Optogenetic Stimulation of Type I GAD65+ Cells in Taste Buds Activates Gustatory Neurons and Drives Appetitive Licking Behavior in Sodium-Depleted Mice.

Mammalian taste buds are comprised of specialized neuroepithelial cells that act as sensors for molecules that provide nutrition (e.g., carbohydrates, amino acids, and salts) and those that are potentially harmful (e.g., certain plant compounds and strong acids). Type II and III taste bud cells (TBCs) detect molecules described by humans as "sweet," "bitter," "umami," and "sour." TBCs that detect metallic ions, described by humans as "salty," are undefined. Historically, type I glial-like TBCs have been thought to play a supportive role in the taste bud, but little research has been done to explore their role in taste transduction. Some evidence implies that type I cells may detect sodium (Na+) via an amiloride-sensitive mechanism, suggesting they play a role in Na+ taste transduction. We used an optogenetic approach to study type I TBCs by driving the expression of the light-sensitive channelrhodopsin-2 (ChR2) in type I GAD65+ TBCs of male and female mice. Optogenetic stimulation of GAD65+ TBCs increased chorda tympani nerve activity and activated gustatory neurons in the rostral nucleus tractus solitarius. "N neurons," whose NaCl responses were blocked by the amiloride analog benzamil, responded robustly to light stimulation of GAD65+ TBCs on the anterior tongue. Two-bottle preference tests were conducted under Na+-replete and Na+-deplete conditions to assess the behavioral impact of optogenetic stimulation of GAD65+ TBCs. Under Na+-deplete conditions GAD65-ChR2-EYFP mice displayed a robust preference for H2O illuminated with 470 nm light versus nonilluminated H2O, suggesting that type I glial-like TBCs are sufficient for driving a behavior that resembles Na+ appetite.SIGNIFICANCE STATEMENT This is the first investigation on the role of type I GAD65+ taste bud cells (TBCs) in taste-mediated physiology and behavior via optogenetics. It details the first definitive evidence that selective optogenetic stimulation of glial-like GAD65+ TBCs evokes neural activity and modulates behavior. Optogenetic stimulation of GAD65+ TBCs on the anterior tongue had the strongest effect on gustatory neurons that responded best to NaCl stimulation through a benzamil-sensitive mechanism. Na+-depleted mice showed robust preferences to "light taste" (H2O illuminated with 470 nm light vs nonilluminated H2O), suggesting that the activation of GAD65+ cells may generate a salt-taste sensation in the brain. Together, our results shed new light on the role of GAD65+ TBCs in gustatory transduction and taste-mediated behavior.

Dysfunction of the endothelium and constriction of the isolated rat's middle cerebral artery in low sodium environment in the presence of vasopressin.

Hyponatraemia, a water-electrolyte disorder diagnosed in patients with subarachnoid haemorrhage (SAH), increases a risk of persistent vasospasm. In majority of cases, hyponatraemia results from inappropriate secretion of vasopressin (AVP). The effect of AVP-associated hyponatraemia on cerebral vasculature is unknown. The present study aimed to elucidate the role of AVP in the response of the middle cerebral artery (MCA) of the rat to hyponatraemia. Isolated, cannulated, and pressurized rat MCAs were perfused/superfused with physiological (Na+  = 144 mmol/L) buffer or low-sodium (Na+  = 121 mmol/L) buffer containing either AVP or angiotensin II (ANG II). ANG II was used to check if the effect of low plasma sodium concentration combined with AVP on the MCA tone is unique to vasopressin. At physiological Na+ concentration, vasopressin (1.4 × 10-11  mol/L) or angiotensin II (10-9  mol/L) resulted in relaxation of the MCA. Substitution of low-sodium for the normal sodium buffer with the same concentration of AVP, resulted in the constriction of the MCA. This effect was absent after removal of the endothelium, administration of vasopressin V1 receptor antagonist or concomitant inhibition of endothelin-1 receptors and synthesis of thromboxane A2. In contrast, no constriction of the MCA in low-sodium buffer was observed when AVP was replaced with ANG II. Our data suggest that presence of vasopressin and low sodium ion concentration results in the change of endothelium phenotype from pro-vasodilatory to pro-vasoconstrictory. This phenomenon may be an overlooked factor contributing to vasospasm in SAH patients with hyponatraemia caused by inappropriate antidiuretic hormone secretion (SIADH).

