBCn2. We propose that upregulation of basolateral NBCn1 and NBCn2 plus the IRBITs in the mTAL: (1) promotes NH4+ shunting by increasing basolateral HCO3- uptake to neutralize apical NH4+ uptake during MAc; (2) inhibits HCO3- reabsorption during MAlk by opposing HCO3- efflux via the basolateral anion exchanger AE2; and (3) inhibits NaCl reabsorption by mediating (with AE2) net NaCl backflux into the mTAL cell during HSD. Thus, NBCn1, NBCn2 and the IRBITs are at the nexus of the regulatory pathways for multiple renal transport processes.

The divergence, actions, roles, and relatives of sodium-coupled bicarbonate transporters.

The mammalian Slc4 (Solute carrier 4) family of transporters is a functionally diverse group of 10 multi-spanning membrane proteins that includes three Cl-HCO3 exchangers (AE1-3), five Na(+)-coupled HCO3(-) transporters (NCBTs), and two other unusual members (AE4, BTR1). In this review, we mainly focus on the five mammalian NCBTs-NBCe1, NBCe2, NBCn1, NDCBE, and NBCn2. Each plays a specialized role in maintaining intracellular pH and, by contributing to the movement of HCO3(-) across epithelia, in maintaining whole-body pH and otherwise contributing to epithelial transport. Disruptions involving NCBT genes are linked to blindness, deafness, proximal renal tubular acidosis, mental retardation, and epilepsy. We also review AE1-3, AE4, and BTR1, addressing their relevance to the study of NCBTs. This review draws together recent advances in our understanding of the phylogenetic origins and physiological relevance of NCBTs and their progenitors. Underlying these advances is progress in such diverse disciplines as physiology, molecular biology, genetics, immunocytochemistry, proteomics, and structural biology. This review highlights the key similarities and differences between individual NCBTs and the genes that encode them and also clarifies the sometimes confusing NCBT nomenclature.

Contribution of Monocarboxylate Transporter 6 to the Pharmacokinetics and Pharmacodynamics of Bumetanide in Mice.

Bumetanide, a sulfamyl loop diuretic, is used for the treatment of edema in association with congestive heart failure. Being a polar, anionic compound at physiologic pH, bumetanide uptake and efflux into different tissues is largely transporter-mediated. Of note, organic anion transporters (SLC22A) have been extensively studied in terms of their importance in transporting bumetanide to its primary site of action in the kidney. The contribution of one of the less-studied bumetanide transporters, monocarboxylate transporter 6 (MCT6; SLC16A5), to bumetanide pharmacokinetics (PK) and pharmacodynamics (PD) has yet to be characterized. The affinity of bumetanide for murine Mct6 was evaluated using Mct6-transfected Xenopus laevis oocytes. Furthermore, bumetanide was intravenously and orally administered to wild-type mice (Mct6+/+) and homozygous Mct6 knockout mice (Mct6-/-) to elucidate the contribution of Mct6 to bumetanide PK/PD in vivo. We demonstrated that murine Mct6 transports bumetanide at a similar affinity compared with human MCT6 (78 and 84 µM, respectively, at pH 7.4). After bumetanide administration, there were no significant differences in plasma PK. Additionally, diuresis was significantly decreased by ∼55% after intravenous bumetanide administration in Mct6-/- mice. Kidney cortex concentrations of bumetanide were decreased, suggesting decreased Mct6-mediated bumetanide transport to its site of action in the kidney. Overall, these results suggest that Mct6 does not play a major role in the plasma PK of bumetanide in mice; however, it significantly contributes to bumetanide's pharmacodynamics due to changes in kidney concentrations. SIGNIFICANCE STATEMENT: Previous evidence suggested that MCT6 transports bumetanide in vitro; however, no studies to date have evaluated the in vivo contribution of this transporter. In vitro studies indicated that mouse and human MCT6 transport bumetanide with similar affinities. Using Mct6 knockout mice, we demonstrated that murine Mct6 does not play a major role in the plasma pharmacokinetics of bumetanide; however, the pharmacodynamic effect of diuresis was attenuated in the knockout mice, likely because of the decreased bumetanide concentrations in the kidney.

