Proton-facilitated ammonia excretion by ionocytes of medaka (Oryzias latipes) acclimated to seawater

2013 ◽  
Vol 305 (3) ◽  
pp. R242-R251 ◽  
Author(s):  
Sian-Tai Liu ◽  
Lin Tsung ◽  
Jiun-Lin Horng ◽  
Li-Yih Lin
Keyword(s):  
Boundary Layer ◽  
Ammonia Excretion ◽  
Oryzias Latipes ◽  
Yolk Sac ◽  
Pcr Analysis ◽  
Selective Electrode ◽  
High Ammonia ◽  
Rh Glycoproteins ◽  
Electrode Technique

The proton-facilitated ammonia excretion is critical for a fish's ability to excrete ammonia in freshwater. However, it remains unclear whether that mechanism is also critical for ammonia excretion in seawater (SW). Using a scanning ion-selective electrode technique (SIET) to measure H+ gradients, an acidic boundary layer was detected at the yolk-sac surface of SW-acclimated medaka ( Oryzias latipes) larvae. The H+ gradient detected at the surface of ionocytes was higher than that of keratinocytes in the yolk sac. Treatment with Tricine buffer or EIPA (a NHE inhibitor) reduced the H+ gradient and ammonia excretion of larvae. In situ hybridization and immunochemistry showed that slc9a2 (NHE2) and slc9a3 (NHE3) were expressed in the same SW-type ionocytes. A real-time PCR analysis showed that transfer to SW downregulated branchial mRNA expressions of slc9a3 and Rhesus glycoproteins ( rhcg1, rhcg2, and rhbg) but upregulated that of slc9a2. However, slc9a3, rhcg1, rhcg2, and rhbg expressions were induced by high ammonia in SW. This study suggests that SW-type ionocytes play a role in acid and ammonia excretion and that the Na+/H+ exchanger and Rh glycoproteins are involved in the proton-facilitated ammonia excretion mechanism.

Functional plasticity of mitochondrion-rich cells in the skin of euryhaline medaka larvae (Oryzias latipes) subjected to salinity changes

2011 ◽  
Vol 300 (4) ◽  
pp. R858-R868 ◽  
Author(s):  
Wan-Ping Shen ◽  
Jiun-Lin Horng ◽  
Li-Yih Lin
Keyword(s):  
Oryzias Latipes ◽  
Yolk Sac ◽  
Accessory Cell ◽  
Functional Plasticity ◽  
Selective Electrode ◽  
Noninvasive Technique ◽  
Salinity Changes ◽  
External Salinity ◽  
Electrode Technique ◽  
Apical Opening

A noninvasive technique, the scanning ion-selective electrode technique (SIET) was applied to measure Na+ and Cl− transport by the yolk-sac skin and individual mitochondrion-rich cells (MRCs) in intact medaka larvae ( Oryzias latipes ). In seawater (SW)-acclimated larvae, significant outward Na+ and Cl− gradients were measured at the yolk-sac surface, indicating secretions of Na+ and Cl− from the yolk-sac skin. With Na+ pump immunostaining and microscopic observation, two groups of MRCs were identified on the yolk-sac skin of SW-larvae. These were single MRCs (s-MRCs), which do not have an accompanying accessory cell (AC), and multicellular complex MRCs (mc-MRCs), which usually consist of an MRC and an accompanying AC. The percentage of mc-MRC was ∼60% in 30 parts per thousand of SW, and it decreased with the decrease of external salinity. By serial SIET probing over the surface of the MRCs and adjacent keratinocytes (KCs), significant outward fluxes of Na+ and Cl− were detected at the apical opening (membrane) of mc-MRCs, whereas only outward Cl− flux, but not Na+ flux, was detected at s-MRCs. Treatment with 100 μM ouabain or bumetanide effectively blocked the Na+ and Cl− secretion. Following freshwater (FW) to SW transfer, Na+ and Cl− secretions by the yolk-sac skin were fully developed in 5 h and 2 h, respectively. In contrast, both Na+ and Cl− secretions downregulated rapidly after SW to FW transfer. Sequential probing at individual MRCs found that Na+ and Cl− secretions declined dramatically after SW to FW transfer and Na+/Cl− uptake was detected at the same s-MRCs and mc-MRCs after 5 h. This study provides evidence demonstrating that ACs are required for Na+ excretion and MRCs possess a functional plasticity in changing from a Na+/Cl−-secreting cell to a Na+/Cl−-absorbing cell.