Acute body sodium depletion induces skin sodium mobilization in female Wistar rats.

NEW FINDINGS: What is the central question of this study? Can Na+ depletion mobilize Na+ from the skin reservoir in ovariectomized rats? Does oestrogen replacement change the amount and the dynamics of skin Na+ storage? Is the reduced salt appetite after Na+ depletion in ovariectomized rats with oestrogen replacement related to changes in the skin Na+ ? What is the main finding and its importance? This work demonstrated that acute body Na+ depletion induced by frusemide mobilized the osmotically inactive skin Na+ reservoir to become osmotically active. Oestrogen treatment decreased the induced Na+ intake in ovariectomized rats but did not modulate the inactive Na+ reservoir in control conditions or its mobilization induced by Na+ depletion. ABSTRACT: Oestradiol, which is an important hormone for water and electrolyte balance, also has a role in the inhibition of induced Na+ appetite. Sodium can be stored in the skin in osmotically active or inactive forms, and this skin Na+ reservoir may be involved in the control of body Na+ levels during physiopathological challenges. In this study, we investigated whether the effect of sodium depletion by frusemide can mobilize Na+ from the skin reservoir and whether oestradiol replacement changes or mobilizes the Na+ reserves in the skin. Ovariectomized Wistar rats were treated with vehicle or oestradiol for 7 days to evaluate the effects of oestrogen on the hydroelectrolyte balance, intake responses and skin Na+ and water content in basal conditions. Furthermore, the effects of oestrogen were evaluated after 24 h frusemide-induced whole-body Na+ depletion. Oestradiol-replaced rats exhibited reduced water intake without any significant changes in salt intake, Na+ excretion or water and Na+ skin content in basal conditions. After sodium depletion, both vehicle- and oestradiol-treated rats exhibited an increase in the osmotically active skin Na+ , which was associated with a decrease of the inactive skin Na+ reservoir. Oestrogen decreased the hypertonic saline intake induced by Na+ depletion, but it was not associated with any significant changes in the skin Na+ reservoir. Thus, sodium depletion is able to change the inactive-active skin Na+ reservoir balance. However, the oestrogenic modulation of sodium appetite after Na+ depletion is probably not related to the action of this hormone in the skin Na+ reservoir balance.

Na restriction activates epithelial Na channels in rat kidney through two mechanisms and decreases distal Na+ delivery.

KEY POINTS: Dietary Na restriction, through the mineralocorticoid aldosterone, acts on epithelial Na channels via both fast (24 h) and slow (5-7 days) mechanisms in the kidney. The fast effect entails increased proteolytic processing and trafficking of channel protein to the apical membrane. It is rapidly reversible by the mineralocorticoid receptor antagonist eplerenone and is largely lost when tubules are studied ex vivo. The slow effect does not require increased processing or surface expression, is refractory to acute eplerenone treatment, and is preserved ex vivo. Both slow and fast effects contribute to Na retention in vivo. Increased Na+ reabsorption in the proximal tubule also promotes Na conservation under conditions of chronic dietary Na restriction, reducing Na+ delivery to the distal nephron. ABSTRACT: Changes in the activity of the epithelial Na channel (ENaC) help to conserve extracellular fluid volume. In rats fed a low-salt diet, proteolytic processing of ENaC increased within 1 day, and was almost maximal after 3 days. The rapid increase in the abundance of cleaved αENaC and γENaC correlated with decreased urinary Na+ excretion and with increased ENaC surface expression. By contrast, ENaC activity, measured ex vivo in isolated cortical collecting ducts, increased modestly after 3 days and required 5 days to reach maximal levels. The mineralocorticoid receptor antagonist eplerenone reversed the increase in cleaved γENaC and induced natriuresis after 1 or 3 days but failed to alter either ENaC currents or Na+ excretion after 7 days of Na restriction. We conclude that Na depletion, through aldosterone, stimulates ENaC via independent fast and slow mechanisms. In vivo, amiloride-induced natriuresis increased after 1 day of Na depletion. By contrast, hydrochlorothiazide (HCTZ)-induced natriuresis decreased gradually over 7 days, consistent with increased ability of ENaC activity to compensate for decreased Na+ reabsorption in the distal convoluted tubule. Administration of amiloride and HCTZ together increased Na+ excretion less in Na-depleted compared to control animals, indicating decreased delivery of Na+ to the distal nephron when dietary Na is restricted. Measurements of creatinine and Li+ clearances indicated that increased Na reabsorption by the proximal tubules is responsible for the decreased delivery. Thus, Na conservation during chronic dietary salt restriction entails enhanced transport by both proximal and distal nephron segments.

