Renal Ammonia Excretion Research Articles

State of knowledge on ammonia handling by the kidney.

The disposal of ammonia, the main proton buffer in the urine, is important for acid-base homeostasis. Renal ammonia excretion is the predominant contributor to renal net acid excretion, both under basal condition and in response to acidosis. New insights into the mechanisms of renal ammonia production and transport have been gained in the past decades. Ammonia is the only urinary solute known to be produced in the kidney and selectively transported through the different parts of the nephron. Both molecular forms of total ammonia, NH3 and NH4+, are transported by specific proteins. Proximal tubular ammoniagenesis and the activity of these transport processes determine the eventual fate of total ammonia produced and excreted by the kidney. In this review, we summarized the state of the art of ammonia handling by the kidney and highlighted the newest processes described in the last decade.

NBCe1-B/C-knockout mice exhibit an impaired respiratory response and an enhanced renal response to metabolic acidosis.

The sodium-bicarbonate cotransporter (NBCe1) has three primary variants: NBCe1-A, -B and -C. NBCe1-A is expressed in renal proximal tubules in the cortical labyrinth, where it is essential for reclaiming filtered bicarbonate, such that NBCe1-A knockout mice are congenitally acidemic. NBCe1-B and -C variants are expressed in chemosensitive regions of the brainstem, while NBCe1-B is also expressed in renal proximal tubules located in the outer medulla. Although mice lacking NBCe1-B/C (KOb/c) exhibit a normal plasma pH at baseline, the distribution of NBCe1-B/C indicates that these variants could play a role in both the rapid respiratory and slower renal responses to metabolic acidosis (MAc). Therefore, in this study we used an integrative physiologic approach to investigate the response of KOb/c mice to MAc. By means of unanesthetized whole-body plethysmography and blood-gas analysis, we demonstrate that the respiratory response to MAc (increase in minute volume, decrease in pCO2) is impaired in KOb/c mice leading to a greater severity of acidemia after 1day of MAc. Despite this respiratory impairment, the recovery of plasma pH after 3-days of MAc remained intact in KOb/c mice. Using data gathered from mice housed in metabolic cages we demonstrate a greater elevation of renal ammonium excretion and greater downregulation of the ammonia recycling enzyme glutamine synthetase in KOb/c mice on day 2 of MAc, consistent with greater renal acid-excretion. We conclude that KOb/c mice are ultimately able to defend plasma pH during MAc, but that the integrated response is disturbed such that the burden of work shifts from the respiratory system to the kidneys, delaying the recovery of pH.

The Role of the Androgen Receptor in Sexual Dimorphisms in Acidosis‐Stimulated Renal Ammonia Metabolism

Renal ammonia excretion is a critical mechanism through which the kidneys maintain acid‐base homeostasis. We have previously identified that ammonia excretion is less in male mice than in female mice under basal conditions, that this correlates with differences in expression of key proteins involved in ammonia metabolism, and involves both testosterone‐dependent and renal androgen receptor (AR)‐dependent signaling pathways. In addition, there are sex differences in the response to acid‐loading. This study’s objective was to determine renal AR’s role in the renal ammonia response to an exogenous acid load. To avoid the known systemic effects of AR blockade/deletion, we studied mice with kidney‐specific AR deletion (KS‐AR‐KO) using Cre/loxP techniques and the Pax8‐Cre transgene; control (C) mice were Pax8‐Cre‐negative littermates. Mice were acid‐loaded by adding HCl to powdered chow for 7 days; control mice had equal amounts of deionized water added. Urine ammonia under basal conditions was ~2 fold greater in male KO (M‐KO) than in male control (M‐C) mice. After acid‐loading, ammonia excretion was significantly greater in M‐KO than in M‐C mice on day 7 (M‐KO, 852±203; M‐C, 588±93 µmol/day; P < 0.05; N= 6 in each group). Basal urine pH was significantly lower in M‐KO than in M‐C mice, but the response to acid‐loading significantly between genotypes. There was no significant difference in serum Na+, K+, and HCO3‐ after acid loading. Expression of phosphoenolpyruvate carboxykinase (PEPCK), a major proximal tubule (PT) ammonia generating protein, was greater under control conditions in M‐KO, but the relative increase with acid‐loading was greater in M‐C mice. Expression of glutamine synthetase, which recycles ammonia, significantly decreased in response to an acid load, with no significant genotype difference. NBCe1‐A expression was significantly less under control conditions in M‐KO, but the relative increase with acid‐loading was greater in M‐KO mice. Expression of NHE3, believed to be the primary mechanism of PT ammonia secretion, increased in response to acid‐loading in M‐KO mice but not in M‐C mice. After acid‐loading, cortical NKCC2, which mediates thick ascending limb ammonia reabsorption, was significantly greater in M‐KO than M‐C mice. The collecting duct secretes the majority of urinary ammonia, and the Rhesus glycoproteins, Rhbg and Rhcg, are the primary collecting duct ammonia transporting proteins. Expression of Rhbg increased in response to an acid load with no significant genotype difference. Renal structural analysis showed AR deletion in male mice significantly decreased kidney size (M‐C, 275±33 mg; M‐KO, 225±32; P<0.05; N= 6 in each group) after acid loading. In summary, KS‐AR‐KO increases the urinary ammonia response to acid‐loading, through coordinated effects on PEPCK, GS, NBCe1‐A, NHE3 and NKCC2. Thus, in male mice renal AR activation inhibits the response to acid‐loading of multiple components of ammonia metabolism and transport, which leads to less ability to increase ammonia excretion.