Ammonia excretion by the skin of zebrafish (Danio rerio) larvae

2008 ◽  
Vol 295 (6) ◽  
pp. C1625-C1632 ◽  
Author(s):  
Tin-Han Shih ◽  
Jiun-Lin Horng ◽  
Pung-Pung Hwang ◽  
Li-Yih Lin
Keyword(s):  
Direct Evidence ◽  
Ammonia Excretion ◽  
Bafilomycin A1 ◽  
Selective Electrode ◽  
Zebrafish Larvae ◽  
Trapping Mechanism ◽  
Morpholino Oligonucleotides ◽  
Electrode Technique ◽  
External Ph ◽  
Freshwater Teleosts

The mechanism of ammonia excretion in freshwater teleosts is not well understood. In this study, scanning ion-selective electrode technique was applied to measure H+ and NH4+ fluxes in specific cells on the skin of zebrafish larvae. NH4+ extrusion was relatively high in H+ pump-rich cells, which were identified as the H+-secreting ionocyte in zebrafish. Minor NH4+ extrusion was also detected in keratinocytes and other types of ionocytes in larval skin. NH4+ extrusion from the skin was tightly linked to acid secretion. Increases in the external pH and buffer concentration (5 mM MOPS) diminished H+ and NH4+ gradients at the larval surface. Moreover, coupled decreases in NH4+ and H+ extrusion were found in larvae treated with an H+-pump inhibitor (bafilomycin A1) or H+-pump gene ( atp6v1a) knockdown. Knockdown of Rhcg1 with morpholino-oligonucleotides also decreased NH4+ excretion. This study demonstrates ammonia excretion in epithelial cells of larval skin through an acid-trapping mechanism, and it provides direct evidence for the involvement of the H+ pump and an Rh glycoprotein (Rhcg1) in ammonia excretion.

scholarly journals Acid secretion by mitochondrion-rich cells of medaka (Oryzias latipes) acclimated to acidic freshwater

2012 ◽  
Vol 302 (2) ◽  
pp. R283-R291 ◽  
Author(s):  
Chia-Cheng Lin ◽  
Li-Yih Lin ◽  
Hao-Hsuan Hsu ◽  
Violette Thermes ◽  
Patrick Prunet ◽  
...  
Keyword(s):  
Acid Secretion ◽  
Mrna Level ◽  
Critical Role ◽  
Mrna Levels ◽  
Selective Electrode ◽  
Embryonic Skin ◽  
Apical Membranes ◽  
Acid Acclimation ◽  
Electrode Technique

In the present study, medaka embryos were exposed to acidified freshwater (pH 5) to investigate the mechanism of acid secretion by mitochondrion-rich (MR) cells in embryonic skin. With double or triple in situ hybridization/immunocytochemistry, the Na+/H+ exchanger 3 (NHE3) and H+-ATPase were localized in two distinct subtypes of MR cells. NHE3 was expressed in apical membranes of a major proportion of MR cells, whereas H+-ATPase was expressed in basolateral membranes of a much smaller proportion of MR cells. Gill mRNA levels of NHE3 and H+-ATPase and the two subtypes of MR cells in yolk sac skin were increased by acid acclimation; however, the mRNA level of NHE3 was remarkably higher than that of H+-ATPase. A scanning ion-selective electrode technique was used to measure H+, Na+, and NH4+ transport by individual MR cells in larval skin. Results showed that Na+ uptake and NH4+ excretion by MR cells increased after acid acclimation. These findings suggested that the NHE3/Rh glycoprotein-mediated Na+ uptake/NH4+ excretion mechanism plays a critical role in acidic equivalent (H+/NH4+) excretion by MR cells of the freshwater medaka.

Ammonium-dependent sodium uptake in mitochondrion-rich cells of medaka (Oryzias latipes) larvae

2010 ◽  
Vol 298 (2) ◽  
pp. C237-C250 ◽  
Author(s):  
Shu-Chen Wu ◽  
Jiun-Lin Horng ◽  
Sian-Tai Liu ◽  
Pung-Pung Hwang ◽  
Zhi-Hong Wen ◽  
...  
Keyword(s):  
In Situ Hybridization ◽  
Real Time Pcr ◽  
Skin Surface ◽  
Selective Electrode ◽  
Trapping Mechanism ◽  
Water Ph ◽  
Acid Secreting ◽  
Apical Membranes ◽  
Electrode Technique