Extrarenal Signs of Proximal Renal Tubular Acidosis Persist in Nonacidemic Nbce1b/c-Null Mice.

BACKGROUND: The SLC4A4 gene encodes electrogenic sodium bicarbonate cotransporter 1 (NBCe1). Inheritance of recessive mutations in SLC4A4 causes proximal renal tubular acidosis (pRTA), a disease characterized by metabolic acidosis, growth retardation, ocular abnormalities, and often dental abnormalities. Mouse models of pRTA exhibit acidemia, corneal edema, weak dental enamel, impacted colons, nutritional defects, and a general failure to thrive, rarely surviving beyond weaning. Alkali therapy remains the preferred treatment for pRTA, but it is unclear which nonrenal signs are secondary to acidemia and which are a direct consequence of NBCe1 loss from nonrenal sites (such as the eye and enamel organ) and therefore require separate therapy. SLC4A4 encodes three major NBCe1 variants: NBCe1-A, NBCe1-B, and NBCe1-C. NBCe1-A is expressed in proximal tubule epithelia; its dysfunction causes the plasma bicarbonate insufficiency that underlies acidemia. NBCe1-B and NBCe1-C exhibit a broad extra-proximal-tubular distribution. METHODS: To explore the consequences of Nbce1b/c loss in the absence of acidemia, we engineered a novel strain of Nbce1b/c-null mice and assessed them for signs of pRTA. RESULTS: Nbce1b/c-null mice have normal blood pH, but exhibit increased mortality, growth retardation, corneal edema, and tooth enamel defects. CONCLUSIONS: The correction of pRTA-related acidemia should not be considered a panacea for all signs of pRTA. The phenotype of Nbce1b/c-null mice highlights the physiologic importance of NBCe1 variants expressed beyond the proximal tubular epithelia and potential limitations of pH correction by alkali therapy in pRTA. It also suggests a novel genetic locus for corneal dystrophy and enamel hypomineralization without acidemia.

Soft drink consumption during and following exercise in the heat elevates biomarkers of acute kidney injury.

The purpose of this study was to test the hypothesis that consuming a soft drink (i.e., a high-fructose, caffeinated beverage) during and following exercise in the heat elevates biomarkers of acute kidney injury (AKI) in humans. Twelve healthy adults drank 2 liters of an assigned beverage during 4 h of exercise in the heat [35.1 (0.1)°C, 61 (5)% relative humidity] in counterbalanced soft drink and water trials, and ≥1 liter of the same beverage after leaving the laboratory. Stage 1 AKI (i.e., increased serum creatinine ≥0.30 mg/dl) was detected at postexercise in 75% of participants in the Soft Drink trial compared with 8% in Water trial ( P = 0.02). Furthermore, urinary neutrophil gelatinase-associated lipocalin (NGAL), a biomarker of AKI, was higher during an overnight collection period after the Soft Drink trial compared with Water in both absolute concentration [6 (4) ng/dl vs. 5 (4) ng/dl, P < 0.04] and after correcting for urine flow rate [6 (7) (ng/dl)/(ml/min) vs. 4 (4) (ng/dl)/(ml/min), P = 0.03]. Changes in serum uric acid from preexercise were greater in the Soft Drink trial than the Water trial at postexercise ( P < 0.01) and 24 h ( P = 0.05). There were greater increases from preexercise in serum copeptin, a stable marker of vasopressin, at postexercise in the Soft Drink trial ( P < 0.02) than the Water trial. These findings indicate that consuming a soft drink during and following exercise in the heat induces AKI, likely via vasopressin-mediated mechanisms.

Increasing Medical Student Interest in Nephrology.