The funny current: Even funnier than 40 years ago. Uncanonical expression and roles of HCN/f channels all over the body

Discovered some 40 years ago, the If current has since been known as the “pacemaker” current due to its role in the initiation and modulation of the heartbeat and of neuronal excitability. But this is not all, the funny current keeps entertaining the researchers; indeed, several data discovering novel and uncanonical roles of f/HCN channel are quickly accumulating. In the present review, we provide an overview of the expression and cellular functions of HCN/f channels in a variety of systems/organs, and particularly in sour taste transduction, hormones secretion, activation of astrocytes and microglia, inhibition of osteoclastogenesis, renal ammonium excretion, and peristalsis in the gastrointestinal and urine systems. We also analyzed the role of HCN channels in sustaining cellular respiration in mitochondria and their participation to mitophagy under specific conditions. The relevance of HCN currents in undifferentiated cells, and specifically in the control of stem cell cycle and in bioelectrical signals driving left/right asymmetry during zygote development, is also considered. Finally, we present novel data concerning the expression of HCN mRNA in human leukocytes.We can thus conclude that the emerging evidence presented in this review clearly points to an increasing interest and importance of the “funny” current that goes beyond its role in cardiac sinoatrial and neuronal excitability regulation.

Differences in acidosis-stimulated renal ammonia metabolism in the male and female kidney.

Renal ammonia excretion is a critical component of acid-base homeostasis, and changes in ammonia excretion are the predominant component of increased net acid excretion in response to metabolic acidosis. We recently reported substantial sex-dependent differences in basal ammonia metabolism that correlate with sex-dependent differences in renal structure and expression of key proteins involved in ammonia metabolism. The purpose of the present study was to investigate the effect of sex on the renal ammonia response to an exogenous acid load. We studied 4-mo-old C57BL/6 mice. Ammonia excretion, which was less in male mice under basal conditions, increased in response to acid loading to a greater extent in male mice, such that maximal ammonia excretion did not differ between the sexes. Fundamental structural sex differences in the nonacid-loaded kidney persisted after acid loading, with less cortical proximal tubule volume density in the female kidney than in the male kidney, whereas collecting duct volume density was greater in the female kidney. To further investigate sex-dependent differences in the response to acid loading, we examined the expression of proteins involved in ammonia metabolism. The change in expression of phosphoenolpyruvate carboxykinase and Rh family B glycoprotein with acid loading was greater in male mice than in female mice, whereas Na+-K+-2Cl- cotransporter and inner stripe of the outer medulla intercalated cell Rh family C glycoprotein expression were significantly greater in female mice than in male mice. There was no significant sex difference in glutamine synthetase, Na+/H+ exchanger isoform 3, or electrogenic Na+-bicarbonate cotransporter 1 variant A protein expression in response to acid loading. We conclude that substantial sex-dependent differences in the renal ammonia response to acid loading enable a similar maximum ammonia excretion response.

SAT-269 URINE ESTIMATED AMMONIUM-TO-CREATININE RATIO IN PATIENTS WITH CHRONIC KIDNEY DISEASE.