In this study, a scanning ion-selective electrode technique (SIET) was applied to measure H+, Na+, and NH4+ gradients and apparent fluxes at specific cells on the skin of medaka larvae. Na+ uptake and NH3/NH4+ excretion were detected at most mitochondrion-rich cells (MRCs). H+ probing at MRCs revealed two group of MRCs, i.e., acid-secreting and base-secreting MRCs. Treatment with EIPA (100 μM) blocked 35% of the NH3/NH4+ secretion and 54% of the Na+ uptake, suggesting that the Na+/H+ exchanger (NHE) is involved in Na+ and NH3/NH4+ transport. Low-Na+ water (<0.001 mM) or high-NH4+ (5 mM) acclimation simultaneously increased Na+ uptake and NH3/NH4+ excretion but decreased or even reversed the H+ gradient at the skin and MRCs. The correlation between NH4+ production and H+ consumption at the skin surface suggests that MRCs excrete nonionic NH3 (base) by an acid-trapping mechanism. Raising the external NH4+ significantly blocked NH3/NH4+ excretion and Na+ uptake. In contrast, raising the acidity of the water (pH 7 to pH 6) enhanced NH3/NH4+ excretion and Na+ uptake by MRCs. In situ hybridization and real-time PCR showed that the mRNAs of the Na+/H+ exchanger ( slc9a3) and Rhesus glycoproteins ( Rhcg1 and Rhbg) were colocalized in MRCs of medaka, and their expressions were induced by low-Na+ acclimation. This study suggests a novel Na+/NH4+ exchange pathway in apical membranes of MRCs, in which a coupled NHE and Rh glycoprotein is involved and the Rh glycoprotein may drive the NHE by generating H+ gradients across apical membranes of MRCs.

Rhcg1 and NHE3b are involved in ammonium-dependent sodium uptake by zebrafish larvae acclimated to low-sodium water

2012 ◽  
Vol 302 (1) ◽  
pp. R84-R93 ◽  
Author(s):  
Tin-Han Shih ◽  
Jiun-Lin Horng ◽  
Sian-Tai Liu ◽  
Pung-Pung Hwang ◽  
Li-Yih Lin
Keyword(s):  
Yolk Sac ◽  
Ion Selective Electrode ◽  
Loss Of Function ◽  
Selective Electrode ◽  
Uptake Mechanism ◽  
Sodium Uptake ◽  
Zebrafish Larvae ◽  
Low Sodium ◽  
Morpholino Oligonucleotides ◽  
Electrode Technique

To investigate whether Na+ uptake by zebrafish is dependent on NH4+ excretion, a scanning ion-selective electrode technique was applied to measure Na+ and NH4+ gradients at the yolk-sac surface of zebrafish larvae. Low-Na+ acclimation induced an inward Na+ gradient (uptake), and a combination of low Na+ and high NH4+ induced a larger inward Na+ gradient. When measuring the ionic gradients, raising the external NH4+ level (5 mM) blocked NH4+ excretion and Na+ uptake; in contrast, raising the external Na+ level (10 mM) simultaneously enhanced Na+ uptake and NH4+ excretion. The addition of MOPS buffer (5 mM), which is known to block NH4+ excretion, also suppressed Na+ uptake. These results showed that Na+ uptake and NH4+ excretion by larval skin are associated when ambient Na+ level is low. Knockdown of Rhcg1 translation with morpholino-oligonucleotides decreased both NH4+ excretion and Na+ uptake by the skin and Na+ content of whole larvae. Knockdown of nhe3b translation or inhibitor (5-ethylisopropyl amiloride) treatment also decreased both the NH4+ excretion and Na+ uptake. This study provides loss-of-function evidence for the involvement of Rhcg1 and NHE3b in the ammonium-dependent Na+ uptake mechanism in zebrafish larvae subjected to low-Na+ water.

scholarly journals Collecting duct-specific Rh C glycoprotein deletion alters basal and acidosis-stimulated renal ammonia excretion

2009 ◽  
Vol 296 (6) ◽  
pp. F1364-F1375 ◽  
Author(s):  
Hyun-Wook Lee ◽  
Jill W. Verlander ◽  
Jesse M. Bishop ◽  
Peter Igarashi ◽  
Mary E. Handlogten ◽  
...  
Keyword(s):  
Plasma Membranes ◽  
Collecting Duct ◽  
Ammonia Excretion ◽  
Pcr Analysis ◽  
Lipid Phase ◽  
Urine Ph ◽  
Phase Diffusion ◽  
Ammonia Transport ◽  
Lipid Diffusion ◽  
Rh Glycoproteins

