Browse
Submit Data
Databases
API
Help

Dataset Information

13 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Na+-H+ exchange in luminal-membrane vesicles from rabbit proximal convoluted and straight tubules in response to metabolic acidosis.

ABSTRACT: Na+-H+-exchanger activity of pars convoluta and pars recta luminal-membrane vesicles prepared from the proximal tubule of acidotic and control rabbits were assayed by a rapid-filtration technique and an Acridine Orange method. Both experimental approaches revealed the existence of an antiporter, sensitive to metabolic acidosis, in pars convoluta membrane vesicles. Kinetic data, obtained with the pH-sensitive dye, showed that the Km for Na+ transport was unchanged by acidosis, whereas Vmax. for exchanger activity was increased, on an average, by 44%. The fluorescence method, in contrast with the rapid-filtration technique, was able to detect exchanger activity in pars recta membrane vesicles. The Km value for the antiporter located in pars recta is comparable with that calculated for pars convoluta membrane vesicles. By contrast, the Vmax. of this exchanger is only about 25% of that found for pars convoluta. Furthermore, metabolic acidosis apparently does not increase Na+-H+-exchanger activity of pars recta luminal-membrane vesicles.

SUBMITTER: Jacobsen C

PROVIDER: S-EPMC1147295 | biostudies-other | 1986 Oct

REPOSITORIES: biostudies-other

ACCESS DATA

Json Xml

Similar Datasets

Response of the mitochondrial proteome of rat renal proximal convoluted tubules to chronic metabolic acidosis.

Project description:Metabolic acidosis is a common clinical condition that is caused by a decrease in blood pH and bicarbonate concentration. Increased extraction and mitochondrial catabolism of plasma glutamine within the renal proximal convoluted tubule generates ammonium and bicarbonate ions that facilitate the excretion of acid and partially restore acid-base balance. Previous studies identified only a few mitochondrial proteins, including two key enzymes of glutamine metabolism, which are increased during chronic acidosis. A workflow was developed to characterize the mitochondrial proteome of the proximal convoluted tubule. Based upon the increase in specific activity of cytochrome c oxidase, the isolated mitochondria were enriched eightfold. Two-dimensional liquid chromatography coupled with mass spectrometry was utilized to compare mitochondrial-enriched samples from control and chronic acidotic rats. Proteomic analysis identified 901 proteins in the control and acidotic samples. Further analysis identified 37 peptides that contain an N-?-acetyl-lysine; of these, 22 are novel sites. Spectral counting analysis revealed 33 proteins that are significantly altered in abundance in response to chronic metabolic acidosis. Western blot analysis was performed to validate the calculated changes in abundance. Thus the current study represents the first comprehensive analysis of the mitochondrial proteome of the rat renal proximal convoluted tubule and its response to metabolic acidosis.

| S-EPMC3543630 | biostudies-literature

Proteomic profiling and pathway analysis of the response of rat renal proximal convoluted tubules to metabolic acidosis.

Project description:Metabolic acidosis is a relatively common pathological condition that is defined as a decrease in blood pH and bicarbonate concentration. The renal proximal convoluted tubule responds to this condition by increasing the extraction of plasma glutamine and activating ammoniagenesis and gluconeogenesis. The combined processes increase the excretion of acid and produce bicarbonate ions that are added to the blood to partially restore acid-base homeostasis. Only a few cytosolic proteins, such as phosphoenolpyruvate carboxykinase, have been determined to play a role in the renal response to metabolic acidosis. Therefore, further analysis was performed to better characterize the response of the cytosolic proteome. Proximal convoluted tubule cells were isolated from rat kidney cortex at various times after onset of acidosis and fractionated to separate the soluble cytosolic proteins from the remainder of the cellular components. The cytosolic proteins were analyzed using two-dimensional liquid chromatography and tandem mass spectrometry (MS/MS). Spectral counting along with average MS/MS total ion current were used to quantify temporal changes in relative protein abundance. In all, 461 proteins were confidently identified, of which 24 exhibited statistically significant changes in abundance. To validate these techniques, several of the observed abundance changes were confirmed by Western blotting. Data from the cytosolic fractions were then combined with previous proteomic data, and pathway analyses were performed to identify the primary pathways that are activated or inhibited in the proximal convoluted tubule during the onset of metabolic acidosis.

| S-EPMC3761203 | biostudies-literature

Proteomic analysis of brush-border membrane vesicles isolated from purified proximal convoluted tubules.

