scholarly journals Identification of renal transporters involved in sulfate excretion in marine teleost fish

2009 ◽  
Vol 297 (6) ◽  
pp. R1647-R1659 ◽  
Author(s):  
Akira Kato ◽  
Min-Hwang Chang ◽  
Yukihiro Kurita ◽  
Tsutomu Nakada ◽  
Maho Ogoshi ◽  
...  
Keyword(s):  
Xenopus Oocytes ◽  
Proximal Tubules ◽  
Border Region ◽  
Marine Teleost ◽  
Renal Transporters ◽  
Electrode Voltage ◽  
Cloned Cdna ◽  
Cdna Fragments ◽  
Immunohistochemical Analyses ◽  
Marine Teleost Fish

Sulfate (SO42−) is the second most abundant anion in seawater (SW), and excretion of excess SO42− from ingested SW is essential for marine fish to survive. Marine teleosts excrete SO42− via the urine produced in the kidney. The SO42− transporter that secretes and concentrates SO42− in the urine has not previously been identified. Here, we have identified and characterized candidates for the long-sought transporters. Using sequences from the fugu database, we have cloned cDNA fragments of all transporters belonging to the Slc13 and Slc26 families from mefugu ( Takifugu obscurus ). We compared Slc13 and Slc26 mRNA expression in the kidney between freshwater (FW) and SW mefugu. Among 14 clones examined, the expression of a Slc26a6 paralog (mfSlc26a6A) was the most upregulated (30-fold) in the kidney of SW mefugu. Electrophysiological analyses of Xenopus oocytes expressing mfSlc26a6A, mfSlc26a6B, and mouse Slc26a6 (mSlc26a6) demonstrated that all transporters mediate electrogenic Cl−/SO42−, Cl−/oxalate2−, and Cl−/ nHCO3− exchanges and electroneutral Cl−/formate− exchange. Two-electrode voltage-clamp experiments demonstrated that the SO42−-elicited currents of mfSlc26a6A is quite large (∼35 μA at +60 mV) and 50- to 200-fold higher than those of mfSlc26a6B and mSlc26a6. Conversely, the currents elicited by oxalate and HCO3− are almost identical among mfSlc26a6A, mfSlc26a6B, and mSlc26a6. Kinetic analysis revealed that mfSlc26a6A has the highest SO42− affinity as well as capacity. Immunohistochemical analyses demonstrated that mfSlc26a6A localizes to the apical (brush-border) region of the proximal tubules. Together, these findings suggest that mfSlc26a6A is the most likely candidate for the major apical SO42− transporter that mediates SO42− secretion in the kidney of marine teleosts.

scholarly journals Extracellular Hco3− Dependence of Electrogenic Na/Hco3 Cotransporters Cloned from Salamander and Rat Kidney

2000 ◽  
Vol 115 (5) ◽  
pp. 533-546 ◽  
Author(s):  
Irina I. Grichtchenko ◽  
Michael F. Romero ◽  
Walter F. Boron
Keyword(s):  
Membrane Potential ◽  
Voltage Clamp ◽  
Xenopus Oocytes ◽  
Rat Kidney ◽  
Outward Current ◽  
Membrane Vesicles ◽  
Disulfonic Acid ◽  
Ambystoma Tigrinum ◽  
Ph Titration ◽  
Electrode Voltage