Sodium Carbonate is Saltier Than Sodium Chloride to Sodium-Depleted Rats.

In a series of behavioral experiments in the 1960s, G.R. Morrison identified several unique features of the taste of Na2CO3 to rats; namely, it is 1) considerably more intense than NaCl at isomolar concentrations, 2) avoided at 10 times lower concentrations than NaCl to thirsty rats, 3) preferred at 10 times lower concentrations than NaCl in sodium-depleted rats. He also demonstrated its qualitatively similarity to NaCl. In Experiment 1, we confirmed and extended many of Morrison's observations. Rats were injected with furosemide on 3 occasions to stimulate a sodium appetite. After each depletion, rats were given a brief-access taste test in a lickometer presenting, in random order, water and 7 concentrations of salt. One test used NaCl (0.028-0.89 M, quarter log steps), another used Na2CO3, and the third used Na2CO3, but at a tenfold lower concentration range (0.0028-0.089 M). Rats licked NaCl in an inverted-U shaped concentration-response function peaking at 0.158-0.281 M. As Morrison's results predicted, rats licked Na2CO3 in nearly identical fashion, but at a tenfold lower concentration range (peak at 0.0158-0.028 M). In a second experiment, furosemide-treated rats were repeatedly tested with the lower Na2CO3 range but mixed in the epithelial sodium channel blocker amiloride at various concentrations (3-300 µM, half log steps). Amiloride reduced licking for Na2CO3 and shifted the peak response rightward up to about half a log unit. Thus, this "super-saltiness" of Na2CO3 to rats is at least partly amiloride-dependent.

The Perceptual Characteristics of Sodium Chloride to Sodium-Depleted Rats.

Three experiments assessed potential changes in the rat's perception of sodium chloride (NaCl) during a state of sodium appetite. In Experiment 1, sodium-sufficient rats licking a range of NaCl concentrations (0.028-0.89M) in 15s trials showed an inverted U-shaped concentration response function peaking at 0.281M. Depleted rats (furosemide) showed an identical function, merely elevated, suggesting altered qualitative or hedonic perception but no change in perceived intensity. In Experiment 2, sodium-depleted rats were tested with NaCl, sodium gluconate, and potassium chloride (KCl; 0.028-0.89M) similar to Experiment 1. KCl was licked at the same rate as water except for a slight elevation at 0.158; sodium gluconate and NaCl were treated similarly, but rats showed more licking for hypertonic sodium gluconate than hypertonic NaCl. Sodium-depleted rats were also tested with NaCl mixed in amiloride (10-300 µM). Amiloride reduced licking but did not alter the shape of the concentration-response function. Collectively, these results suggest that transduction of sodium by epithelial sodium channels (which are blocked by amiloride and are more dominant in sodium gluconate than NaCl transduction) is crucial for the perception of sodium during physiological sodium depletion. In Experiment 3, sodium-deplete rats were tested with NaCl as in Experiment 1 but after taste aversion conditioning to 0.3M NaCl or sucrose. Rats conditioned to avoid NaCl but not sucrose failed to express a sodium appetite, strongly suggesting that NaCl does not undergo a change in taste quality during sodium appetite-rats show no confusion between sucrose and NaCl in this paradigm.