BACKGROUND: Interest in nephrology careers is declining, possibly due to perceptions of the field and/or training aspects. Understanding practices of medical schools successfully instilling nephrology interest could inform efforts to attract leading candidates to the specialty. METHODS: The American Society of Nephrology Workforce Committee's Best Practices Project was one of several initiatives to increase nephrology career interest. Board-certified nephrologists graduating medical school between 2002 and 2009 were identified in the American Medical Association Masterfile and their medical schools ranked by production. Renal educators from the top 10 producing institutions participated in directed focus groups inquiring about key factors in creating nephrology career interest, including aspects of their renal courses, clinical rotations, research activities, and faculty interactions. Thematic content analysis of the transcripts (with inductive reasoning implementing grounded theory) was performed to identify factors contributing to their programs' success. RESULTS: The 10 schools identified were geographically representative, with similar proportions of graduates choosing internal medicine (mean 26%) as the national graduating class (26% in the 2017 residency Match). Eighteen educators from 9 of these 10 institutions participated. Four major themes were identified contributing to these schools' success: (1) nephrology faculty interaction with medical students; (2) clinical exposure to nephrology and clinical relevance of renal pathophysiology materials; (3) use of novel educational modalities; and (4) exposure, in particular early exposure, to the breadth of nephrology practice. CONCLUSION: Early and consistent exposure to a range of clinical nephrology experiences and nephrology faculty contact with medical students are important to help generate interest in the specialty.

Mouse models of SLC4-linked disorders of HCO3--transporter dysfunction.

The SLC4 family Cl-/[Formula: see text] cotransporters (NBCe1, NBCe2, NBCn1, and NBCn2) contribute to a variety of vital physiological processes including pH regulation and epithelial fluid secretion. Accordingly, their dysfunction can have devastating effects. Disorders such as epilepsy, hemolytic anemia, glaucoma, hearing loss, osteopetrosis, and renal tubular acidosis are all genetically linked to SLC4-family gene loci. This review summarizes how studies of Slc4-modified mice have enhanced our understanding of the etiology of SLC4-linked pathologies and the interpretation of genetic linkage studies. The review also surveys the novel disease signs exhibited by Slc4-modified mice which could either be considered to presage their description in humans, or to highlight interspecific differences. Finally, novel Slc4-modified mouse models are proposed, the study of which may further our understanding of the basis and treatment of SLC4-linked disorders of [Formula: see text]-transporter dysfunction.

Quality measures in acute kidney injury.

PURPOSE OF REVIEW: Quality measure assessment and reporting is evolving in end-stage renal disease care and is inchoate in ambulatory nephrology clinic care. Acute kidney injury (AKI) quality measures have not received sufficient attention, yet deserve consideration in view of the substantial proportion of effort nephrology providers devote to AKI care. RECENT FINDINGS: Accumulating literature permits consideration of timing of nephrology consultation, follow-up after AKI hospitalization, early detection, medication dosing, hospital readmissions and length of stay, cost, and mortality as potential AKI quality measures. SUMMARY: We review candidate AKI quality measures and assess the strength of evidence supporting the use of each measure as a standard for AKI care.

Education in Nephrology Fellowship: A Survey-Based Needs Assessment.

Educational needs assessments for nephrology fellowship training are limited. This study assessed fellows' perceptions of current educational needs and interest in novel modalities that may improve their educational experience and quantified educational resources used by programs and fellows. We distributed a seven-question electronic survey to all United States-based fellows receiving complimentary American Society of Nephrology (ASN) membership at the end of the 2015-2016 academic year in conjunction with the ASN Nephrology Fellows Survey. One third (320 of 863; 37%) of fellows in Accreditation Council for Graduate Medical Education-accredited positions responded. Most respondents rated overall quality of teaching in fellowship as either "good" (37%) or "excellent" (44%), and most (55%) second-year fellows felt "fully prepared" for independent practice. Common educational resources used by fellows included UpToDate, Journal of the American Society of Nephrology/Clinical Journal of the American Society of Nephrology, and Nephrology Self-Assessment Program; others-including ASN's online curricula-were used less often. Fellows indicated interest in additional instruction in several core topics, including home dialysis modalities, ultrasonography, and pathology. Respondents strongly supported interventions to improve pathology instruction and increase time for physiology and clinical review. In conclusion, current nephrology fellows perceive several gaps in training. Innovation in education and training is needed to better prepare future nephrologists for the growing challenges of kidney care.

