Gene expression profiling of proximal and distal renal tubules in Atlantic salmon (Salmo salar) acclimated to fresh water and seawater

Euryhaline teleost kidneys undergo a major functional switch from being filtratory in freshwater (FW) to being predominantly secretory in seawater (SW) conditions. The transition involves both vascular and tubular effects. There is consensus that the glomerular filtration rate is greatly reduced upon exposure to hyperosmotic conditions. Yet, regulation at the tubular level has only been examined sporadically in a few different species. This study aimed to obtain a broader understanding of transcriptional regulation in proximal versus distal tubular segments during osmotic transitions. Proximal and distal tubule cells were dissected separately by laser capture microdissection, RNA was extracted, and relative mRNA expression levels of >30 targets involved in solute and water transport were quantified by quantitative PCR in relation to segment type in fish acclimated to FW or SW. The gene categories were aquaporins, solute transporters, fxyd proteins, and tight junction proteins. aqp8bb1, aqp10b1, nhe3, sglt1, slc41a1, cnnm3, fxyd12a, cldn3b, cldn10b, cldn15a, and cldn12 were expressed at a higher level in proximal compared with distal tubules. aqp1aa, aqp1ab, nka-a1a, nka-a1b, nkcc1a, nkcc2, ncc, clc-k, slc26a6C, sglt2, fxyd2, cldn3a, and occln were expressed at a higher level in distal compared with proximal tubules. Expression of aqp1aa, aqp3a1, aqp10b1, ncc, nhe3, cftr, sglt1, slc41a1, fxyd12a, cldn3a, cldn3b, cldn3c, cldn10b, cldn10e, cldn28a, and cldn30c was higher in SW- than in FW-acclimated salmon, whereas the opposite was the case for aqp1ab, slc26a6C, and fxyd2. The data show distinct segmental distribution of transport genes and a significant regulation of tubular transcripts when kidney function is modulated during salinity transitions.

Download Full-text

The characterisation and uptake of paraquat in cultured baboon kidney proximal tubule cells (bPTC)

Human & Experimental Toxicology ◽

10.1191/096032701672136818 ◽

2001 ◽

Vol 20 (2) ◽

pp. 90-99 ◽

Cited By ~ 10

Author(s):

R Machaalani ◽

V Lazzaro ◽

G G Duggin

Keyword(s):

Proximal Tubule ◽

Mdck Cells ◽

Distal Tubule ◽

Genetic Mutation ◽

Immunoperoxidase Staining ◽

Renal Tubules ◽

Trypan Blue Exclusion ◽

Proximal Tubule Cells ◽

Morphological Studies ◽

Tubule Cells

A primary culture of baboon proximal tubule cells (bPTC) was prepared and characterised using LLC-PK1 cells of proximal tubule origin and MDCK cells of distal tubule origin, as positive and negative references, respectively. The proximal tubular origin of the bPTC was determined by morphological studies, immunoperoxidase staining and the expression of proximal tubule markers alkaline phosphatase and gammaglutamyltransferase. The hypothesis that paraquat (PQ) is transported by the bPTC was investigated. The cytotoxic threshold for PQ in these cells was determined and compared to the LLC-PK1 and MDCK cells. Furthermore, this study investigated the transport of the monovalent cation tetraethyl ammonium (TEA) and the polyvalent cation cimetidine in the bPTC and demonstrated their effect on the cellular uptake of PQ. The cytotoxic threshold of PQ in the bPTC, determined by cellular viability studies using the method of Trypan blue exclusion, is 0.05 mM at 2 h incubation. The LC50 after 24 his 76, 61 and 455 pM for the bPTC, LLC-PK1 and MDCK cells, respectively. This indicates that proximal tubule cells are more susceptible to PQ toxicity compared to distal tubule cells, which is consistent with clinical PQ toxicity where renal damage is found predominantly in the proximal renal tubules. The cations PQ and cimetidine were actively transported by the bPTC. The uptake of PQ (0.05 mM) commenced after 15 min whereas cimetidine (0.5 mM) uptake was evident after 2 min. Furthermore, cimetidine was shown to compete with PQ for uptake in the bPTC. Coincubating PQ (0.05 mM) and cimetidine (0.5 mM) for 60 min resulted in an approximate 50% decrease in PQ uptake. The cation TEA was not transported by the bPTC suggesting either a genetic mutation or complete absence of the transporter for TEA in the cells. The results suggest that PQ may be transported by the same cation transporter as cimetidine and not TEA, indicating PQ uptake in the bPTC to be via a polyvalent organic cation transporter.