Towards a geography of omnivory: Omnivores increase carnivory when sodium is limiting.

Towards understanding the geography of omnivory, we tested three hypotheses that predict the proportion of animal tissue consumed: the sodium limitation hypothesis predicts that omnivores increase animal consumption in Na-poor environments because Na bioaccumulates from plants to predators; thus, heterotrophs are Na-rich sources. The nitrogen limitation and habitat productivity hypotheses use the same logic to predict more animal consumption in N-poor and productive environments respectively. Omnivory is a common trophic strategy, but what determines the balance of plant and animal tissue omnivores consume is relatively unexplored. Most of what we know comes from single populations at local scales. Here we quantitatively test these three hypotheses at a large geographic scale and across 20 species of omnivorous ants. We tested each hypothesis using N stable isotopes (δ15 N) to quantify the degree of carnivory in ant populations in 20 forests that span 12° latitude from Georgia to Maine, USA. We used the difference in δ15 N between 20 ant conspecifics in 10 genera between two paired forests (10 pairs of 20 forests) that consisted of a coastal and inland forests on the same latitude to determine if the proportion of animal tissue consumed could be predicted based on Na, N or net primary productivity. Sodium gradients accounted for 18% of the variation in δ15 N, 45% if one outlier ant species was omitted. In contrast, the nitrogen limitation and habitat productivity hypotheses, which predict more animal consumption in N-poor and more productive environments respectively, failed to vary with δ15 N. Our results reveal a geography of omnivory driven in part by access to Na.

Enamel alteration following tooth bleaching and remineralization.

The purpose of this study was to compare the effects of professional tooth whitening agents containing highly concentrated hydrogen peroxide (with and without laser activation), on the enamel surface; and the potential of four different toothpastes to remineralize any alterations. The study was performed on 50 human molars, divided in two groups: treated with Opalescence(®) Boost and Mirawhite(®) Laser Bleaching. Furthermore, each group was divided into five subgroups, a control one and 4 subgroups remineralized with: Mirasensitive(®) hap+, Mirawhite(®) Gelleѐ, GC Tooth Mousse™ and Mirafluor(®) C. The samples were analysed by SEM/3D-SEM-micrographs, SEM/EDX-qualitative analysis and SEM/EDX-semiquantitative analysis. The microphotographs show that both types of bleaching cause alterations: emphasized perikymata, erosions, loss of interprizmatic substance; the laser treatment is more aggressive and loss of integrity of the enamel is determined by shearing off the enamel rods. In all samples undergoing remineralization deposits were observed, those of toothpastes based on calcium phosphate technologies seem to merge with each other and cover almost the entire surface of the enamel. Loss of integrity and minerals were detected only in the line-scans of the sample remineralized with GC Tooth Mousse™. The semiquantitative EDX analysis of individual elements in the surface layer of the enamel indicates that during tooth-bleaching with HP statistically significant loss of Na and Mg occurs, whereas the bleaching in combination with a laser leads to statistically significant loss of Ca and P. The results undoubtedly confirm that teeth whitening procedures lead to enamel alterations. In this context, it must be noted that laser bleaching is more aggressive for dental substances. However, these changes are reversible and can be repaired by application of remineralization toothpastes.

Safety and Efficacy of Low-osmolarity ORS vs. Modified Rehydration Solution for Malnourished Children for Treatment of Children with Severe Acute Malnutrition and Diarrhea: A Randomized Controlled Trial.

World Health Organization-recommended rehydration solution for malnourished children (ReSoMal) for rehydrating severe acute malnourished children is not available in India. In present study, 110 consecutive children aged 6-59 months with severely acute malnourishment and acute diarrhea were randomized to low-osmolarity oral rehydration solution (ORS) (osmolarity: 245, sodium: 75) with added potassium (20 mmol/l) or modified ReSoMal (osmolarity: 300, sodium: 45). In all, 15.4% of modified ReSoMal group developed hyponatremia as compared with 1.9% in low-osmolarity ORS, but none developed severe hyponatremia or hypernatremia. Both groups had equal number of successful rehydration (52 each). Both types of ORS were effective in correcting hypokalemia and dehydration, but rehydration was achieved in shorter duration with modified ReSoMal.