Project description:The renal proximal convoluted tubule is the primary site of water, electrolyte and nutrient reabsorption and of active secretion of selected molecules. Proteins in the apical brush-border membrane facilitate these functions and initiate some of the cellular responses to altered renal physiology. The current study uses two-dimensional liquid chromatography/mass spectrometry to compare brush border membrane vesicles isolated from rat renal cortex (BBMV(CTX)) and from purified proximal convoluted tubules (BBMV(PCT)). Both proteomic data and Western blot analysis indicate that the BBMV(CTX) contain apical membrane proteins from cortical cells other than the proximal tubule. This heterogeneity was greatly reduced in the BBMV(PCT). Proteomic analysis identified 193 proteins common to both samples, 21 proteins unique to BBMV(CTX), and 57 proteins unique to BBMV(PCT). Spectral counts were used to quantify relative differences in protein abundance. This analysis identified 42 and 50 proteins that are significantly enriched (p values <or=0.001) in the BBMV(CTX) and BBMV(PCT), respectively. These data were validated by measurement of gamma-glutamyltranspeptidase activity and by Western blot analysis. The combined results establish that BBMV(PCT) are primarily derived from the proximal convoluted tubule (S1 and S2 segments), whereas BBMV(CTX) include proteins from the proximal straight tubule (S3 segment). Analysis of functional annotations indicated that BBMV(PCT) are enriched in mitochondrial proteins and enzymes involved in glucose and organic acid metabolism. Thus the current study reports a detailed proteomic analysis of the brush-border membrane of the rat renal proximal convoluted tubule and provides a database for future hypothesis-driven research.

| S-EPMC2886813 | biostudies-other

Bioprinting of 3D Convoluted Renal Proximal Tubules on Perfusable Chips.

Project description:Three-dimensional models of kidney tissue that recapitulate human responses are needed for drug screening, disease modeling, and, ultimately, kidney organ engineering. Here, we report a bioprinting method for creating 3D human renal proximal tubules in vitro that are fully embedded within an extracellular matrix and housed in perfusable tissue chips, allowing them to be maintained for greater than two months. Their convoluted tubular architecture is circumscribed by proximal tubule epithelial cells and actively perfused through the open lumen. These engineered 3D proximal tubules on chip exhibit significantly enhanced epithelial morphology and functional properties relative to the same cells grown on 2D controls with or without perfusion. Upon introducing the nephrotoxin, Cyclosporine A, the epithelial barrier is disrupted in a dose-dependent manner. Our bioprinting method provides a new route for programmably fabricating advanced human kidney tissue models on demand.

| S-EPMC5057112 | biostudies-literature

Potassium channels in the luminal membrane of rabbit proximal straight tubule. Evidence from vesicle studies.

Project description:The characteristics of 86Rb+ fluxes through K+ channels in luminal-membrane vesicles isolated from the pars recta of rabbit proximal tubule were studied. In KCl-loaded vesicles from the pars recta, transient accumulation of 86Rb+ is observed which is modestly inhibited by BaCl2 and blocked by CdCl2. The isotope accumulation is driven by an electrical diffusion potential, as shown in experiments using either these membrane vesicles loaded with different anions, or an outwardly directed Li+ gradient with a Li+ ionophore. The vesicles containing the channel show a cation selectivity with the order K+ greater than Rb+ greater than choline+ greater than or equal to Li+ greater than Na+. The CdCl2-sensitive 86Rb+ flux is dependent on intravesicular Ca2+. Increasing concentrations of Ca2+ gradually decreased the 86Rb+ uptake and at 1 microM-Ca2+ the CdCl2-sensitive isotope flux is nearly abolished.

| S-EPMC1133257 | biostudies-other

Proteomic profiling of the mitochondrial inner membrane of rat renal proximal convoluted tubules.

Project description:The proximal convoluted tubule is the primary site of renal fluid, electrolyte, and nutrient reabsorption, processes that consume large amounts of adenosine-5'-triphosphate. Previous proteomic studies have profiled the adaptions that occur in this segment of the nephron in response to the onset of metabolic acidosis. To extend this analysis, a proteomic workflow was developed to characterize the proteome of the mitochondrial inner membrane of the rat renal proximal convoluted tubule. Separation by LC coupled with analysis by MS/MS (LC-MS/MS) confidently identified 206 proteins in the combined samples. Further proteomic analysis identified 14 peptides that contain an N-?-acetyl-lysine, seven of which are novel sites. This study provides the first proteomic profile of the mitochondrial inner membrane proteome of this segment of the rat renal nephron. The MS data have been deposited in the ProteomeXchange with the identifier PXD000121.

| S-EPMC3826448 | biostudies-literature

Decreased NHE8 isoform expression and defective acidification in proximal convoluted tubules of senile rats.