NH3movement across plasma membranes has traditionally been ascribed to passive, lipid-phase diffusion. However, ammonia-specific transporters, Mep/Amt proteins, are present in primitive organisms and mammals express orthologs of Mep/Amt proteins, the Rh glycoproteins. These findings suggest that the mechanisms of NH3movement in mammalian tissues should be reexamined. Rh C glycoprotein (Rhcg) is expressed in the collecting duct, where NH3secretion is necessary for both basal and acidosis-stimulated ammonia transport. To determine whether the collecting duct secretes NH3via Rhcg or via lipid-phase diffusion, we generated mice with collecting duct-specific Rhcg deletion (CD-KO). CD-KO mice had loxP sites flanking exons 5 and 9 of the Rhcg gene (Rhcgfl/fl) and expressed Cre-recombinase under control of the Ksp-cadherin promoter (Ksp-Cre). Control (C) mice were Rhcgfl/flbut Ksp-Cre negative. We confirmed kidney-specific genomic recombination using PCR analysis and collecting duct-specific Rhcg deletion using immunohistochemistry. Under basal conditions, urinary ammonia excretion was less in KO vs. C mice; urine pH was unchanged. After acid-loading for 7 days, CD-KO mice developed more severe metabolic acidosis than did C mice. Urinary ammonia excretion did not increase significantly on the first day of acidosis in CD-KO mice, despite an intact ability to increase urine acidification, whereas it increased significantly in C mice. On subsequent days, urinary ammonia excretion slowly increased in CD-KO mice, but was always significantly less than in C mice. We conclude that collecting duct Rhcg expression contributes to both basal and acidosis-stimulated renal ammonia excretion, indicating that collecting duct ammonia secretion is, at least in part, mediated by Rhcg and not solely by lipid diffusion.

Calcitropic gene expression suggests a role for the intraplacental yolk sac in maternal-fetal calcium exchange

2002 ◽  
Vol 282 (3) ◽  
pp. E721-E732 ◽  
Author(s):  
Christopher S. Kovacs ◽  
Linda L. Chafe ◽  
Mandy L. Woodland ◽  
Kirsten R. McDonald ◽  
Neva J. Fudge ◽  
...  
Keyword(s):  
Time Frame ◽  
Yolk Sac ◽  
Late Gestation ◽  
Calcium Sensing ◽  
Parathyroid Hormone Related Protein ◽  
Active Calcium ◽  
Calcium Exchange ◽  
Calbindin D ◽  
Pthrp Receptor

The expression of calcitropic genes and proteins was localized within murine placenta during late gestation (the time frame of active calcium transfer) with an analysis of several gene-deletion mouse models by immunohistochemistry and in situ hybridization. Parathyroid hormone-related protein (PTHrP), the PTH/PTHrP receptor, calcium receptor, calbindin-D9k, Ca2+-ATPase, and vitamin D receptor were all highly expressed in a localized structure of the murine placenta, the intraplacental yolk sac, compared with trophoblasts. In the PTHrP gene-deleted or Pthrp-null placenta in which placental calcium transfer is decreased, calbindin-D9k expression was downregulated in the intraplacental yolk sac but not in the trophoblasts. These observations indicated that the intraplacental yolk sac contains calcium transfer and calcium-sensing capability and that it is a probable route of maternal-fetal calcium exchange in the mouse.

Localized Corrosion of Binary Mg-Nd Alloys in Chloride-Containing Electrolyte Using a Scanning Vibrating Electrode Technique

CORROSION ◽  
2012 ◽  
Vol 68 (6) ◽  
pp. 489-498 ◽  
Author(s):  
G. Williams ◽  
K. Gusieva ◽  
N. Birbilis
Keyword(s):  
Localized Corrosion ◽  
Radial Expansion ◽  
Alloying Additions ◽  
Corrosion Rates ◽  
Surface Breakdown ◽  
Density Values ◽  
Intact Surface ◽  
Mean Current ◽  
Electrode Technique