Quercetin, Morin, Luteolin, and Phloretin Are Dietary Flavonoid Inhibitors of Monocarboxylate Transporter 6.

Monocarboxylate transporter 6 (MCT6; SLC16A5) has been recognized for its role as a xenobiotic transporter, with characterized substrates probenecid, bumetanide, and nateglinide. To date, the impact of commonly ingested dietary compounds on MCT6 function has not been investigated, and therefore, the objective of this study was to evaluate a variety of flavonoids for their potential MCT6-specific interactions. Flavonoids are a large group of polyphenolic phytochemicals found in commonly consumed plant-based products that have been recognized for their dietary health benefits. The uptake of bumetanide in human MCT6 gene-transfected Xenopus laevis oocytes was significantly decreased in the presence of a variety of flavonoids (e.g., quercetin, luteolin, phloretin, and morin), but was not significantly affected by flavonoid glycosides (e.g., naringin, rutin, phlorizin). The IC50 values of quercetin, phloretin, and morin were determined to be 25.3 ± 3.36, 17.3 ± 2.37, and 33.1 ± 3.29 µM, respectively. The mechanism of inhibition of phloretin was reversible and competitive, with a Ki value of 22.8 µM. Furthermore, typical MCT substrates were also investigated for their potential interactions with MCT6. Substrates of MCTs 1, 2, 4, 8, and 10 did not cause any significant decrease in MCT6-mediated bumetanide uptake, suggesting that MCT6 has distinct compound selectivity. In summary, these results suggest that dietary aglycon flavonoids may significantly alter the pharmacokinetics and pharmacodynamics of bumetanide and other MCT6-specific substrates, and may represent potential substrates for MCT6.

Enhancing Nephrology Career Interest through the ASN Kidney TREKS Program.

The Kidney Tutored Research and Education for Kidney Students (TREKS) Program is a product of the American Society of Nephrology (ASN) Workforce Committee that seeks to connect medical and graduate students to nephrology. This program starts with a weeklong camp-like course introducing participants to renal physiology through classic and modern experiments. Next, each student is matched with a nephrology mentor at his or her home institution to foster a better understanding of a nephrology career. Lastly, the students are encouraged to participate in scholarly activities and attend the ASN Kidney Week. Now in its third year, with a total of 84 participants, survey data suggest early success of the program, with a self-reported 40% increased interest in nephrology fellowship and/or research careers. In addition, students give high ratings to the course components and mentorship pairings. Continued student tracking will be necessary to determine the long-term program effect.

Mouse Slc4a11 expressed in Xenopus oocytes is an ideally selective H+/OH- conductance pathway that is stimulated by rises in intracellular and extracellular pH.

The SLC4A11 gene encodes the bicarbonate-transporter-related protein BTR1, which is mutated in syndromes characterized by vision and hearing loss. Signs of these diseases [congenital hereditary endothelial dystrophy (CHED) and Harboyan syndrome] are evident in mouse models of Slc4a11 disruption. However, the intrinsic activity of Slc4a11 remains controversial, complicating assignment of its (patho)physiological role. Most studies concur that Slc4a11 transports H+ (or the thermodynamically equivalent species OH-) rather than HCO3-, but disparities have arisen as to whether the transport is coupled to another species such as Na+ or NH3/NH4+ Here for the first time, we examine the action of mouse Slc4a11 in Xenopus oocytes. We simultaneously monitor changes in intracellular pH, membrane potential, and conductance as we alter extracellular pH, revealing the electrical and chemical driving forces that underlie the observed ion fluxes. We find that mSlc4a11 is an ideally selective H+/OH- conductive pathway, the action of which is uncoupled from the cotransport of any other ion. We also find that the activity of mSlc4a11 is independently enhanced by both extracellular and intracellular alkalinization, suggesting OH- as the most likely substrate and providing a novel explanation for the apparent NH3-dependence of Slc4a11-mediated currents reported by others. We suggest that the unique properties of Slc4a11 action underlie its value as a pH regulator in corneal endothelial cells.

A novel mutant Na+ /HCO3- cotransporter NBCe1 in a case of compound-heterozygous inheritance of proximal renal tubular acidosis.