Aldosterone increases oxygen consumption of rectal epithelia of normal, sodium-deprived and sodiumloaded rats.

INTRODUCTION: The colonic epithelium is a classical aldosterone target, but the effect of the hormone on the oxygen consumption rate (QO2 ) of this tissue is unknown. Objectives. We aimed at assessing, in the rectal epithelium of rats fed with diets of different sodium content, the effect of epithelial sodium channel (ENaC) blockade on short-circuit current (ISC ) and QO2 , and the acute effect of aldosterone incubation on ISC and QO2 . METHODS: Adult male rats were fed with normal, low or high-sodium diets for 8 days. Plasma sodium and serum aldosterone were measured. Isolated mucosa preparations from the rectal portion of the colon were mounted in Ussing chambers modified to measure ISC and QO2. RESULTS: Baseline ISC and QO2 were highest in sodium-deprived rats. Both were proportionally reduced by amiloride (0.1 mM) in this group and in the normal sodium group, but not in sodium-loaded rats. In separate experiments, incubation with aldosterone (10 nM) for 7 h increased ISC and QO2 in all groups; increases were larger in the normal and sodium-loaded groups. Amiloride decreased both ISC and QO2 , abolishing the differences between groups. Linear regression of the decrease in QO2 and ISC after amiloride showed the steepest slope for the sodium-deprived group and the flattest one for the sodium-loaded group. CONCLUSIONS: Baseline epithelial QO2 of sodium-deprived and control rats is reduced by ENaC blockade. Aldosterone increased QO2 proportionally to ISC augmentation in all groups, but the coupling between aerobic metabolism and electrogenic transport seems more efficient in sodium-deprived animals.

Sodium homeostasis and bone.

PURPOSE OF REVIEW: Sodium balance is primarily regulated through the renin-angiotensin-aldosterone system. Extracellular fluid (ECF) sodium concentrations ([Na]) reflect the overall body sodium content, but are also influenced by the osmoregulatory system, which is regulated by the posterior pituitary hormone arginine vasopressin (AVP). Consequently, changes in total body sodium content are not always accurately reflected by the ECF [Na]. This review summarizes the growing evidence base suggesting that skeletal bone, which is rich in sodium, may play a key role in overall body sodium homeostasis. RECENT FINDINGS: Hyponatremia, even when relatively mild, leads to increased morbidity and mortality in diverse clinical scenarios. In particular, hyponatremia has been shown to increase gait instability, falls, and fracture risk. It now appears likely that at least part of the fracture risk is because of the adverse effects of hyponatremia on bone density and quality. The mechanisms through which this occurs are not yet completely understood, but prominently involve increased bone osteoclast formation and activity. An additional direct effect of AVP on bone remodeling has also been recently suggested. SUMMARY: Recent evidence expands upon the previously accepted concepts of body sodium homeostasis and suggests that sodium balance can be augmented by inputs from skeletal bone, which acts as a sodium-rich reservoir that can be deployed during times of sodium deficiency. However, this evolutionarily adaptive mechanism to maintain sodium homeostasis during times of environmental sodium deprivation also has adverse consequences by negatively impacting bone quality and increasing fracture risk.

Effect of sodium deficiency on growth of surgical infants: a retrospective observational study.