The influence of neodymium (Nd) alloying additions in the 0.47 wt% to 3.53 wt% range on the localized corrosion behavior of Mg, when freely corroding in aqueous sodium chloride (NaCl) electrolyte, is investigated using an in situ scanning vibrating electrode technique (SVET). For all samples, the point of surface breakdown is an intense focal anode that expands radially with respect to time, revealing a cathodically activated interior, which is galvanically coupled with the local anode at the perimeter. However, for Nd compositions of ≤0.74%, radial expansion ceases within ca. 2 h of initiation, whereupon dark filiform-like corrosion features are observed, which traverse over the exposed Mg surface. For Nd additions of ≥1.25%, the radial expansion continues with time up to a point where the entire intact surface becomes consumed. The intensity of the local anode ring of circular corroded regions is seen to increase as more cathodically activated corroded surface becomes exposed. Mean current density values measured within these corroded areas increase progressively with Nd content, leading to a progressive rise in localized corrosion rates. The cathodic activation of corroded regions is proposed to derive from an enrichment of noble, Nd-rich intermetallic grains caused as the alpha-Mg phase becomes attacked at local anode sites.

scholarly journals Airborne determination of the temporo-spatial distribution of benzene, toluene, nitrogen oxides and ozone in the boundary layer across Greater London, UK

2015 ◽  
Vol 15 (9) ◽  
pp. 5083-5097 ◽  
Author(s):  
M. D. Shaw ◽  
J. D. Lee ◽  
B. Davison ◽  
A. Vaughan ◽  
R. M. Purvis ◽  
...  
Keyword(s):  
Boundary Layer ◽  
Nitrogen Oxides ◽  
Linear Regression Analysis ◽  
Proton Transfer Reaction ◽  
Mixing Ratios ◽  
Spatially Resolved ◽  
Dual Channel ◽  
Benzene Toluene

Abstract. Highly spatially resolved mixing ratios of benzene and toluene, nitrogen oxides (NOx) and ozone (O3) were measured in the atmospheric boundary layer above Greater London during the period 24 June to 9 July 2013 using a Dornier 228 aircraft. Toluene and benzene were determined in situ using a proton transfer reaction mass spectrometer (PTR-MS), NOx by dual-channel NOx chemiluminescence and O3 mixing ratios by UV absorption. Average mixing ratios observed over inner London at 360 ± 10 m a.g.l. were 0.20 ± 0.05, 0.28 ± 0.07, 13.2 ± 8.6, 21.0 ± 7.3 and 34.3 ± 15.2 ppbv for benzene, toluene, NO, NO2 and NOx respectively. Linear regression analysis between NO2, benzene and toluene mixing ratios yields a strong covariance, indicating that these compounds predominantly share the same or co-located sources within the city. Average mixing ratios measured at 360 ± 10 m a.g.l. over outer London were always lower than over inner London. Where traffic densities were highest, the toluene / benzene (T / B) concentration ratios were highest (average of 1.8 ± 0.5 ppbv ppbv-1), indicative of strong local sources. Daytime maxima in NOx, benzene and toluene mixing ratios were observed in the morning (~ 40 ppbv NOx, ~ 350 pptv toluene and ~ 200 pptv benzene) and in the mid-afternoon for ozone (~ 40 ppbv O3), all at 360 ± 10 m a.g.l.

The Evolution of Asymmetries in the Tropical Cyclone Boundary Layer Wind Field during Landfall

2021 ◽  
Keyword(s):  
Boundary Layer ◽  
Tropical Cyclone ◽  
Wind Field ◽  
Surface Wind ◽  
Frictional Stress ◽  
Homogeneous Surface ◽  
Sudden Decrease ◽  
The Right ◽  
Tropical Cyclone Boundary Layer

Abstract The evolution of the tropical cyclone boundary layer (TCBL) wind field before landfall is examined in this study. As noted in previous studies, a typical TCBL wind structure over the ocean features a supergradient boundary layer jet to the left of motion and Earth-relative maximum winds to the right. However, the detailed response of the wind field to frictional convergence at the coastline is less well known. Here, idealized numerical simulations reveal an increase in the offshore radial and vertical velocities beginning once the TC is roughly 200 km offshore. This increase in the radial velocity is attributed to the sudden decrease in frictional stress once the highly agradient flow crosses the offshore coastline. Enhanced advection of angular momentum by the secondary circulation forces a strengthening of the supergradient jet near the top of the TCBL. Sensitivity experiments reveal that the coastal roughness discontinuity dominates the friction asymmetry due to motion. Additionally, increasing the inland roughness through increasing the aerodynamic roughness length enhances the observed asymmetries. Lastly, a brief analysis of in-situ surface wind data collected during the landfall of three Gulf of Mexico hurricanes is provided and compared to the idealized simulations. Despite the limited in-situ data, the observations generally support the simulations. The results here imply that assumptions about the TCBL wind field based on observations from over horizontally-homogeneous surface types - which have been well-documented by previous studies - are inappropriate for use near strong frictional heterogeneity.

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