Renal ammonium (NH4+) excretion plays a critical role in the elimination of acid. Recent studies have reported that the impairment in urinary NH4+ excretion is an important determinant of the development of metabolic acidosis and is an independent factor for predicting loss of renal function. However, urine ammonium measurements are not widely available in routine diagnostic laboratories, and its clinical significance is still unknown. We hypothesized that urine estimated ammonium-to-creatinine ratio (u-eNH4+/u-Cr), as an indicator of urinary NH4+excretion, would be surrogates for early metabolic acidosis in patients with CKD. We measured u-eNH4+/u-Cr in outpatients without overt metabolic acidosis (5.0< [urinary pH] <7.0, [HCO3-] >20 mEq/L, no medication of bicarbonate). The estimated ammonium concentration (u-eNH4+) from the osmolar gap was calculated as u-eNH4+=0.5x(osmolality-2[Na++K+]-[urea]/2.8-[glucose]/18). A total of 168 outpatients were identified (Mean age; 63.1 ± 18.0 years old, Male; 59.5%, CKD; 54.8%, Hypertensives; 74.4%, Diabetes; 35.1%). Although u-eNH4+/u-Cr showed a positive correlation with eGFR (P = 0.0151) (Fig.1), no associations were observed between eNH4+/u-Cr with age, BMI, uPCR, pH, HCO3-, BE, or Anion gap. Interestingly, eNH4+/u-Cr was significantly lower in CKD stage 3-5 than in non-CKD groups (Fig 2), even though there were no overt metabolic acidosis.Fig. 2View Large Image Figure ViewerDownload Hi-res image Download (PPT) u-eNH4+/u-Cr is easily measured in clinical practice and would be more tightly linked with ammonium excretion than serum bicarbonate and other acid-base indicators.

Sex Differences in Acidosis‐Stimulated Renal Ammonia Metabolism

Renal ammonia excretion is a critical component in maintaining acid‐base homoeostasis and changes in ammonia excretion are the predominant component of increased net acid excretion in response to metabolic acidosis. We reported recently substantial sex‐dependent differences in basal ammonia metabolism that correlate with sex‐dependent structural differences and differences in the expression of key proteins involved in ammonia metabolism. This study's objective was to determine the effect of sex on the renal ammonia response to an exogenous acid load. We compared 4‐month old C57BL/6 male (M) and female (F) mice. Mice were acid‐loaded by adding HCl to powdered chow for 7 days, and control mice had equal amounts of deionized water added. Basal ammonia excretion, factored for body weight, was ~2‐fold greater in female than male mice, but the response to acid‐loading did not differ significantly. There was no significant sex difference in serum Na+, K+, and HCO3− after an acid load. The basal fundamental structural differences between the female and male kidney were maintained after an acid load. After acid‐loading, in the female kidney, proximal tubules account for a lower percentage of the cortical parenchyma than in the male kidney (F, 43±2; M, 61±3%; P &lt; 0.001), whereas the collecting ducts account for a greater percentage of the renal parenchyma (F, 16±1; M, 11±1%; P &lt; 0.01) which are similar to basal conditions. Cortical PCT and PST cell height increased in response to acid‐loading, and the increase was greater in M than F kidney. In the outer medulla (OM), PST cell height was similar in M and F, and sex did not alter the increase with acid‐loading. Similarly, intercalated cell (IC) size in the collecting duct in the OM was greater in F than in M kidney, and increased with acid‐loading in both sexes; sex did not alter the acid‐loading increase. Cortical expression of phosphoenol pyruvate, a major proximal tubule (PT) ammonia generating protein, was greater under control conditions in F, but the relative increase with acid‐loading was greater in M. Expression of glutamine synthetase, which recycles ammonia, significantly decreased in response to an acid load, with no significant sex difference. Expression of total cortical NBCe1, but not NBCe1‐A, increased in response to acid‐loading. Expression of NHE3, believed to be the major mechanism of PT ammonia secretion, increased in response to an acid load in M, but not in F. Cortical NKCC2, which mediates thick ascending limb ammonia reabsorption, was significantly greater in F than M after an acid load. The collecting duct secretes the majority of urinary ammonia and the Rhesus glycoproteins, Rhbg and Rhcg, are the primary collecting duct ammonia transporting proteins. Acid‐loading increased cortical Rhbg expression in M, but not in F in the cortex, whereas in the OM, Rhbg expression increased similarly in both M and F. Rhcg expression in intercalated cells in the inner stripe of the outer medulla was significantly greater in F than in M mice after an acid load. We conclude that there are substantial sex‐dependent differences in the mechanism of renal ammonia response to acid‐loading that enable a similar maximum ammonia excretion response.Support or Funding InformationNIH R01DK045788, R01DK107798 and 5T32DK104721This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Резекция печени изменяет механизмы выведения аммиака почками крыс с хроническим тетрахлорметановым гепатитом