BACKGROUND/AIM: Sodium is thought to be critical to growth. Infants who have an ileostomy may suffer from growth faltering, as sodium losses from stomas may be excessive. Urinary sodium measurements may indicate which patients could benefit from sodium supplementation; however, there is no consensus on what level of urinary sodium should be the cutoff for intervention. Our aim was to determine whether there is a relationship between urinary sodium and growth in infants undergoing ileostomy, colostomy and cystostomy. METHODS: Following audit approval, a retrospective observational study of patient notes and chemical pathology data was carried out. All infants <1 year of age that had an ileostomy, colostomy, or cystostomy procedure between February 1997 and January 2014 were included. Patients' weights, urinary and serum sodium and potassium levels and clinical variables were recorded until discharge. Weights were converted to Z-scores for analysis. RESULTS: Forty patients were identified whose notes were available for review and who had at least three urinary sodium measurements. During their hospital stay, 11 (28%) maintained weight within normal limits (Z-scores -2 to +2, 15 (38%) were moderately malnourished (-3 to -2) and 14 (35%) severely malnourished (<-3). Thirty patients had at least one urinary sodium <10 mmol/litre, six patients had their lowest recorded urinary sodium between 10 and 30 mmol/litre and only four patients had all their urinary sodium measurements >30 mmol litre. Electrolyte data were not normally distributed so that correlations between electrolytes and growth were tested using the non-parametric Spearman rank correlation coefficient. Urinary sodium levels positively correlated with growth (r = 0.3071, p < 0.0001), as did serum sodium levels (r = 0.2620, p = 0.0059) whereas there was no relationship between urine or serum potassium and growth. CONCLUSIONS: Poor growth is frequent in this group of patients and appears to be linked with sodium levels. Further work is necessary to draw up guidelines for appropriate sodium supplementation.

Adaptive appetites for salted and unsalted food in rats: differential effects of sodium depletion, DOCA, and dehydration.

Most ingested sodium is contained in food. The aim was to investigate whether sodium depletion, dehydration, or DOCA alters intakes of salted and unsalted foods by rats given choices of two foods: salted (0.2-0.5% Na) and unsalted food containing either similar or different other dietary components. Diuretic-induced (furosemide or acetazolamide, two treatments on successive days) sodium depletion always caused pronounced falls in intake of unsalted food within 24 h, continuing at least another 2 days (e.g., 20.9 ± 1.6 pretreatment to 14.8 ± 1.2, 10.6 ± 1.5, and 14.3 ± 1.3 g/day for 3 days of depletion). Intake and preference for salted food increased after 24-72 h (e.g., 6.5 ± 1.2 pretreatment to 7.1 ± 1.1, 16.4 ± 2.3, and 17.0 ± 1.5 g/day at 1, 2, and 3 days of depletion). Valsartan (10 mg/day) blocked the increased intake of salted food but not the reduced intake of unsalted food. DOCA (2 mg/day) caused equivalent increase and decrease in intakes of salted and unsalted food, respectively. Water-deprived rats reduced intake (e.g., 14.2 ± 3.1 to 3.2 ± 2.0 g/day) of and preference for salted food (e.g., 56 ± 13% to 21 ± 11%) after 2 days of dehydration but did not consistently reduce intake of unsalted food. Total food ingested/day fell in both sodium-depleted and dehydrated rats. Rats regulate intakes of different foods to balance sodium needs, osmoregulatory homeostasis, and energy requirements. Reduced appetite for unsalted food may be a homeostatic response to sodium depletion, which together with subsequent generation of appetite for salted food, drives animals to ingest sodium-containing food, thereby restoring sodium balance.

Development of an animal model of nephrocalcinosis via selective dietary sodium and chloride depletion.

BACKGROUND: Nephrocalcinosis (NC) is an important clinical problem seen in critically ill preterm neonates treated with loop diuretics. No reliable animal models are available to study the pathogenesis of NC in preterm infants. The purpose of this study was to develop a reproducible and clinically relevant animal model of NC for these patients and to explore the impact of extracellular fluid (ECF) volume contraction induced by sodium and chloride depletion in this process. METHODS: Three-week-old weanling Sprague-Dawley rats were fed diets deficient in either chloride or sodium or both. A subgroup of rats from each dietary group was injected daily with furosemide (40 mg/kg i.p.). RESULTS: Rats fed a control diet, with or without furosemide, or a chloride-depleted diet alone, did not develop NC. By contrast, 50% of the rats injected with furosemide and fed the chloride-depleted diet developed NC. Moreover, 94% of the rats fed the combined sodium- and chloride-depleted diet developed NC, independently of furosemide use. NC was associated with the development of severe ECF volume contraction; hypochloremic, hypokalemic, metabolic alkalosis; increased phosphaturia; and growth retardation. CONCLUSION: Severe ECF volume contraction induced by chronic sodium and chloride depletion appears to play an important role in the pathogenesis of NC.