Project description:We have previously found that aged rats show decreased proximal acidification without changes in NHE3 or V-H(+) ATPase expression in brush border membrane vesicles. However, we did not identify any mechanism underlying these observations. The aim of the present work was to evaluate some of the regulatory systems of proximal acidification that could be affected by aging. We measured plasma concentrations of parathyroid hormone (PTH) and the amount of cAMP in the renal cortex of young and old Wistar rats. PTH plasma concentration was increased in old rats, whereas, although it showed a tendency to increase, the cAMP content in the renal cortex of old rats was not significantly different compared with the cortex of young rats. We measured the abundance of NHE8 isoforms of the Na(+)/H(+) exchanger in brush border membrane vesicles from proximal convoluted tubules (PCT) of young and old rats by western blot analysis. We performed RT-PCR experiments in renal cortex homogenates from both experimental groups to evaluate mRNA expression of NHE3, NHE8 and H(+)ATPase. In senile rats, we detected a decreased abundance (at both gene expression and protein level) of the NHE8 isoform. These results could explain previous observations in which proximal tubule acidification appears affected in aged rats through a decrease in the activity of ethylisopropyl amiloride (EIPA)- and Bafilomycin-sensitive components, without changes in the NHE3 and V-H(+)ATPase abundance in the apical membrane of the PCT.

| S-EPMC2645991 | biostudies-other

Proteomic profiling of the effect of metabolic acidosis on the apical membrane of the proximal convoluted tubule.

Project description:The physiological response to the onset of metabolic acidosis requires pronounced changes in renal gene expression. Adaptations within the proximal convoluted tubule support the increased extraction of plasma glutamine and the increased synthesis and transport of glucose and of NH(4)(+) and HCO(3)(-) ions. Many of these adaptations involve proteins associated with the apical membrane. To quantify the temporal changes in these proteins, proteomic profiling was performed using brush-border membrane vesicles isolated from proximal convoluted tubules (BBMV(PCT)) that were purified from normal and acidotic rats. This preparation is essentially free of contaminating apical membranes from other renal cortical cells. The analysis identified 298 proteins, 26% of which contained one or more transmembrane domains. Spectral counts were used to assess changes in protein abundance. The onset of acidosis produced a twofold, but transient, increase in the Na(+)-dependent glucose transporter and a more gradual, but sustained, increase (3-fold) in the Na(+)-dependent lactate transporter. These changes were associated with the loss of glycolytic and gluconeogenic enzymes that are contained in the BBMV(PCT) isolated from normal rats. In addition, the levels of γ-glutamyltranspeptidase increased twofold, while transporters that participate in the uptake of neutral amino acids, including glutamine, were decreased. These changes could facilitate the deamidation of glutamine within the tubular lumen. Finally, pronounced increases were also observed in the levels of DAB2 (3-fold) and myosin 9 (7-fold), proteins that may participate in endocytosis of apical membrane proteins. Western blot analysis and accurate mass and time analyses were used to validate the spectral counting.

| S-EPMC3378174 | biostudies-literature

CD8<sup>+</sup> T cells stimulate Na-Cl co-transporter NCC in distal convoluted tubules leading to salt-sensitive hypertension.

Project description:Recent studies suggest a role for T lymphocytes in hypertension. However, whether T cells contribute to renal sodium retention and salt-sensitive hypertension is unknown. Here we demonstrate that T cells infiltrate into the kidney of salt-sensitive hypertensive animals. In particular, CD8+ T cells directly contact the distal convoluted tubule (DCT) in the kidneys of DOCA-salt mice and CD8+ T cell-injected mice, leading to up-regulation of the Na-Cl co-transporter NCC, p-NCC and the development of salt-sensitive hypertension. Co-culture with CD8+ T cells upregulates NCC in mouse DCT cells via ROS-induced activation of Src kinase, up-regulation of the K+ channel Kir4.1, and stimulation of the Cl- channel ClC-K. The last event increases chloride efflux, leading to compensatory chloride influx via NCC activation at the cost of increasing sodium retention. Collectively, these findings provide a mechanism for adaptive immunity involvement in the kidney defect in sodium handling and the pathogenesis of salt-sensitive hypertension.

| S-EPMC5227995 | biostudies-literature

Identification of renal distal convoluted tubules enriched genes

Project description:The renal distal convoluted tubule DCT is a short part of the distal nephron with specific functions transporting ions and thereby modifying Na, Ca, Mg, K balance A transcriptomic analysis of the DCT was performed to identify specific gene expression which would implicate those genes in specific DCT function

2013-11-19 | GSE51935 | GEO

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data