Цель исследования - изучение влияния резекции печени (РП) на аммиакэкскретирующую функцию почек при хроническом тетрахлорметановом гепатите. Методика. Опыты выполнены на 265 беспородных белых крысах (самках) массой 180-220 г. Хронический гепатит воспроизводили подкожным введением 50% раствора тетрахлорметана (CCl) на оливковом масле (0,1 мл/100 г массы тела, через сутки, c двумя двухнедельными перерывами между 6, 7 и 13-14 инъекциями). На 65-е сут. (последние) введения тетрахлорметана, удаляли часть левой доли печени (15-20% массы органа). На 3-и, 7-е и 14-е сут. после РП или лапаротомии («ложнооперированные» животные) в почках, артериальной и венозной крови, моче исследовали содержание аммиака, глутамина и мочевины. Результаты. Прогрессирование эндогенной аммиачной интоксикации после РП на фоне тетрахлорметанового гепатита сопровождается повышенной экскрецией аммиака почками. Однако это не устраняет артериальную гипераммониемию и не предотвращает накопление почками аммиака. Инкреция глютамина из почек в кровоток прекращается. К 14-м сут. послеоперационного периода возрастает потребление глютамина из артериальной крови, что приводит к его накоплению в почках. Стимулируя выведение мочевины из организма с мочой, РП одновременно активирует её образование в почках, с дальнейшим поступлением как в кровоток, так и в мочу. В зависимости от сроков послеоперационного периода это сопровождается изменением скорости реабсорбции мочевины в почках. Заключение. Полученные результаты свидетельствуют, что при РП на фоне тетрахлорметанового гепатита почки не предотвращают прогрессирование эндогенной аммиачной интоксикации, патологическое накопление аммиака и глутамина её клетками, но сохраняют способность принимать участие в регуляции повышенного содержания мочевины в артериальной крови.Mechanical (resection) or toxic (hepatitis) liver damage alone has an ambiguous effect on renal ammonia excretion during development of endogenous ammonia intoxication. The aim. The study investigated the effect of liver resection (LR) on renal ammonia excretion in chronic tetrachlorocarbon (CCl)-induced hepatitis. Methods. Experiments were conducted on 240 mongrel white rats (females) weighing 180-220 g. Chronic hepatitis was induced by subcutaneous injection of 50% solution of carbon tetrachloride (CCl) in olive oil (0.1 ml/100g body weight per day with two two-week breaks between injections 6-7 and 13-14). LR with removal of a part of the left lobe (15-20% of body weight) was performed on the 65th (last) day of CCl injections. The animals were examined on the 3rd, 7th and 14th day after LR or laparotomy (sham operation). Contents of ammonia (AM), glutamine (GN), and urea were measured in the kidney, arterial (AB) and venous ( v.renlis ) blood, and urine. Results. Progression of endogenous ammonia intoxication after LR associated with CCl-induced hepatitis and increased renal excretion of Am involves three mechanisms: 1) excretion of Am that is delivered to kidneys in the free form with AB; 2) stimulation of renal tubule secretion of Am that had formed in kidneys by deamidation of «arterial» Gn; and 3) contrary to rules, partial reabsorption of Am from collecting tubules into the blood. However, this does not eliminate arterial hyperammonemia or prevent accumulation of Am in kidneys. The stimulatory effect of LR in CCl-induced hepatitis on Gn incretion from kidneys to the circulation stops by the 14 day after surgery, and the accompanying increased consumption of Gn from AВ results in Gn accumulation in kidneys. LR stimulates urea excretion with urine and simultaneously activates kidney formation of urea, which further enters the bloodstream and urine. Depending on the postoperative period this is associated with changes in the rate of urea reabsorption in kidneys. Conclusions. In RP associated with CCl-induced hepatitis, kidneys cannot prevent progression of endogenous ammonia intoxication and pathological accumulation of ammonia and glutamine in kidney cells but retain the ability to participate in the regulation of the increased urea level in AB.

Effects of Dietary Arginine, Ornithine, and Zeolite Supplementation on Uremic Toxins in Cats.