We studied the extracellular [HCOabstract 3 −] dependence of two renal clones of the electrogenic Na/HCO3 cotransporter (NBC) heterologously expressed in Xenopus oocytes. We used microelectrodes to measure the change in membrane potential (ΔVm) elicited by the NBC cloned from the kidney of the salamander Ambystoma tigrinum (akNBC) and by the NBC cloned from the kidney of rat (rkNBC). We used a two-electrode voltage clamp to measure the change in current (ΔI) elicited by rkNBC. Briefly exposing an NBC-expressing oocyte to HCOabstract 3 −/CO2 (0.33–99 mM HCOabstract 3−, pHo 7.5) elicited an immediate, DIDS (4,4-diisothiocyanatostilbene-2,2-disulfonic acid)-sensitive and Na+-dependent hyperpolarization (or outward current). In ΔVm experiments, the apparent Km for HCOabstract 3− of akNBC (10.6 mM) and rkNBC (10.8 mM) were similar. However, under voltage-clamp conditions, the apparent Km for HCOabstract 3− of rkNBC was less (6.5 mM). Because it has been reported that SOabstract 3=/HSO abstract 3− stimulates Na/HCO3 cotransport in renal membrane vesicles (a result that supports the existence of a COabstract 3= binding site with which SOabstract 3= interacts), we examined the effect of SOabstract 3=/HSO abstract 3− on rkNBC. In voltage-clamp studies, we found that neither 33 mM SOabstract 4= nor 33 mM SOabstract 3 =/HSOabstract 3− substantially affects the apparent Km for HCO abstract 3−. We also used microelectrodes to monitor intracellular pH (pHi) while exposing rkNBC-expressing oocytes to 3.3 mM HCOabstract 3 −/0.5% CO2. We found that SO abstract 3=/HSOabstract 3 − did not significantly affect the DIDS-sensitive component of the pHi recovery from the initial CO2 -induced acidification. We also monitored the rkNBC current while simultaneously varying [CO2]o, pHo, and [COabstract 3=]o at a fixed [HCOabstract 3−]o of 33 mM. A Michaelis-Menten equation poorly fitted the data expressed as current versus [COabstract 3=]o . However, a pH titration curve nicely fitted the data expressed as current versus pHo. Thus, rkNBC expressed in Xenopus oocytes does not appear to interact with SOabstract 3 =, HSOabstract 3−, or COabstract 3=.

Oil Spill Dispersant Use: Toxicity on Marine Teleost Fish

2018 ◽  
pp. 71-82
Author(s):  
Thomas Milinkovitch ◽  
Stéphane Le Floch ◽  
Hélène Thomas-Guyon
Keyword(s):  
Oil Spill ◽  
Teleost Fish ◽  
Marine Teleost ◽  
Marine Teleost Fish

scholarly journals Ca2+-driven intestinal HCO3− secretion and CaCO3 precipitation in the European flounder in vivo: influences on acid-base regulation and blood gas transport

2010 ◽  
Vol 298 (4) ◽  
pp. R870-R876 ◽  
Author(s):  
Christopher A. Cooper ◽  
Jonathan M. Whittamore ◽  
Rod W. Wilson
Keyword(s):  
Teleost Fish ◽  
Gas Transport ◽  
Driving Forces ◽  
Marine Teleost ◽  
Acid Excretion ◽  
Acid Base ◽  
Marine Teleost Fish ◽  
Net Acid Excretion

Marine teleost fish continuously ingest seawater to prevent dehydration and their intestines absorb fluid by mechanisms linked to three separate driving forces: 1) cotransport of NaCl from the gut fluid; 2) bicarbonate (HCO3−) secretion and Cl− absorption via Cl−/HCO3− exchange fueled by metabolic CO2; and 3) alkaline precipitation of Ca2+ as insoluble CaCO3, which aids H2O absorption). The latter two processes involve high rates of epithelial HCO3− secretion stimulated by intestinal Ca2+ and can drive a major portion of water absorption. At higher salinities and ambient Ca2+ concentrations the osmoregulatory role of intestinal HCO3− secretion is amplified, but this has repercussions for other physiological processes, in particular, respiratory gas transport (as it is fueled by metabolic CO2) and acid-base regulation (as intestinal cells must export H+ into the blood to balance apical HCO3− secretion). The flounder intestine was perfused in vivo with salines containing 10, 40, or 90 mM Ca2+. Increasing the luminal Ca2+ concentration caused a large elevation in intestinal HCO3− production and excretion. Additionally, blood pH decreased (−0.13 pH units) and plasma partial pressure of CO2 (Pco2) levels were elevated (+1.16 mmHg) at the highest Ca perfusate level after 3 days of perfusion. Increasing the perfusate [Ca2+] also produced proportional increases in net acid excretion via the gills. When the net intestinal flux of all ions across the intestine was calculated, there was a greater absorption of anions than cations. This missing cation flux was assumed to be protons, which vary with an almost 1:1 relationship with net acid excretion via the gill. This study illustrates the intimate link between intestinal HCO3− production and osmoregulation with acid-base balance and respiratory gas exchange and the specific controlling role of ingested Ca2+ independent of any other ion or overall osmolality in marine teleost fish.