Download Full-text

Whole-cell potassium currents in single early distal tubule cells

AJP Renal Physiology ◽

10.1152/ajprenal.1988.255.4.f699 ◽

1988 ◽

Vol 255 (4) ◽

pp. F699-F703 ◽

Cited By ~ 3

Author(s):

M. Hunter ◽

H. Oberleithner ◽

R. M. Henderson ◽

G. Giebisch

Keyword(s):

Intracellular Ph ◽

Potassium Conductance ◽

Distal Tubule ◽

Patch Clamp Technique ◽

Whole Cell ◽

Whole Cell Patch Clamp ◽

Cell Current ◽

Tubule Cells ◽

Distal Tubules ◽

Cellular Acidification

The apical membrane potassium conductance of amphibian early distal tubules is sensitive to changes in the intracellular pH, with cellular acidification causing a decreased conductance. With the whole-cell patch-clamp technique, we have measured the total potassium conductance of single isolated early distal tubule cells of the frog. With symmetrical potassium gluconate solutions, the whole-cell current was found to be strongly rectifying, with an inward conductance of 12.9 nS (at intracellular pH between 7.6 and 8.0) and an outward conductance of 1.0 nS. The inward current was almost totally inhibited by the addition of 10 mM Ba2+ to the bath solution. The use of pipette solutions with pH between 7.0 and 8.0 showed a positive correlation between intracellular pH and conductance. In contrast, acidification of the extracellular solution caused no significant change in conductance.

Download Full-text

Effect of uroguanylin on potassium and bicarbonate transport in rat renal tubules

Canadian Journal of Physiology and Pharmacology ◽

10.1139/y06-044 ◽

2006 ◽

Vol 84 (10) ◽

pp. 1003-1010 ◽

Cited By ~ 14

Author(s):

José Benedito Oliveira Amorim ◽

Raif Musa-Aziz ◽

Lucilia M.A. Lessa ◽

Gerhard Malnic ◽

Manassés Claudino Fonteles

Keyword(s):

Rat Kidney ◽

Distal Tubule ◽

Bicarbonate Transport ◽

Control Group ◽

Renal Tubules ◽

Stationary Concentration ◽

K Channels ◽

Series Of Experiments ◽

Distal Tubules

The effect of uroguanylin (UGN) on K+ and H+ secretion in the renal tubules of the rat kidney was studied using in vivo stationary microperfusion. For the study of K+ secretion, a tubule was punctured to inject a column of FDC-green-colored Ringer's solution with 0.5 mmol KCl/L ± 10−6 mol UGN/L, and oil was used to block fluid flow. K+ activity and transepithelial potential differences (PD) were measured with double microelectrodes (K+ ion-selective resin vs. reference) in the distal tubules of the same nephron. During perfusion, K+ activity rose exponentially, from 0.5 mmol/L to stationary concentration, allowing for the calculation of K+ secretion (JK). JK increased from 0.63 ± 0.06 nmol·cm–2·s–1 in the control group to 0.85 ± 0.06 in the UGN group (p < 0.01). PD was –51.0 ± 5.3 mV in the control group and –50.3 ± 4.98 mV in the UGN group. In the presence of 10−7 mol iberiotoxin/L, the UGN effect was abolished: JK was 0.37 ± 0.038 nmol·cm–2·s–1 in the absence of, and 0.38 ± 0.025 in the presence of, UGN, indicating its action on maxi-K channels. In another series of experiments, renal tubule acidification was studied, using a similar method: proximal and distal tubules were perfused with solutions containing 25 mmol NaHCO3/L. Acidification half-time was increased both in proximal and distal segments and, as a consequence, bicarbonate reabsorption decreased in the presence of UGN (in proximal tubules, from 2.40 ± 0.26 to 1.56 ± 0.21 nmol·cm–2·s–1). When the Na+/H+ exchanger was inhibited by 10−4 mol hexamethylene amiloride (HMA)/L, the control and UGN groups were not significantly different. In the late distal tubule, after HMA, UGN significantly reduced JHCO3–, indicating an effect of UGN on H+-ATPase. These data show that UGN stimulated JK+ by acting on maxi-K channels, and decreased JHCO3– by acting on NHE3 in proximal and H+-ATPase in distal tubules.