To test if arginine and ornithine, both components of the Krebs-Henseleit cycle, or zeolite, a potential ammonium absorber, can modulate the excretion of harmful bacterial metabolites, intestinal microbial protein fermentation was stimulated by feeding a high-protein (60.3%) diet as a single daily meal to 10 adult cats. The diet was supplemented without or with arginine (+50, 75, 100% compared to arginine in the basal diet), ornithine (+100, 150, 200% compared to arginine in the basal diet), or zeolite (0.125, 0.25, 0.375 g/kg body weight/day). The cats received each diet for 11 days. Urine, feces, and blood were collected during the last 4 days. Arginine and ornithine enhanced the postprandial increase of blood urea, but renal urea excretion was not increased. Zeolite decreased renal ammonium excretion and fecal biogenic amines. The data indicate an increased detoxification rate of ammonia by arginine and ornithine supplementation. However, as urea was not increasingly excreted, detrimental effects on renal function cannot be excluded. Zeolite had beneficial effects on the intestinal nitrogen metabolism, which should be further evaluated in diseased cats. Clinical studies should investigate whether dietary arginine and ornithine might improve hepatic ammonia detoxification or could be detrimental for renal function.

Urine Ammonium Predicts Clinical Outcomes in Hypertensive Kidney Disease.

Metabolic acidosis is associated with poor outcomes in CKD. Because impaired renal ammonium excretion is important in the pathogenesis of acidosis, urine ammonium excretion might be a better and perhaps earlier acid-base indicator of risk than serum bicarbonate, particularly in patients without acidosis. We evaluated the association between baseline ammonium excretion and clinical outcomes in African American Study of Kidney Disease and Hypertension participants (n=1044). Median daily ammonium excretion was 19.5 (95% confidence interval [95% CI], 6.5 to 43.2) mEq. In Cox regression models (adjusted for demographics, measured GFR, proteinuria, body mass index, net endogenous acid production, and serum potassium and bicarbonate), hazard ratios of the composite outcome of death or dialysis were 1.46 (95% CI, 1.13 to 1.87) in the low tertile and 1.14 (95% CI, 0.89 to 1.46) in the middle tertile of daily ammonium excretion compared with the high tertile. Among participants without acidosis at baseline, the adjusted hazard ratio for those with ammonium excretion <20 mEq/d was 1.36 (95% CI, 1.09 to 1.71) compared with those with ammonium excretion ≥20 mEq/d. Additionally, compared with participants in the high ammonium tertile, those in the low ammonium tertile had higher adjusted odds of incident acidosis at 1 year (adjusted odds ratio, 2.56; 95% CI, 1.04 to 6.27). In conclusion, low ammonium excretion is associated with death and renal failure in hypertensive kidney disease, even among those without acidosis. Low ammonium excretion could identify patients with CKD and normal bicarbonate levels who might benefit from alkali before acidosis develops.

Ammonia Transporters and Their Role in Acid-Base Balance.

Acid-base homeostasis is critical to maintenance of normal health. Renal ammonia excretion is the quantitatively predominant component of renal net acid excretion, both under basal conditions and in response to acid-base disturbances. Although titratable acid excretion also contributes to renal net acid excretion, the quantitative contribution of titratable acid excretion is less than that of ammonia under basal conditions and is only a minor component of the adaptive response to acid-base disturbances. In contrast to other urinary solutes, ammonia is produced in the kidney and then is selectively transported either into the urine or the renal vein. The proportion of ammonia that the kidney produces that is excreted in the urine varies dramatically in response to physiological stimuli, and only urinary ammonia excretion contributes to acid-base homeostasis. As a result, selective and regulated renal ammonia transport by renal epithelial cells is central to acid-base homeostasis. Both molecular forms of ammonia, NH3 and NH4+, are transported by specific proteins, and regulation of these transport processes determines the eventual fate of the ammonia produced. In this review, we discuss these issues, and then discuss in detail the specific proteins involved in renal epithelial cell ammonia transport.

Loss of function mutation of the Slc38a3 glutamine transporter reveals its critical role for amino acid metabolism in the liver, brain, and kidney.