scholarly journals Gene Expression Levels and Immunolocalization of Organic Ion Transporters in the Human Kidney

2002 ◽  
Vol 13 (4) ◽  
pp. 866-874
Author(s):  
Hideyuki Motohashi ◽  
Yuji Sakurai ◽  
Hideyuki Saito ◽  
Satohiro Masuda ◽  
Yumiko Urakami ◽  
...  
Keyword(s):  
Basolateral Membrane ◽  
Mrna Level ◽  
Human Kidney ◽  
Proximal Tubules ◽  
Mrna Levels ◽  
Ion Transporters ◽  
Ion Transporter ◽  
Transporter Family ◽  
Immunohistochemical Analyses ◽  
Gene Expression Levels

ABSTRACT. Renal excretion of organic anions and cations is mediated by the organic ion transporter family (SLC22A). In this study, the mRNA levels of the organic ion transporters were quantified by real-time PCR in normal parts of renal tissues from seven nephrectomized patients with renal cell carcinoma, and the distributions and localization of human (h)OAT1, hOAT3, and hOCT2 proteins were investigated by immunohistochemical analyses in the human kidney. The expression level of hOAT3 mRNA was the highest among the organic ion transporter family, followed by that of hOAT1 mRNA. The hOCT2 mRNA level was the highest in the human OCT family, and the level of hOCTN2 mRNA was higher than that of hOCTN1. hOCT1 mRNA showed the lowest level of expression in organic ion transporter family. hOAT1, hOAT3, and hOCT2 proteins were detected in crude membranes from the kidney of all patients by Western blot analyses, whereas hOCT1 protein could not be detected. Immunohistochemical analyses showed that both hOAT1 and hOAT3 were localized to the basolateral membrane of the proximal tubules in the cortex, and hOCT2 was localized to the basolateral membrane of the proximal tubules in both the cortex and medullary ray. Immunohistochemical analyses of serial sections indicated that hOAT1, hOAT3, and hOCT2 were coexpressed in a portion of the proximal tubules. These results suggest that hOAT1, hOAT3, and hOCT2 play predominant roles in the transport of organic ions across the basolateral membrane of human proximal tubules.

scholarly journals pH sensitivity of ammonium transport by Rhbg

2010 ◽  
Vol 299 (6) ◽  
pp. C1386-C1397 ◽  
Author(s):  
Nazih L. Nakhoul ◽  
Solange M. Abdulnour-Nakhoul ◽  
Eric Schmidt ◽  
Rienk Doetjes ◽  
Edd Rabon ◽  
...  
Keyword(s):  
Xenopus Oocytes ◽  
Ph Sensitivity ◽  
Ammonium Transport ◽  
Amine Hydrochloride ◽  
Membrane Glycoprotein ◽  
Induced Current ◽  
Inward Current ◽  
Gas Channel ◽  
Methyl Amine ◽  
Electrode Voltage

Rhbg is a membrane glycoprotein that is involved in NH3/NH4+ transport. Several models have been proposed to describe Rhbg, including an electroneutral NH4+/H+ exchanger, a uniporter, an NH4+ channel, or even a gas channel. In this study, we characterized the pH sensitivity of Rhbg expressed in Xenopus oocytes. We used two-electrode voltage clamp and ion-selective microelectrodes to measure NH4+-induced [and methyl ammonium (MA+)] currents and changes in intracellular pH (pHi), respectively. In oocytes expressing Rhbg, 5 mM NH4Cl (NH3/NH4+) at extracellular pH (pHo) of 7.5 induced an inward current, decreased pHi, and depolarized the cell. Raising pHo to 8.2 significantly enhanced the NH4+-induced current and pHi changes, whereas decreasing bath pH to 6.5 inhibited these changes. Lowering pHi (decreased by butyrate) also inhibited the NH4+-induced current and pHi decrease. In oocytes expressing Rhbg, 5 mM methyl amine hydrochloride (MA/MA+), often used as an NH4Cl substitute, induced an inward current, a pHi increase (not a decrease), and depolarization of the cell. Exposing the oocyte to MA/MA+ at alkaline bath pH (8.2) enhanced the MA+-induced current, whereas lowering bath pH to 6.5 inhibited the MA+ current completely. Exposing the oocyte to MA/MA+ at low pHi abolished the MA+-induced current and depolarization; however, pHi still increased. These data indicate that 1) transport of NH4+ and MA/MA+ by Rhbg is pH sensitive; 2) electrogenic NH4+ and MA+ transport are stimulated by alkaline pHo but inhibited by acidic pHi or pHo; and 3) electroneutral transport of MA by Rhbg is likely but is less sensitive to pH changes.