Download Full-text

Dietary HCO3 reduces distal tubule acidification by increasing cellular HCO3 secretion

AJP Renal Physiology ◽

10.1152/ajprenal.1996.271.1.f132 ◽

1996 ◽

Vol 271 (1) ◽

pp. F132-F142 ◽

Cited By ~ 5

Author(s):

D. E. Wesson

Keyword(s):

Carbonic Anhydrase ◽

Distal Tubule ◽

Carbonic Anhydrase Activity ◽

Anesthetized Rats ◽

Nephron Segment ◽

Tubule Cells ◽

Distal Tubules

We examined the components of net HCO3 reabsorption (H+/HCO3 secretion and transepithelial HCO3 permeability) in in vivo perfused distal tubules of anesthetized rats to determine the mechanisms by which dietary HCO3 reduces acidification in this nephron segment. Animals eating a minimum electrolyte diet drank either (in mM) 80 NaHCO3, 80 NaCl, or 40 Na2SO4 for 7-10 days and were compared with controls drinking distilled H2O. On perfusion with a HCO3- and Cl- -containing solution, net HCO3 reabsorption was lower than control in only the NaHCO3 animals (14.4 +/- 1.3 vs. 4.1 +/- 0.5 pmol.mm-1.min-1, P < 0.001). On perfusion with a 0 HCO3-0 Cl- solution, distal tubule luminal HCO3 accumulation (JHCO3) was higher in NaHCO3 animals than control (-13.7 +/- 1.3 vs. -4.7 +/- 0.7 pmol.mm-1.min-1, P < 0.002). Despite a higher JHCO3, estimated transepithelial HCO3 permeability in the NaHCO3 animals was similar to control [0.52 +/- 0.06 vs. 0.36 +/- 0.04 x 10(-7) cm2/s, P = not significant (NS)]. Luminal acetazolamide (Az) reduced JHCO3 in NaHCO3 animals to a level similar to control (-6.2 +/- 0.6 vs. -4.0 +/- 0.5 pmol.mm-1.min-1, P = NS) in this nephron segment containing cells with cytoplasmic but no luminal carbonic anhydrase activity. Including Cl- in the initial perfusate increased JHCO3 in NaHCO3 animals only (-20.8 +/- 1.9 vs. -13.7 +/- 1.3 pmol.mm-1.min-1, P < 0.02), and this increase was inhibited by luminal Az. Calculated H+ secretion was similar among groups. Together, the data indicate that dietary HCO3 reduces distal tubule acidification by increasing Az-sensitive generation of HCO3 by distal tubule cells that enters the lumen by a mechanism augmented by luminal Cl-.

Download Full-text

Adaptation of distal convoluted tubule of rats. II. Effects of chronic thiazide infusion

AJP Renal Physiology ◽

10.1152/ajprenal.1991.261.1.f137 ◽

1991 ◽

Vol 261 (1) ◽

pp. F137-F143 ◽

Cited By ~ 9

Author(s):

P. Morsing ◽

H. Velazquez ◽

F. S. Wright ◽

D. H. Ellison

Keyword(s):

Distal Tubule ◽

Chronic Administration ◽

Transport Capacity ◽

Binding Studies ◽

Nacl Transport ◽

Tubule Cells ◽

Distal Tubules ◽

Cortical Membranes ◽

Apical Membranes

Mammalian distal tubules adapt structurally and functionally when NaCl concentration in tubule fluid is altered chronically. These experiments were designed to test the hypothesis that chronic administration of hydrochlorothiazide (HCTZ), a drug that blocks Na and Cl uptake across apical membranes of rat distal tubule cells, would reduce intrinsic transport capacity of distal tubules and reduce the number of thiazide-sensitive transporters. Osmotic pumps were implanted into rats to deliver 3.75 mg/day HCTZ or vehicle for 10-14 days. All animals were offered a solution containing 0.8% NaCl and 0.1% KCl as drinking fluid. Free-flow micropuncture after 10–14 days indicated that Na and Cl delivery to distal tubule was not significantly different in HCTZ- and vehicle-treated animals. Microperfusion in vivo with an artificial interstitial solution, with no thiazide, indicated that 10-14 days of HCTZ infusion did reduce Na transport capacity of distal tubules from 390 +/- 32 to 203 +/- 24 pmol/min (P less than 0.01). In contrast, the number of thiazide-sensitive NaCl transporters, determined as high-affinity receptors for [3H]metolazone in renal cortical membranes, was higher in HCTZ group than in controls (2.2 +/- 0.4 vs. 1.0 +/- 0.1 pmol/mg protein, P less than 0.01). These data support the hypothesis that chronic blockade of NaCl entry across apical membranes of distal tubule cells reduces NaCl transport capacity, an effect that occurs despite an increase in the number of thiazide receptors. They indicate that thiazide receptor binding studies should be interpreted in combination with direct functional measurements.