KEY POINTS: The inheritance of two defective alleles of SLC4A4, the gene that encodes the widely-expressed electrogenic sodium bicarbonate cotransporter NBCe1, results in the bicarbonate-wasting disease proximal renal tubular acidosis (pRTA). In the present study, we report the first case of compound-heterozygous inheritance of pRTA (p.Arg510His/p.Gln913Arg) in an individual with low blood pH, blindness and neurological signs that resemble transient ischaemic attacks. We employ fluorescence microscopy on non-polarized (human embryonic kidney) and polarized (Madin-Darby canine kidney) renal cell lines and electrophysiology on Xenopus oocytes to characterize the mutant transporters (R510H and Q913R). Both mutant transporters exhibit enhanced intracellular retention in renal cells, an observation that probably explains the HCO3- transport deficit in the individual. Both mutants retain a close-to-normal per molecule Na+ /HCO3- cotransport activity in Xenopus oocytes, suggesting that they are suitable candidates for folding-correction therapy. However, Q913R expression is uniquely associated with a depolarizing, HCO3- independent, Cl- -conductance in oocytes that could have pathological consequences if expressed in the cells of patients. ABSTRACT: Proximal renal tubular acidosis (pRTA) is a rare, recessively-inherited disease characterized by abnormally acidic blood, blindness, as well as below average height and weight. pRTA is typically associated with homozygous mutation of the solute carrier 4 family gene SLC4A4. SLC4A4 encodes the electrogenic sodium bicarbonate cotransporter NBCe1, a membrane protein that acts to maintain intracellular and plasma pH. We present the first description of a case of compound-heterozygous inheritance of pRTA. The individual has inherited two mutations in NBCe1: p.Arg510His (R510H) and p.Gln913Arg (Q913R), one from each parent. In addition to the usual features of pRTA, the patient exhibits unusual signs, such as muscle spasms and fever. We have recreated these mutant transporters for expression in model systems. We find that both of the mutant proteins exhibit substantial intracellular retention when expressed in mammalian renal cell lines. When expressed in Xenopus oocytes, we find that the R510H and Q913R-mutant NBCe1 molecules exhibit apparently normal Na+ /HCO3- cotransport activity but that Q913R is associated with an unusual HCO3- independent anion-leak. We conclude that a reduced accumulation of NBCe1 protein in the basolateral membrane of proximal-tubule epithelia is the most probable cause of pRTA in this case. We further note that the Q913R-associated anion-leak could itself be pathogenic if expressed in the plasma membrane of mammalian cells, compromising the benefit of strategies aiming to enhance mutant NBCe1 accumulation in the plasma membrane.

Na+-H+ exchanger-1 (NHE1) regulation in kidney proximal tubule.

The ubiquitously expressed plasma membrane Na(+)-H(+) exchanger NHE1 is a 12 transmembrane-spanning protein that directs important cell functions such as homeostatic intracellular volume and pH control. The 315 amino acid cytosolic tail of NHE1 binds plasma membrane phospholipids and multiple proteins that regulate additional, ion-translocation independent functions. This review focuses on NHE1 structure/function relationships, as well as the role of NHE1 in kidney proximal tubule functions, including pH regulation, vectorial Na(+) transport, cell volume control and cell survival. The implications of these functions are particularly critical in the setting of progressive, albuminuric kidney diseases, where the accumulation of reabsorbed fatty acids leads to disruption of NHE1-membrane phospholipid interactions and tubular atrophy, which is a poor prognostic factor for progression to end stage renal disease. This review amplifies the vital role of the proximal tubule NHE1 Na(+)-H(+) exchanger as a kidney cell survival factor.

Improving the Nephrology Match: the Path Forward.

The Fellowship Match process was designed to provide applicants and program directors with an opportunity to consider all their options before making decisions about post-residency training. In a Match, applicants can choose the programs that best suit their career goals, and program directors can consider all candidates before preparing a rank order list. The Match is a contract, requiring obligations of both programs and applicants to achieve success, ensure uniformity, and standardize participation.