ACID-BASE REGULATION, BRANCHIAL TRANSFERS AND RENAL OUTPUT IN A MARINE TELEOST FISH (THE LONG-HORNED SCULPIN MYOXOCEPHALUS OCTODECIMSPINOSUS) DURING EXPOSURE TO LOW SALINITIES

1994 ◽  
Vol 193 (1) ◽  
pp. 79-95 ◽  
Author(s):  
J Claiborne ◽  
J Walton ◽  
D Compton-Mccullough
Keyword(s):  
Teleost Fish ◽  
Sea Water ◽  
Carbon Dioxide Concentration ◽  
Marine Teleost ◽  
Total Acid ◽  
Low Salinity ◽  
External Salinity ◽  
First Time ◽  
Marine Teleost Fish

A number of studies have implied a linkage between acid­base and ion exchanges in both freshwater and seawater fish, although little is known about the branchial and renal acid­base transfers involved as the animals move between different salinities. To investigate the role of these transfers in a marine teleost fish as it is exposed to a dilute environment, we measured plasma acid­base values and net movements from fish to water of NH4+, HCO3- and H+ in long-horned sculpin (Myoxocephalus octodecimspinosus) placed in 100 %, 20 %, 8 % or 4 % sea water for 24­48 h. Renal excretion of H+ was also monitored in fish exposed to 4 % sea water. Sculpin proved to be somewhat euryhaline for they were able to maintain plasma ion and acid­base transfers in hypo-osmotic (20 %) sea water, but could not tolerate greater dilutions for more than several days. Plasma pH and carbon dioxide concentration (CCO2) increased in the 20 % and 8 % dilution groups, with CCO2 nearly doubling (control, 4.56 mmol l-1; 8 % group, 8.56 mmol l-1) as a result of a combined increase in the partial pressure of plasma CO2 (PCO2) and [HCO3-]. During a 44­46 h exposure, HCO3- transfers increased progressively in the most dilute water, with animals in the 8 % and 4 % groups exhibiting a net H+ loss that was smaller than that of seawater fish (control, 5.1 mmol kg-1; 8 %, 0.9 mmol kg-1; 4 %, -2.9 mmol kg-1). Animals exposed to 4 % sea water for 24 h and then returned to normal sea water had a variable plasma pH, an elevated CCO2 and a net efflux of H+ that effectively stopped (control, 0.10 mmol kg-1 h-1; 4 %, 0.02 mmol kg-1 h-1; seawater recovery, 0.20 mmol kg-1 h-1) during the low-salinity period. Renal acid excretion remained relatively constant throughout the experiment but only made up a significant portion (approximately 40 %) of the total acid transfers during the 4 % dilution period (control rate approximately 3 µmol kg-1 h-1: 3 % of branchial rate). We postulate that the increase in plasma CCO2 during exposure to low salinity may be due to mobilization of base from the intracellular bone compartment. The decrease in external salinity could induce base loss by alteration of gill ion exchanges (Na+/H+, Cl-/HCO3-) and/or changes in branchial HCO3- permeability. For the first time, we have shown that the effects of a dilute environment on acid­base transfers may be an important limitation to the survival of a euryhaline species in brackish or fresh water.

Inhibitory Effect of Glucocorticoids on Human-cloned 5-hydroxytryptamine3AReceptor Expressed in Xenopus  Oocytes

2004 ◽  
Vol 101 (3) ◽  
pp. 660-665 ◽  
Author(s):  
Takahiro Suzuki ◽  
Masahiro Sugimoto ◽  
Hideki Koyama ◽  
Takashi Mashimo ◽  
Ichiro Uchida
Keyword(s):  
Xenopus Oocytes ◽  
Inhibitory Concentration ◽  
Inhibitory Effect ◽  
Combined Effect ◽  
Induced Current ◽  
Direct Effects ◽  
Receptor Antagonists ◽  
Electrode Voltage ◽  
Direct Inhibitory Effect ◽  
Voltage Clamping