Glutamine, the most abundant amino acid in mammals, is critical for cell and organ functions. Its metabolism depends on the ability of cells to take up or release glutamine by transporters located in the plasma membrane. Several solute carrier (SLC) families transport glutamine, but the SLC38 family has been thought to be mostly responsible for glutamine transport. We demonstrate that despite the large number of glutamine transporters, the loss of Snat3/Slc38a3 glutamine transporter has a major impact on the function of organs expressing it. Snat3 mutant mice were generated by N-ethyl-N-nitrosurea (ENU) mutagenesis and showed stunted growth, altered amino acid levels, hypoglycemia, and died around 20 days after birth. Hepatic concentrations of glutamine, glutamate, leucine, phenylalanine, and tryptophan were highly reduced paralleled by downregulation of the mTOR pathway possibly linking reduced amino acid availability to impaired growth and glucose homeostasis. Snat3-deficient mice had altered urea levels paralleled by dysregulation of the urea cycle, gluconeogenesis, and glutamine synthesis. Mice were ataxic with higher glutamine but reduced glutamate and gamma-aminobutyric acid (GABA) levels in brain consistent with a major role of Snat3 in the glutamine-glutamate cycle. Renal ammonium excretion was lower, and the expression of enzymes and amino acid transporters involved in ammoniagenesis were altered. Thus, SNAT3 is a glutamine transporter required for amino acid homeostasis and determines critical functions in various organs. Despite the large number of glutamine transporters, loss of Snat3 cannot be compensated, suggesting that this transporter is a major route of glutamine transport in the liver, brain, and kidney.

IMPACT OF HYPERBARIC OXYGENATION ON RENAL AMMONIA EXCRETION DURING EXPERIMENTAL LIVER RESECTION

Objective: to study the impact of hyperbaric oxygenation (HBO) on renal ammonia excretion during liver resection (LR). Material and methods. Experiments were carried out on 138 pubescent rats (females). Three HBO sessions were performed at 3 ATA lasting 50 min once daily after LR (15—20% of the liver weight). The kidney, blood (aorta, v. renalis), and urine were examined. The levels of ammonia, glutamine, and urea were measured. Results. By eliminating postoperative arterial hyperammonemia, HBO stimulates urinary ammonia ion excretion. This is achieved through elimination of the inhibitory effect of LR under HBO on ammonia secretion into the renal tubules and through activation of intracellular ammoniagenesis in nephrocytes, including that uncoupled with arterial glutamine deamidation. HBO potentiates simultaneously the activating effect of LR on the glutamine cycle in the kidneys mainly through the hyperproduction of renal glutamine and its influx from them into the bloodstream. HPO enhances the stimulatory effect of LR on urea reabsorption in the kidneys, but, unlike non-oxygenated rats, there was no reduction in urine urea excretion due to increased arterial blood urea concentrations. Termination of the body's exposure to hyperoxia restores urea reabsorption in the kidneys with the persisting stimulatory impact of HBO on the formation of urea by nephrocytes and its incretion from the latter into the bloodstream.

Effect of dietary protein restriction on renal ammonia metabolism.

Dietary protein restriction has multiple benefits in kidney disease. Because protein intake is a major determinant of endogenous acid production, it is important that net acid excretion change in parallel during protein restriction. Ammonia is the primary component of net acid excretion, and inappropriate ammonia excretion can lead to negative nitrogen balance. Accordingly, we examined ammonia excretion in response to protein restriction and then we determined the molecular mechanism of the changes observed. Wild-type C57Bl/6 mice fed a 20% protein diet and then changed to 6% protein developed an 85% reduction in ammonia excretion within 2 days, which persisted during a 10-day study. The expression of multiple proteins involved in renal ammonia metabolism was altered, including the ammonia-generating enzymes phosphate-dependent glutaminase (PDG) and phosphoenolpyruvate carboxykinase (PEPCK) and the ammonia-metabolizing enzyme glutamine synthetase. Rhbg, an ammonia transporter, increased in expression in the inner stripe of outer medullary collecting duct intercalated cell (OMCDis-IC). However, collecting duct-specific Rhbg deletion did not alter the response to protein restriction. Rhcg deletion did not alter ammonia excretion in response to dietary protein restriction. These results indicate 1) dietary protein restriction decreases renal ammonia excretion through coordinated regulation of multiple components of ammonia metabolism; 2) increased Rhbg expression in the OMCDis-IC may indicate a biological role in addition to ammonia transport; and 3) Rhcg expression is not necessary to decrease ammonia excretion during dietary protein restriction.

Physiological and molecular responses of the goldfish (Carassius auratus) kidney to metabolic acidosis, and potential mechanisms of renal ammonia transport.