Functional analysis of the human concentrative nucleoside transporter-1 variant hCNT1S546P provides insight into the sodium-binding pocket.

SLC28 genes, encoding concentrative nucleoside transporter proteins (CNT), show little genetic variability, although a few single nucleotide polymorphisms (SNPs) have been associated with marked functional disturbances. In particular, human CNT1S546P had been reported to result in negligible thymidine uptake. In this study we have characterized the molecular mechanisms responsible for this apparent loss of function. The hCNT1S546P variant showed an appropriate endoplasmic reticulum export and insertion into the plasma membrane, whereas loss of nucleoside translocation ability affected all tested nucleoside and nucleoside-derived drugs. Site-directed mutagenesis analysis revealed that it is the lack of the serine residue itself responsible for the loss of hCNT1 function. This serine residue is highly conserved, and mutation of the analogous serine in hCNT2 (Ser541) and hCNT3 (Ser568) resulted in total and partial loss of function, respectively. Moreover, hCNT3, the only member that shows a 2Na(+)/1 nucleoside stoichiometry, showed altered Na(+) binding properties associated with a shift in the Hill coefficient, consistent with one Na(+) binding site being affected by the mutation. Two-electrode voltage-clamp studies using the hCNT1S546P mutant revealed the occurrence of Na(+) leak, which was dependent on the concentration of extracellular Na(+) indicating that, although the variant is unable to transport nucleosides, there is an uncoupled sodium transport.

Loss of claudin-15, but not claudin-2, causes Na+ deficiency and glucose malabsorption in mouse small intestine.

BACKGROUND & AIMS: In the small intestine, the paracellular transport of Na(+) is thought to be critical for luminal Na(+)-homeostasis and the transcellular absorption of nutrients by Na(+)-driven transporters. Na(+) is supplied to the intestinal lumen from the submucosa and serum through tight junctions, which form a paracellular barrier between the cells of epithelial sheets. However, the molecular basis for this paracellular transport of Na(+) is not well understood. Here, we examined this mechanism by performing loss-of-function studies of claudin-2 and claudin-15, two tight-junctional membrane proteins that are specifically and age-dependently expressed in the villi and/or crypts of small intestinal epithelia. METHODS: Knockout mice for claudin-2 or claudin-15 were subjected to histologic, cell biologic, electrophysiologic, and physiologic analyses. RESULTS: Examination of the knockout mice revealed that both claudin-2 and claudin-15 play crucial roles in the transepithelial paracellular channel-like permselectivity for extracellular monovalent cations, particularly Na(+), in infants and adults. Especially in Cldn15(-/-) adults, the luminal Na(+) concentration in the small intestine measured directly in vivo was abnormally low, and glucose absorption was impaired, as assessed by the oral glucose tolerance test and estimation of unabsorbed glucose. CONCLUSIONS: We propose that the "Na(+)-leaky" claudin-15 is indispensable in vivo for the paracellular Na(+) permeability, luminal Na(+)-homeostasis, and efficient glucose absorption in the small intestine, but claudin-2 is indispensable for only the first of these functions. Claudin-15 knockout leads to Na(+) deficiency and glucose malabsorption in the mouse adult small intestine.

Sodium shortage as a constraint on the carbon cycle in an inland tropical rainforest.