Download Full-text

Cellular actions of thiazide diuretics in the distal tubule.

Journal of the American Society of Nephrology ◽

10.1681/asn.v15832 ◽

1990 ◽

Vol 1 (5) ◽

pp. 832-836

Author(s):

B A Stanton

Keyword(s):

Apical Membrane ◽

Basolateral Membrane ◽

Distal Tubule ◽

Thiazide Diuretics ◽

Exchange Inhibitor ◽

Disulphonic Acid ◽

Tubule Cells ◽

Distal Tubules ◽

Tubule Fluid

Thiazide diuretics inhibit electroneutral NaCl reabsorption across the distal tubule of the salamander, Amphiuma, and hyperpolarize the basolateral membrane voltage (Vbl) of distal tubule cells. The objective of this study was to determine whether thiazides hyperpolarize Vbl by reducing intracellular Cl- activity (Acl). To this end, distal tubules were perfused in vitro, and electrophysiological techniques were used to measure Acl and Vbl. Hydrochlorothiazide in tubular fluid reduced ACl from 17.0 to 12.6 mM and hyperpolarized Vbl by 16 mV. Reduction of Cl- in tubular fluid from 84 to 8 mM also decreased Acl from 16.1 to 9.9 mM and hyperpolarized Vbl by 12 mV. Because a previous study suggested that electroneutral NaCl reabsorption is mediated by Na+/H(+)-Cl-/HCO3- exchangers in the apical membrane, the Cl-/HCO3- exchange inhibitor, 4,4-diisothiocyanostilbene-2-2-disulphonic acid (DIDS) was added to tubule fluid. DIDS reduced Acl from 15.0 to 11.6 and hyperpolarized Vbl by 10 mV. DIDS and hydrochlorothiazide were not additive, inhibition of Na+/H+ exchange by amiloride (10(-3) M) and Na+ replacement with N-methyl-D-glucamine also reduced Acl from 17.4 to 12.9 and hyperpolarized Vbl by 16 mV. The hyperpolarization of Vbl in each experiment is referable to the fall in Acl. These data show that thiazide diuretics regulate ACl and that the hyperpolarization of Vbl is referable to the thiazide-induced reduction of Acl.

Download Full-text

PROBLEMS OF SPECIFICITY IN THE USE OF A STRONTIUM CAPTURE TECHNIQUE FOR THE CYTOCHEMICAL LOCALIZATION OF OUABAIN-SENSITIVE, POTASSIUM-DEPENDENT PHOSPHATASE IN MAMMALIAN RENAL TUBULES

Journal of Histochemistry & Cytochemistry ◽

10.1177/22.12.1163 ◽

1974 ◽

Vol 22 (12) ◽

pp. 1163-1168 ◽

Cited By ~ 29

Author(s):

J. A. FIRTH

Keyword(s):

Alkaline Phosphatase ◽

Proximal Tubule ◽

Adenosine Triphosphatase ◽

Plasma Membranes ◽

Distal Tubule ◽

Renal Tubules ◽

Cytochemical Localization ◽

Nitrophenyl Phosphate ◽

Basal Plasma ◽

Tubule Cells

The p-nitrophenyl phosphate-strontium procedure for the localization of the phosphatase component of Na-K-activated adenosine triphosphatase was evaluated using rat renal cortex as a test tissue. The results obtained by light microscopy were unexpected in that reaction product was found only on the brush borders of proximal tubule cells; this reaction was ouabain-resistant, K-independent and partially Mg-dependent, but could be completely inhibited by l-tetramisole. Electron microscopy showed that a reaction was also present on the cytoplasmic surfaces of the lateral and basal plasma membranes of the proximal and distal tubule cells. That seen in the distal tubule was sensitive to ouabain but not to l-tetramisole, whereas that in the proximal tubule showed a mixture of ouabain-sensitive and l-tetramisole-sensitive components. It is concluded that the procedure as originally described is not specific, demonstrating alkaline phosphatase as well as Na-K-adenosine triphosphatase, but that this problem may be overcome by the use of an alkaline phosphatase inhibitor.

Download Full-text