Background Methylprednisolone, dexamethasone, and other glucocorticoids have been found effective against nausea and vomiting induced by chemotherapy and surgery. Although the specific 5-hydroxytriptamine3 (5-HT3) receptor antagonists such as ondansetron and ramosetron are used as antiemetics, reports show that the use of 5-HT3 receptor antagonists with some glucocorticoids brings additional effects. Glucocorticoids are reported to be antiemetic. The effect of glucocorticoids on 5-HT3 receptor, however, has not been well characterized. This study was designed to examine whether dexamethasone and methylprednisolone had direct effects on human-cloned 5-HT3A receptor expressed in Xenopus oocytes. Methods Homomeric human-cloned 5-HT3A receptor was expressed in Xenopus oocytes. The authors used the two-electrode voltage-clamping technique to study the effect of methylprednisolone and dexamethasone on 5-HT-induced current. Results Both dexamethasone and methylprednisolone concentration-dependently attenuated 5-HT-induced current. Dexamethasone inhibited 2 microm 5-HT-induced current, which was equivalent to EC30 concentration for 5-HT3A receptor, with an inhibitory concentration 50% of 5.29 +/- 1.02 microm. Methylprednisolone inhibited 2 microm 5-HT-induced current with an inhibitory concentration 50% of 1.07 +/- 0.15 mm. The mode of inhibition with either dexamethasone or methylprednisolone was noncompetitive and voltage-independent. When administered together with the 5-HT3 receptor antagonists, ramosetron or metoclopramide, both glucocorticoids showed an additive effect on 5-HT3 receptor. Conclusion The glucocorticoids had a direct inhibitory effect on 5-HT3 receptors. The combined effect of glucocorticoids and the 5-HT3 receptor antagonists seems additive.

scholarly journals Calcium Current Activated by Depletion of Calcium Stores in Xenopus Oocytes

1997 ◽  
Vol 109 (6) ◽  
pp. 703-715 ◽  
Author(s):  
Yong Yao ◽  
Roger Y. Tsien
Keyword(s):  
Xenopus Oocytes ◽  
Voltage Dependence ◽  
Calcium Stores ◽  
Kinase C ◽  
Electrode Voltage ◽  
Activation Mechanisms ◽  
Electrophysiological Characterization ◽  
Inwardly Rectifying ◽  
A Current

Ca2+ currents activated by depletion of Ca2+ stores in Xenopus oocytes were studied with a two-electrode voltage clamp. Buffering of cytosolic Ca2+ with EGTA and MeBAPTA abolished ICl(Ca) and unmasked a current in oocytes that was activated by InsP3 or ionomycin in minutes and by thapsigargin or the chelators themselves over hours. At −60 mV in 10 mM extracellular CaCl2, the current was typically around −90 or −160 nA in oocytes loaded with EGTA or MeBAPTA, respectively. This current was judged to be a Ca2+-selective current for the following reasons: (a) it was inwardly rectifying and reversed at membrane potentials usually more positive than +40 mV; (b) it was dependent on extracellular [CaCl2] with Km = 11.5 mM; (c) it was highly selective for Ca2+ against monovalent cations Na+ and K+, because replacing Na+ and K+ by N-methyl-d-glucammonium did not reduce the amplitude or voltage dependence of the current significantly; and (d) Ca2+, Sr2+, and Ba2+ currents had similar instantaneous conductances, but Sr2+ and Ba2+ currents appeared to inactivate more strongly than Ca2+. This Ca2+ current was blocked by metal ions with the following potency sequence: Mg2+ << Ni2+ ≈ Co2+ ≈ Mn2+ < Cd2+ << Zn2+ << La3+. It was also inhibited by niflumic acid, which is commonly used to block ICl(Ca). PMA partially inhibited the Ca2+ current, and this effect was mostly abolished by calphostin C, indicating that the Ca2+ current is sensitive to protein kinase C. These results are the first detailed electrophysiological characterization of depletion-activated Ca2+ current in nondialyzed cells. Because exogenous molecules and channels are easy to introduce into oocytes and the distortions in measuring ICl(Ca) can now be bypassed, oocytes are now a superior system in which to analyze the activation mechanisms of capacitative Ca2+ influx.

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