Relative to the gills, the mechanisms by which the kidney contributes to ammonia and acid-base homeostasis in fish are poorly understood. Goldfish were exposed to a low pH environment (pH4.0, 48 h), which induced a characteristic metabolic acidosis and an increase in total plasma [ammonia] but reduced plasma ammonia partial pressure (PNH3). In the kidney tissue, total ammonia, lactate and intracellular pH remained unchanged. The urinary excretion rate of net base under control conditions changed to net acid excretion under low pH, with contributions from both the NH4 (+) (∼30%) and titratable acidity minus bicarbonate (∼70%; TA-HCO3 (-)) components. Inorganic phosphate (Pi), urea and Na(+) excretion rates were also elevated while Cl(-) excretion rates were unchanged. Renal alanine aminotransferase activity increased under acidosis. The increase in renal ammonia excretion was due to significant increases in both the glomerular filtration and the tubular secretion rates of ammonia, with the latter accounting for ∼75% of the increase. There was also a 3.5-fold increase in the mRNA expression of renal Rhcg-b (Rhcg1) mRNA. There was no relationship between ammonia secretion and Na(+) reabsorption. These data indicate that increased renal ammonia secretion during acidosis is probably mediated through Rhesus (Rh) glycoproteins and occurs independently of Na(+) transport, in contrast to branchial and epidermal models of Na(+)-dependent ammonia transport in freshwater fish. Rather, we propose a model of parallel H(+)/NH3 transport as the primary mechanism of renal tubular ammonia secretion that is dependent on renal amino acid catabolism.

Impaired expression of key molecules of ammoniagenesis underlies renal acidosis in a rat model of chronic kidney disease.

Advanced chronic kidney disease (CKD) is associated with the development of renal metabolic acidosis. Metabolic acidosis per se may represent a trigger for progression of CKD. Renal acidosis of CKD is characterized by low urinary ammonium excretion with preserved urinary acidification indicating a defect in renal ammoniagenesis, ammonia excretion or both. The underlying molecular mechanisms, however, have not been addressed to date. We examined the Han:SPRD rat model and used a combination of metabolic studies, mRNA and protein analysis of renal molecules involved in acid-base handling. We demonstrate that rats with reduced kidney function as evident from lower creatinine clearance, lower haematocrit, higher plasma blood urea nitrogen, creatinine, phosphate and potassium had metabolic acidosis that could be aggravated by HCl acid loading. Urinary ammonium excretion was highly reduced whereas urinary pH was more acidic in CKD compared with control animals. The abundance of key enzymes and transporters of proximal tubular ammoniagenesis (phosphate-dependent glutaminase, PEPCK and SNAT3) and bicarbonate transport (NBCe1) was reduced in CKD compared with control animals. In the collecting duct, normal expression of the B1 H(+)-ATPase subunit is in agreement with low urinary pH. In contrast, the RhCG ammonia transporter, critical for the final secretion of ammonia into urine was strongly down-regulated in CKD animals. In the Han:SPRD rat model for CKD, key molecules required for renal ammoniagenesis and ammonia excretion are highly down-regulated providing a possible molecular explanation for the development and maintenance of renal acidosis in CKD patients.

Effects of chronic lithium administration on renal acid excretion in humans and rats.

Lithium therapy's most common side effects affecting the kidney are nephrogenic diabetes insipidus (NDI) and chronic kidney disease. Lithium may also induce a distal renal tubular acidosis. This study investigated the effect of chronic lithium exposure on renal acid–base homeostasis, with emphasis on ammonia and citrate excretion. We compared 11 individuals on long‐term lithium therapy with six healthy individuals. Under basal conditions, lithium‐treated individuals excreted significantly more urinary ammonia than did control subjects. Following an acute acid load, urinary ammonia excretion increased approximately twofold above basal rates in both lithium‐treated and control humans. There were no significant differences between lithium‐treated and control subjects in urinary pH or urinary citrate excretion. To elucidate possible mechanisms, rats were randomized to diets containing lithium or regular diet for 6 months. Similar to humans, basal ammonia excretion was significantly higher in lithium‐treated rats; in addition, urinary citrate excretion was also significantly greater. There were no differences in urinary pH. Expression of the critical ammonia transporter, Rhesus C Glycoprotein (Rhcg), was substantially greater in lithium‐treated rats than in control rats. We conclude that chronic lithium exposure increases renal ammonia excretion through mechanisms independent of urinary pH and likely to involve increased collecting duct ammonia secretion via the ammonia transporter, Rhcg.