Sodium (Na) is uncommon in plants but essential to the metabolism of plant consumers, both decomposers and herbivores. One consequence, previously unexplored, is that as Na supplies decrease (e.g., from coastal to inland forests), ecosystem carbon should accumulate as detritus. Here, we show that adding NaCl solution to the leaf litter of an inland Amazon forest enhanced mass loss by 41%, decreased lignin concentrations by 7%, and enhanced decomposition of pure cellulose by up to 50%, compared with stream water alone. These effects emerged after 13-18 days. Termites, a common decomposer, increased 7-fold on +NaCl plots, suggesting an agent for the litter loss. Ants, a common predator, increased 2-fold, suggesting that NaCl effects cascade upward through the food web. Sodium, not chloride, was likely the driver of these patterns for two reasons: two compounds of Na (NaCl and NaPO(4)) resulted in equivalent cellulose loss, and ants in choice experiments underused Cl (as KCl, MgCl(2), and CaCl(2)) relative to NaCl and three other Na compounds (NaNO(3), Na(3)PO(4), and Na(2)SO(4)). We provide experimental evidence that Na shortage slows the carbon cycle. Because 80% of global landmass lies >100 km inland, carbon stocks and consumer activity may frequently be regulated via Na limitation.

Inhibition of K+ secretion in the distal nephron in nephrotic syndrome: possible role of albuminuria.

Nephrotic syndrome features massive proteinuria and retention of sodium which promotes ascite formation. In the puromycin aminonucleoside-induced rat model of nephrotic syndrome, sodium retention originates from the collecting duct where it generates a driving force for potassium secretion. However, there is no evidence for urinary potassium loss or hypokalaemia in the nephrotic syndrome. We therefore investigated the mechanism preventing urinary potassium loss in the nephrotic rats and, for comparison, in hypovolaemic rats, another model displaying increased sodium reabsorption in collecting ducts. We found that sodium retention is not associated with urinary loss of potassium in either nephrotic or hypovolaemic rats, but that different mechanisms account for potassium conservation in the two models. Collecting ducts from hypovolaemic rats displayed high expression of the potassium-secreting channel ROMK but no driving force for potassium secretion owing to low luminal sodium availability. In contrast, collecting ducts from nephrotic rats displayed a high driving force for potassium secretion but no ROMK. Down-regulation of ROMK in nephrotic rats probably stems from phosphorylation of ERK arising from the presence of proteins in the luminal fluid. In addition, nephrotic rats displayed a blunted capacity to excrete potassium when fed a potassium-rich diet, and developed hyperkalaemia. As nephrotic patients were found to display plasma potassium levels in the normal to high range, we would recommend not only a low sodium diet but also a controlled potassium diet for patients with nephrotic syndrome.

Ghrelin reduces hypertonic saline intake in a variety of natriorexigenic conditions.

Ghrelin is a gut peptide that has been studied extensively for its role in food intake and energy balance. More recent studies show that ghrelin reduces water intake in rats and some non-mammalian species. Despite the importance of the regulation of NaCl intake in body fluid homeostasis, the effects of ghrelin on saline intake have not been investigated. Accordingly, we tested the effect of ghrelin on water and 1.8% NaCl intake in two-bottle test conditions with the following five stimuli that increase hypertonic saline intake: central angiotensin II administration; 24 h fluid deprivation; water deprivation followed by partial rehydration; dietary sodium deficiency; and polyethylene glycol administration combined with dietary sodium deficiency. We found that ghrelin attenuated saline intake stimulated by angiotensin II, by water deprivation followed by partial rehydration and by dietary sodium deficiency. We did not detect an effect of ghrelin on saline intake after 24 h fluid deprivation without partial rehydration or after the combination of polyethylene glycol and dietary sodium deficiency. The finding that ghrelin reduced hypertonic saline intake in some, but not all, natriorexigenic conditions mirrors the previously published findings that in one-bottle tests of drinking, ghrelin reduces water intake in only some conditions. The results provide evidence for a new role for ghrelin in the regulation of body fluid homeostasis.

Protocol for sodium depletion and measurement of sodium appetite in mice.

Sodium appetite is a state that motivates animals to consume normally unappetizing concentrations of sodium. Here we describe a protocol to induce sodium appetite in mice by furosemide-induced diuresis and measure sodium intake using volumetric drinking tubes. This protocol induces sodium appetite rapidly and can be used to assess the effect of various treatments on sodium appetite. This protocol does not require electronic equipment and can be implemented easily. For complete details on the use and execution of this protocol, please refer to Park et al. (2020).