Urea and Ammonia Metabolism and the Control of Renal Nitrogen Excretion

Renal nitrogen metabolism primarily involves urea and ammonia metabolism, and is essential to normal health. Urea is the largest circulating pool of nitrogen, excluding nitrogen in circulating proteins, and its production changes in parallel to the degradation of dietary and endogenous proteins. In addition to serving as a way to excrete nitrogen, urea transport, mediated through specific urea transport proteins, mediates a central role in the urine concentrating mechanism. Renal ammonia excretion, although often considered only in the context of acid-base homeostasis, accounts for approximately 10% of total renal nitrogen excretion under basal conditions, but can increase substantially in a variety of clinical conditions. Because renal ammonia metabolism requires intrarenal ammoniagenesis from glutamine, changes in factors regulating renal ammonia metabolism can have important effects on glutamine in addition to nitrogen balance. This review covers aspects of protein metabolism and the control of the two major molecules involved in renal nitrogen excretion: urea and ammonia. Both urea and ammonia transport can be altered by glucocorticoids and hypokalemia, two conditions that also affect protein metabolism. Clinical conditions associated with altered urine concentrating ability or water homeostasis can result in changes in urea excretion and urea transporters. Clinical conditions associated with altered ammonia excretion can have important effects on nitrogen balance.

Rh versus pH: the role of Rhesus glycoproteins in renal ammonia excretion during metabolic acidosis in a freshwater teleost fish.

Increased renal ammonia excretion in response to metabolic acidosis is thought to be a conserved response in vertebrates. We tested the hypothesis that Rhesus (Rh) glycoproteins in the kidney of the freshwater common carp, Cyprinus carpio, play a crucial role in regulating renal ammonia excretion during chronic metabolic acidosis. Exposure to water pH 4.0 (72 h) resulted in a classic metabolic acidosis with reduced plasma arterial pH and [HCO3(-)], no change in PCO2 and large changes in renal function. Urine [NH4(+)] as well as [titratable acidity-HCO3(-)] rose significantly over the acid exposure, but the profound reduction (fivefold) in urine flow rates eliminated the expected elevations in renal ammonia excretion. Low urine flow rates may be a primary strategy to conserve ions, as urinary excretion rates of Na(+), Cl(-) and Ca(2+) were significantly lower during the acid exposure relative to the control period. Interestingly, renal Rhcg1 mRNA and protein levels were elevated in acid-exposed relative to control groups, along with mRNA levels of several ion transporters, including the Na(+)/H(+) exchanger, H(+)-ATPase and Na(+)/K(+)-ATPase. Immunofluorescence microscopy showed a strong apical Rhcg1 signal in distal tubules. Taken together, these data show that renal Rh glycoproteins and associated ion transporters are responsive to metabolic acidosis, but conservation of ions through reduced urine flow rates takes primacy over renal acid-base regulation in the freshwater C. carpio. We propose that an 'acid/base-ion balance' compromise explains the variable renal responses to metabolic acidosis in freshwater teleosts.

Dysregulation of the Glutamine Transporter Slc38a3 (SNAT3) and Ammoniagenic Enzymes in Obese, Glucose-Intolerant Mice

Background/Aims: Uric acid nephrolithiasis is prevalent among patients with type 2 diabetes and metabolic syndrome; it is correlated with an acidic urine and lower urinary ammonium excretion and is likely associated with insulin resistance. Insulin stimulates ammoniagenesis in renal cell lines via increased phosphate-dependent glutaminase (PDG) activity and glutamine metabolism. Ammonium excretion into the proximal tubule is mediated at least in part by the Na⁺/H⁺-exchanger NHE3 and in the collecting duct involving the Rhesus protein RhCG. Here we tested, whether obesity and insulin resistance in a diet-induced mouse model could contribute to deranged ammonium excretion. Methods: Obesity was induced by diet in mice and the impact on key molecules of proximal tubular ammoniagenesis and urinary acid excretion tested. Results: Diet-induced obesity was confirmed by pathological intraperitoneal glucose tolerance tests (IPGTT). Three groups of mice were compared: control mice; obese, glucose-intolerant with abnormal IPGTT (O-GI); or moderate weight with normal IPGTT (Non-Responders, NR). Basal urinary ammonium excretion did not differ among groups. However, acid loading increased urinary ammonium excretion in all groups, but to a lesser extent in the O-GI group. SNAT3 mRNA expression was enhanced in both obese groups. PDG expression was elevated only in acid-loaded O-GI mice, whereas PEPCK was enhanced in both O-GI and NR groups given NH₄CI. NHE activity in the brush border membrane of the proximal tubule was strongly reduced in the O-GI group whereas RhCG expression was similar. Conclusion: In sum, obesity and glucose intolerance impairs renal ammonium excretion in response to NH₄CI feeding most likely through reduced NHE activity. The stimulation of SNAT3 and ammoniagenic enzyme expression may be compensatory but futile.